CN110545841A - 猪冠状病毒疫苗 - Google Patents

猪冠状病毒疫苗 Download PDF

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CN110545841A
CN110545841A CN201880008260.XA CN201880008260A CN110545841A CN 110545841 A CN110545841 A CN 110545841A CN 201880008260 A CN201880008260 A CN 201880008260A CN 110545841 A CN110545841 A CN 110545841A
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pdcov
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pedv
immunogenic composition
porcine
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S·阿克曼
J·R·赫尔曼
L·A·赫尔南德斯
L·A·霍布斯
A·V·耶尔
S·奥康纳
A·R·帕特森
E·M·沃恩
J·G·维多利亚
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Boehringer Ingelheim Animal Health USA Inc
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Abstract

本发明涉及用于保护猪不受与冠状病毒感染相关的疾病的疫苗，所述冠状病毒感染包括猪流行性腹泻病毒(PEDV)和/或猪丁型冠状病毒(PDCoV)。疫苗通常包括灭活/杀死的PEDV(例如，化学灭活的PED病毒)、和/或重组PEDV抗原、和/或佐剂灭活/杀死的PDCoV(例如，化学灭活的PDCoV病毒)、和/或重组PDCoV抗原和佐剂。还提供了用于保护猪不受与PEDV和/或PDCoV相关的疾病的方法，以及产生猪流行性腹泻病毒和/或猪丁型冠状病毒疫苗的方法。

Description

猪冠状病毒疫苗
序列表
本申请包含根据37 C.F.R.1.821–1.825的序列表。大小为480kb、名称为10-0176-WO-SEQ的序列表的ASCII文本文件的内容于2017年1月23日创建，并且经由EFS-Web以电子方式随同提交，该申请整体引入本文作为参考。
技术领域
本发明涉及免疫原性组合物，其保护猪免于由特异性的猪流行性腹泻病毒(PEDV)疫苗和猪丁型冠状病毒(PDCoV)引起的疾病，以及提供PEDV和PDCoV抗原两者的组合疫苗。由于小于10日龄仔猪中的高死亡率(高达100％)，经常一起存在的流行性肠道疾病是美国养猪业的主要经济关注点。
背景技术
猪流行性腹泻病毒是有包膜的正义单链RNA病毒，其引起猪中的急性腹泻、呕吐和脱水。它首先在欧洲得到鉴定，但在许多亚洲国家，包括朝鲜、中国、日本、菲律宾和泰国，已变得越来越成问题。在2013年4月，PEDV在美国中西部的猪中出现，迅速蔓延到全国各地。到2013年10月，PEDV在18个州的猪群中检测到。PEDV感染的经济影响已经很大。已鉴定了PEDV的北美分离株(Huang等人2013；Stevenson等人2013)，然而，在美国没有商购可得的完全批准的疫苗。相应地，一直需要开发能够保护猪不受与PEDV相关的疾病的疫苗。开发针对新出现的北美PEDV毒株有效的疫苗将是有利的，所述疫苗可以经由粘膜途径(经口或鼻内)以及经由肠胃外方法(例如，肌内、皮下或静脉内)施用。
PEDV是甲型冠状病毒属(Alphacoronavirus)的冠状病毒亚科(Coronavirinae)的成员(Bridgen等人1993)，并且在1971年首次在英国得到鉴定，并且后来在其它国家如比利时、中国、匈牙利、意大利、日本、朝鲜和泰国发现(Oldham J.1972；Pensaert和De BouckP.1978；Chen等人2008；Nagy等人1996；Martelli等人2008；Takahashi等人1983；Chae等人2000；和Puranaveja等人2009)。该家族的其它成员包括猪呼吸道冠状病毒(PRCV)、血凝性脑脊髓炎冠状病毒(PHE)和传染性胃肠炎病毒(TGEV)。尽管PEDV和TGEV病毒是相关的并且临床体征非常相似，但不存在免疫交叉保护。
PEDV是具有大约28kb正义单链RNA基因组的有包膜病毒，具有5'帽和3'多腺苷酸化尾。(Pensaert和De Bouck P.1978)。基因组包含5'非翻译区(UTR)、3'UTR和至少7个开放读码框(ORF)，其编码四种结构蛋白(刺突(S)、包膜(E)、膜(M)和核衣壳(N))和三种非结构蛋白(复制酶1a和1b以及ORF3)；这些以5’–复制酶(1a/1b)-S-ORF3-E-M-N-3’的次序排列在基因组上(Oldham J.1972；和Bridgen等人1993)。表征的前三种新出现的北美PEDV基因组序列：Minnesota MN(GenBank：KF468752.1)、Iowa IA1(GenBank：KF468753.1)和Iowa IA2(GenBank：KF468754.1)，具有28,038个核苷酸(nt)的相同大小，排除聚腺苷尾，并且与原型PEDV CV777毒株(GenBank：AF353511.1)共享基因组组构。这三种北美PEDV序列共享99.8％至99.9％的核苷酸同一性。特别地，毒株MN和IA2跨越整个基因组仅具有11个核苷酸差异。
PEDV S蛋白是由1,383个氨基酸(aa)组成的I型糖蛋白。基于其与其它冠状病毒的S蛋白的同源性，S蛋白可以分为S1(1-789aa)和S2(790-1,383aa)结构域(Chang等人；2002；Cruz等人，1994；Godet等人1994；Jackwood等人2001；Sturman和Holmes；1984；以及Sun等人2008)。冠状病毒中的S蛋白是表面抗原，其在调节与宿主细胞受体糖蛋白的相互作用以介导病毒进入，并且刺激天然宿主中的中和抗体的诱导中起作用。因此，S糖蛋白是用于开发针对PEDV的有效疫苗的主要靶。
PEDV M蛋白是最丰富的包膜组分，其在病毒组装过程中起重要作用，并且还诱导中和病毒的抗体。同样地，对于核衣壳提供结构基础、与病毒粒子RNA结合PEDV N蛋白，对于诱导细胞介导的免疫也可能是重要的(Saif，L.1993)。
PEDV中唯一的辅助基因是ORF3。虽然辅助基因一般在野毒株中得到维持，但ORF3的改变被认为影响毒力；细胞培养适应已用于改变ORF3基因，以便减少毒力(Song等人2003)。事实上，通过对ORF3基因的调查，研究人员已描绘了自2006年以来中国免疫猪群中的PEDV新基因群的出现。这些毒株的系统发育研究和PEDV在中国的地理重现已证实引起破坏性肠道疾病的那些野毒株在ORF3中与欧洲毒株和疫苗株在遗传上不同(Park等人2011)。
众所周知确实存在具有不同水平毒力的PEDV的不同毒株。在20世纪80年代和90年代期间，PEDV在整个欧洲在国家如比利时、英国、德国、法国、荷兰和瑞士流行。欧洲报告病例的频率随后逐渐减少和/或由PEDV引起的疾病对于开始疫苗的商业开发不具有足够的经济重要性(Song和Park 2012)。自20世纪80年代以来，中国已记录了PEDV的爆发，与大规模腹泻爆发相关的自2010年以来出现的PEDV变异株已变得更急性和严重。因此，疫苗开发试验主要在亚洲国家完成(Song和Park 2012)。自2010年以来出现的变异据报道在哺乳仔猪中具有80-90％的发病率和50-90％的死亡率(Bi等人2012；Pan等人2012；和Li等人1012)。最近的证据提示，中国新出现的PEDV毒力形式可能是活疫苗株进化的结果(Chen等人2010)。
作为影响猪肠的肠道疾病，PEDV经由粪-口暴露传播。受污染的卡车和设备是幼稚动物的常见感染源。PEDV感染的临床体征类似于传染性胃肠炎病毒(TGEV)感染(Pijpers等人1993)。在3周龄和更年幼的猪中，早在PEDV感染后24小时可以看到临床体征(包括急性水样、腹泻、呕吐和脱水)，导致100％的死亡率。受PEDV感染的饲养者和生长猪以及母猪和公猪可能发展腹泻和呕吐。动物也可以显示厌食症的体征，并且可能是嗜睡的。对老龄猪的全面影响尚未确定，但很可能饲料效率减少、上市天数增加、以及受感染动物对继发感染的易感性。对于母猪，身体状况的减少可能负面影响繁殖性能。报告已指示存在PEDV可以变成北美兽群中的地方性流行病的迹象，导致持续性腹泻和其它挑战。
感染PEDV的动物肠道中的总体和组织学变化在美国与在中国观察到的那些是相似的；基本上该病毒破坏猪肠的绒毛，使得无法吸收营养素。Huang等人2012报道死于明尼苏达州和爱荷华州爆发的疾病的动物具有局限于小肠的肉眼可见的病理性病变，并且小肠的特征在于由黄色流体扩张的半透明薄壁。组织学评估揭示具有绒毛钝化和融合的小肠区域以及固有层绒毛的最低限度成淋巴细胞浸润。
Huang等人2013表征了来自美国的输出爆发的三种不同的PEDV毒株-一种来自明尼苏达州，并且两种来自爱荷华州，分别命名为MN(GenBank登录号：KF468752)以及IA1(GenBank登录号：KF468753)和IA2(GenBank登录号：KF48754)。Huang的系统发育调查将PEDV毒株分组为两个不同的基因群，分别命名为基因群1(G1)和基因群2(G2)。刺突基因的N末端结构域(NTD)中的显著变化区分了基因群1和2。Huang等人2013提出N末端结构域(NTD)中的第二缺失区域(DR2)看来具有比DR1更高的抗原变化程度，提示新出现的北美毒株可能与G1a疫苗株“在抗原上”更不相关。
基因群1包括至少三个簇1a、1b和R。亚群1a包括早期欧洲、中国和朝鲜分离株，例如原型CV777毒株(比利时，1978，GenBank：AF353511.1)和毒株LZC(甘肃，中国，2006；GenBank：EF185992)和SM98(朝鲜，1998；GenBank：GU937797.1)。亚组1b含有5个毒株—一个来自韩国(DR13减毒疫苗株，GenBank：JQ023162.1)，并且其它来自中国，通过nsp3中常见的“遗传标记”8-aa缺失和在C末端处的大ORF3缺失联系。组“R”与其它基因群的重组体相关。然而，新近出现的PEDV毒株，包括自2010年以来在中国和自2013年以来在北美出现的那些，属于基因群G2a。中国毒株AH2012(GenBank登录号：KC210145)和北美毒株共享几个独特的核苷酸变化，并且在基因群2a中聚簇在一起。对AH2012的核苷酸同一性对于毒株MN和IA2为99.6％，并且对于毒株IA1为99.5％。研究人员已推测，AH2012样病毒可能传播到中国东部地区，然后运输至美国，并且很可能是最接近北美株的祖先。基因群2a的成员与基因群1a的原型PEDV毒株CV777仅共享大约96.9％的相似性(Bridgen等人1993；Huang等人2013；GenBank：AF353511.1)。像这样，基于历史CV777衍生的G1a毒株或DR13衍生的G1b毒株的减毒PEDV疫苗可以与新近出现的中国和北美G2a PEDV毒株在抗原上较不相关，并且因此可能是弱疫苗候选物。
密切相关的北美分离株US/Colorado/2013(GenBank登录号：KF272920.1)也已通过Marthaler等人，2013得到报道。如同上述北美分离株，CO/13的完全PEDV基因组与GenBank中可获得的其它完全PEDV基因组具有96.5％至99.5％的核苷酸同一性，与中国毒株AH2012(GenBank登录号KC210145)具有最高的核苷酸同一性(99.5％)。它是2a基因群的成员。通过CO/13的完全基因组与PEDV参考毒株CV777的完全基因组的比较，证实CO/13含有1-nt插入(在位置48处)和5'UTR中5个核苷酸的缺失(在位置73和83至86处)。这种北美病毒显示出在基因组位置20,696和21,125处的S1内增加的差异，与几种插入/缺失仅共享82％的核苷酸同一性。
已开发了在其基因组序列、递送模式和功效方面不同的几种PEDV疫苗。欧洲CV777毒株的细胞培养适应已在其中PEDV爆发严重的亚洲国家使用。这些自20世纪90年代以来一直在使用。
在20世纪80年代早期，日本研究人员从受感染猪的腹泻中分离出致病的PED病毒株83P-5。Kusanagi等人1989在Vero细胞中分离且适应了该毒株。自1997年以来，适应PEDV(P-5V)(83P-5)的细胞培养的减毒病毒疫苗已在日本的母猪中使用。第100次传代的83P-5毒株在日本被Nisseiken Co.，Ltd.(Sato等人2011)批准用作减毒PEDV疫苗。已报道83P-5毒株的适应和减弱显示S蛋白的细胞外部分中的突变，与减毒DR13毒株的那种具有序列相似性(Sato等人2011；参见在第100次传代时的毒株83P-5刺突基因序列，GenBank：AB548621.1)。尽管这种后来的日本疫苗被视为有效的，但并非所有母猪都能够对其仔猪传递免疫力(Usami等人1998)。日本毒株和欧洲毒株是基因群G1a或G1b的成员。如上文讨论的，与新近出现的中国品系的基因群2a相比，这些减毒的疫苗株与趋异的北美毒株更不相关。
用减毒的朝鲜PEDV毒株DR13(传代水平100)(GenBank：JQ023162.1)(基因群G1b的成员)的经口疫苗接种已显示作为疫苗是有效的。自2004年以来，该病毒株在韩国被批准并且用作经口疫苗，并且在2011年在菲律宾登记且商业化(Song和Park2012)。然而，据报道减毒的DR13不显著改变受攻击仔猪中病毒脱落的持续时间-这提示免疫保护是不完全的。此外，用高度减毒的PEDV经口免疫仅在非常高剂量的疫苗中赋予保护(Song和Park 2012)。
其它已知的疫苗包括 PT-100(ChoongAng Vaccine Laboratories，韩国)，组合杀死的PEDV和TGEV疫苗，以及 PED，杀死的PEDV疫苗。通过ChoonAngVaccine Laboratories提供的毒株和亚型是未知的。另外，另一家韩国公司KomipharmInternational Co.提供以商品名销售的一系列杀死的、活的和组合疫苗，其包括基因群G1a的PEDV毒株SM98P。中国的Qilu Animal Health Products Factory也在中国上市了含有PEDV和TGEV的组合杀死的疫苗，其毒株和亚型未知。
新近出现的猪丁型冠状病毒(porcine deltacorona virus)(PDCoV)，也称为猪丁型冠状病毒(swine deltacorona virus)SDCoV或SDCV，并且在本文中可互换使用，类似于猪流行性腹泻病毒(PEDV)，并且经常牵涉新生动物中的发病率，且促成该疾病对养猪业的全球传播和影响。PDCoV在PEDV之后的短时间内影响美国，具有类似的发病模式。在美国，与PEDV不同，丁型冠状病毒看来不引起仔猪中的死亡率，但引起仔猪中的急性发病率包括严重腹泻、呕吐和重量减轻。虽然比PEDV感染更温和，但PDCoV感染可以导致新生儿中30-40％的死亡率。在具有PEDV、轮状病毒和TGEV的共感染中可以看到更严重的疾病(Marthaler等人，2014a；和Wang等人2014)。另外，在2015年5月，在美国检测到第三种新试剂，猪正呼肠孤病毒(PORV)，当在共感染中存在时潜在地促成疾病严重性。虽然需要PDCoV和其它共感染试剂的发病机制和临床牵涉的更多了解，但PEDV和PDCoV的这种最近共出现已潜在地增强了疾病对畜群的严重性和影响。
猪丁型冠状病毒(PDCoV)是冠状病毒科(Coronaviridae)、丁型冠状病毒属(deltacoronavirus)的成员。该病毒在2009年收集的猪直肠拭子样品中首次在香港SAR得到鉴定(Woo，P.C.等人，Discovery of seven novel Mammalian and aviancoronaviruses in the genus deltacoronavirus supports bat coronaviruses as thegene source of alphacoronavirus and betacoronavirus and avian coronavirusesas the gene source of gammacoronavirus and deltacoronavirus.J Virol，2012.86(7)：第3995-4008页)，但不知道与任何疾病有关。SDCV，在本文中称为PDCoV，在2014年2月在俄亥俄州和印第安纳州首次在美国猪群中检测到，并且迅速传播到其它州(Hu，H.等人，Isolation and Characterization of Porcine Deltacoronavirus from Pigs withDiarrhea in the United States.J Clin Microbiol，2015.53(5)：第1537-1548页；Chen，Q.等人，Pathogenicity and pathogenesis of a United States porcinedeltacoronavirus cell culture isolate in 5-day-old neonatal piglets.Virology，2015.482：第51-59页；Li，G.等人，Full-Length Genome Sequence of PorcineDeltacoronavirus Strain USA/IA/2014/8734.Genome Announc，2014.2(2)；Marthaler，D.等人，Rapid detection，complete genome sequencing，and phylogenetic analysisof porcine deltacoronavirus.Emerg Infect Dis，2014a.20(8)：第1347-50页；Marthaler，D.等人，Complete Genome Sequence of Strain SDCV/USA/Illinois121/2014，a Porcine Deltacoronavirus from the United States.Genome Announc，2014b.2(2)；Wang，L.，B.Byrum和Y.Zhang，Detection and genetic characterization ofdeltacoronavirus in pigs，Ohio，USA，2014.Emerg Infect Dis，2014.20(7)：第1227-30页；Jung，K.等人，Pathogenicity of 2porcine deltacoronavirus strains ingnotobiotic pigs.Emerg Infect Dis，2015.21(4)：第650-4页)。PDCoV也在2014年4月在朝鲜(Lee，S.和C.Lee，Complete Genome Characterization of Korean PorcineDeltacoronavirus Strain KOR/KNU14-04/2014.Genome Announc，2014.2(6))、中国大陆(Dong，N.等人，Porcine Deltacoronavirus in Mainland China.Emerg Infect Dis，2015.21(12)：第2254-5页；和Song，D.等人，Newly Emerged Porcine DeltacoronavirusAssociated With Diarrhoea in Swine in China：Identification，Prevalence andFull-Length Genome Sequence Analysis.Transbound Emerg Dis，2015.62(6)：第575-80页)、以及日本(ISERPD 2015)检测到。
在鉴定后，三个研究小组已确认，用来自临床病例的细胞培养物衍生材料和/或组织匀浆物的感染导致已知菌和/或常规仔猪中的急性胃肠道临床体征(Chen等人，2015；Ma，Y.等人，Origin，evolution，and virulence of porcine deltacoronaviruses in theUnited States.MBio，2015.6(2)；和Vitosh-Sillman，S.等人Histopathological andimmunohistochemical characterization of pigs experimentally infected withporcine deltacoronavirus.in American Association of SwineVeterinarians.2015.Orlando，FL)。PDCoV已显示在临床体征发作后8-9天引起分娩室中的母猪和仔猪中的急性腹泻(Li等人2014)，连同呕吐和脱水(Ma等人2015)。该病毒具有与猪肠道冠状病毒(PEDV)、传染性胃肠炎冠状病毒(TGEV)和猪正呼肠孤病毒(PORV)相似的临床体征和症状。
通过Marx等人(WO201600757A2)的最近公开内容描述了据称保护猪免于由猪流行性腹泻病毒(PEDV)和PDCoV引起的疾病的免疫原性组合物。然而，很明显，当用减毒的欧洲INDEL毒株Calaf14攻击(在对照动物中仅引起23.8％的死亡率)时，基于北美科罗拉多株病毒的Marx等人的PEDV疫苗仅是部分保护性的(例如，当比较死亡率、重量减轻或腹泻的临床症状中的适度减少时)。此外，虽然Marx等人还描述了丁型冠状病毒疫苗，用单价PDCoV疫苗或二价PEDV和PDCoV疫苗未证实疾病的临床症状的保护或减少
因此，需要PEDV和PDCoV疫苗，其能够减少由PEDV和PDCoV引起的疾病的临床体征，并且在免疫动物中诱导保护性免疫，包括减少免疫动物中的病毒脱落。
发明内容
本发明提供了克服本领域缺陷的免疫原性组合物、疫苗和相关方法。本发明涉及免疫原性组合物，其包括冠状病毒科的有包膜(+)单链RNA病毒的灭活/杀死和/或重组形式：猪流行性腹泻病毒、或PEDV和/或猪丁型冠状病毒、或PDCoV。特别地，该申请提供了免疫原性组合物，其减少了猪中与PEDV和PDCoV感染相关的疾病的临床症状。本文PDCoV分离株(NSVL)(SEQ ID NO：1和2)以及分离株PDCoV 2.0307和PDCoV 5.0327(分别为SEQ ID NO：5、6、9和10)具有与其它北美和亚洲毒株的那些相似的遗传谱。本文PEDV分离株BI1251-125-10(在本文中称为“125-10”)(SEQ ID NO：29、32)是强毒的北美RNA病毒株，具有与基因群2a报道的其它北美PEDV的那些相似的遗传谱。PEDV衍生物分离株“125-10p30”(SEQ ID NO：34、36)是传代减毒病毒，也具有与北美基因群2a一致的遗传谱。
本发明的免疫原性组合物和疫苗包含灭活/杀死的PDCoV(例如，化学灭活的NSVL分离株(SEQ ID NO：1和2)，和/或分离株PDCoV 2.0307和PDCoV 5.0.327(分别为SEQ IDNO：5和6以及SEQ ID NO：9和10)，和/或灭活/杀死的PEDV(例如，化学灭活的PEDV分离株“125-10”或“125-10p30”(分别为SEQ ID NO：29或32和SEQ ID NO：33或36))，并且通常还包括佐剂。疫苗还可以包括其它组分，例如防腐剂、抗微生物剂、稳定剂例如可以增加疫苗的贮存期限的稳定剂、乳状液以及针对其它猪病原体的抗原。
本发明的免疫原性组合物和疫苗包含刺突抗原，其经由表达PDCoV和/或PEDV刺突蛋白，例如PDCoV刺突蛋白(例如，包含编码氨基酸序列SEQ ID NO：4、8、12、18或28的核酸序列SEQ ID NO：3、7、11、17或27)、和/或PEDV刺突核酸序列(包含编码氨基酸序列SEQ ID NO：31、35、47或53的SEQ ID NO：30、34、46或52)的重组杆状病毒在昆虫细胞的一个非限制性实例中表达，并且通常还包括佐剂。疫苗还可以包括其它组分，例如防腐剂、稳定剂以及针对其它猪病原体的抗原。
适用于本发明的优选PDCoV和/或PEDV刺突核酸序列是编码刺突多肽的多核苷酸，所述多核苷酸与PDCoV刺突多肽SEQ ID NO：3、7、11、17或其功能片段，和/或PEDV刺突SEQID NO：30、34、46或52或其功能片段至少具有至少80％、81％、82％、83％、84％、85％、86％、87％、88％或89％，更优选至少90％，91％，92％，93％或94％，且最优选至少95％、96％、96.1％、96.2％、96.3％、96.4％、96.5％、96.6％、96.7％、96.8％、96.9％、97％、97.1％、97.2％、97.3％、97.4％、97.41％、97.42％、97.43％、97.44％、97.45％、97.46％、97.47％、97.48％、97.49％、97.5％、97.51％、97.52％、97.53％、97.54％、97.55％、97.56％、97.57％、97.58％、97.59％、97.6％、97.61％、97.62％、97.63％、97.64％、97.65％、97.66％、97.67％、97.68％、97.69％97.7％、97.71％、97.72％、97.73％、97.74％、97.75％、97.76％、97.77％、97.78％、97.79％、97.8％、97.81％、97.82％、97.83％、97.84％、97.85％、97.86％、97.87％、97.88％、97.89％、97.9％、97.91％、97.92％、97.93％、97.94％、97.95％、97.96％、97.97％、97.98％、97.99％、98％、98.01％、98.02％、98.03％、98.04％、98.05％、98.06％、98.07％、98.08％、98.09％、98.1％、98.11％、98.12％、98.13％、98.14％、98.15％、98.16％、98.17％、98.18％、98.19％、98.2％、98.21％、98.22％、98.23％、98.24％、98.25％、98.26％、98.27％、98.28％、98.29％、98.3％、98.31％、98.32％、98.33％、98.34％、98.35％、98.36％、98.37％、98.38％、98.39％、98.4％、98.41％、98.42％、98.43％、98.44％、98.45％、98.46％、98.47％、98.48％、98.49％,98.5％、98.51％、98.52％、98.53％、98.54％、98.55％、98.56％、98.57％、98.58％、98.59％、98.6％、98.61％、98.62％、98.63％、98.64％、98.65％、98.66％、98.67％、98.68％、98.69％、98.7％、98.71％、98.72％、98.73％、98.74％、98.75％、98.76％、98.77％、98.78％、98.79％、98.8％、98.81％、98.82％、98.83％、98.84％、98.85％、98.86％、98.87％、98.88％、98.89％、98.9％、98.91％、98.92％、98.93％、98.94％、98.95％、98.96％、98.97％、98.98％、98.99％、99％、99.01％、99.02％、99.03％、99.04％、99.05％、99.06％、99.07％、99.08％、99.09％、99.1％、99.11％、99.12％、99.13％、99.14％、99.15％、99.16％、99.17％、99.18％、99.19％、99.2％、99.21％、99.22％、99.23％、99.24％、99.25％、99.26％、99.27％、99.28％、99.29％、99.3％、99.31％、99.32％、99.33％、99.34％、99.35％、99.36％、99.37％、99.38％、99.39％、99.4％、99.41％、99.42％、99.43％、99.44％、99.45％、99.46％、99.47％、99.48％、99.49％、99.5％、99.51％、99.52％、99.53％、99.54％、99.55％、99.56％、99.57％、99.58％、99.59％、99.6％,99.61％、99.62％、99.63％、99.64％、99.65％、99.66％、99.67％、99.68％、99.69％、99.7％、99.71％、99.72％、99.73％、99.74％、99.75％、99.76％、99.77％、99.78％、99.79％、99.8％、99.81％、99.82％、99.83％99.84％、99.85％、99.86％、99.87％、99.88％、99.89％、99.9％、99.91％、99.92％、99.93％、99.94％、99.95％、99.96％、99.97％、99.98％和99.99％的序列同一性。如本文使用的，特别理解术语“与SEQ ID NO：X的序列同一性”或“相同的SEQ ID NO：X”分别等价于术语“在SEQ ID NO：X的长度上与SEQ ID NO：X的序列的序列同一性”或“在SEQ ID NO：X的长度上与SEQ ID NO：X的序列相同”。
适用于本发明的优选刺突多肽是具有PDCoV刺突SEQ ID NO：4、8、12、18或28中所示序列的多肽和/或与SEQ ID NO：4、8、12、18、28、31、35、47和/或53具有至少80％的同源性的PEDV刺突SEQ ID NO：31、35，47或53，例如与SEQ ID NO：4、8、12、18、28、31、35、47和/或53具有至少85％的同源性，例如与SEQ ID NO：4、8、12、18、28、31、35、47和/或53具有至少85％的同源性，例如与SEQ ID NO：4、8、12、18、28、31、35、47和/或53具有至少90％的同源性，例如与SEQ ID NO：4、8、12、18、28、31、35、47和/或53具有至少95％、至少98％或至少99％的同源性。
术语“疫苗”和“免疫原性组合物”在本文中以广义进行定义，以指能够在用疫苗接种的动物中刺激免疫应答的可施用形式的任何类型的生物制剂。疫苗一般而言可以基于病毒本身(例如，杀死/灭活的或减毒的)或病毒的免疫原性(抗原性)组分。在本发明的一个实施方案中，疫苗(免疫原性组合物)优选包括以杀死/灭活形式的病毒剂或作为亚单位疫苗呈递的病毒的抗原部分。在本文中，当涉及免疫原性组合物使用时，术语“临床症状的减少”指与所讨论的疾病或状况相关的任何症状的减轻或改善(或部分或完全)。因此，通过本免疫原性组合物减少由PDCoV和/或PEDV感染的猪的临床症状一般导致病毒脱落和/或与由PDCoV和/或PEDV感染相关的一种或多种临床症状的减少(例如，小于10日龄的猪中的急性水样腹泻、急性呕吐、脱水、厌食、嗜睡、抑郁和高死亡率)。同样地，通过本免疫原性组合物减少由PDCoV和/或PEDV感染的猪的临床症状一般导致病毒脱落和/或与由PDCoV和/或PEDV感染相关的一种或多种临床症状的减少(例如，小于10日龄的猪中的急性水样腹泻、急性呕吐、脱水、厌食、嗜睡、抑郁和高死亡率)
本领域技术人员将理解，本文使用的组合物可以掺入已知的可注射、生理学上可接受的无菌溶液内。为了制备用于肠胃外注射或输注的即用型溶液，水性等渗溶液例如盐水或血浆蛋白质溶液可容易获得。另外，本发明的免疫原性和疫苗组合物可以包括兽医学上可接受的载体、稀释剂、等渗剂、稳定剂或佐剂。
本发明的方法包括但不限于在对象中引发针对单独或与彼此组合或其它共感染的PDCoV感染和/或PEDV感染的免疫应答的方法，其包括给对象施用免疫原性组合物的步骤，所述免疫原性组合物包含灭活/杀死的PDCoV和/或PEDV、减毒的PDCoV和/或PEDV、和/或衍生自PDCoV和/或PEDV的重组刺突抗原。优选地，引发针对超过一种血清型、或PDCoV或PEDV的毒株的免疫应答。本发明的组合物可以用于预防PDCoV感染、PEDV感染和/或PEDV/PDCoV共感染。优选地，此类免疫应答减少与一种或多种PDCoV基因型和/或PEDV基因型/基因组感染相关或由其引起的一种或多种临床体征的发生率或严重性。
在本文中，可以对其施用本发明的组合物的合适对象和有需要的对象包括需要预防性治疗病毒相关感染、疾病或状况的动物。其中通过使用本发明的组合物或方法刺激免疫应答的动物包括家畜例如猪、牛、家禽(例如鸡、鸭、鹅或火鸡)、山羊和绵羊，以及驯养动物例如小鼠、兔子、犬、猫和马。优选的动物包括猪科动物、鼠科动物、马科动物、兔类动物和牛科动物。最优选地，在猪中刺激免疫应答。
本发明还提供了减少与PDCoV感染和/或PEDV感染相关或由其引起的一种或多种临床体征的发生率或严重性的方法，所述方法包括施用本发明的免疫原性组合物的步骤，所述免疫原性组合物包含灭活/杀死的PDCoV疫苗、灭活/杀死的PEDV疫苗、和/或与衍生自如本文提供的PDCoV和/或PEDV的重组刺突抗原组合，且优选载体分子，使得PDCoV和/或PEDV感染的临床体重的发生率或严重性相对于未接受如本文提供的免疫原性组合物的对象减少至少10％，优选至少20％，甚至更优选至少30％，甚至更优选至少50％，甚至更优选至少70％，最优选100％。此类临床特征包括水样腹泻、呕吐和脱水。这些临床症状中的任一种都可以起因于由具有2a的基因群或者任何其它PEDV基因群包括G1a、G1b或G2b的PEDV的感染。同样地，这些临床体征中的任一种都可以起因于由不同基因型的PDCoV的感染和/或共感染。
在一个实施方案中，本免疫原性组合物包括化学灭活形式的PEDV。包含化学灭活的PDCoV(SEQ ID NO：1或2、5或6、9或10)病毒的疫苗是特别期望的。另外，本免疫原性组合物可以包括化学灭活形式的PEDV。包含化学灭活的PEDV(SEQ ID NO：29或32或SEQ ID NO：33或36)病毒的疫苗是特别期望的。本领域技术人员已知的各种化学灭活剂可以用于灭活病毒。乙烯亚胺和相关衍生物，例如二乙烯亚胺(“BEI”)和乙酰乙烯亚胺，是用于灭活SDCo和/或PED病毒的合适化学灭活剂的实例。其它化学灭活剂，例如β-丙内酯或醛(例如甲醛和戊二醛)也可以用于灭活病毒。
本免疫原性组合物和/或疫苗一般包括期望地可以具有生物粘附特性的佐剂，特别是在病毒被设计为能够鼻内施用的情况下。合适佐剂的实例包括交联的烯属不饱和羧酸聚合物，例如交联的丙烯酸聚合物。如本文使用的，术语“交联的丙烯酸聚合物”指由单体混合物形成的聚合物和共聚物，所述单体混合物包括丙烯酸作为混合物中的占优势单体。合适的交联丙烯酸聚合物的实例包括以商品名934P和971(可从B.F.Goodrich Co.，Cleveland，Ohio获得)商购可得的那些。用于本疫苗中的一种特别合适的佐剂是其布氏粘度不超过约20,000cPs的交联的丙烯酸聚合物，(如在20rpm下测量为以pH 7.5的1.0重量％水溶液)。当需要生物粘附性佐剂时，利用如在两片新鲜切除的兔胃组织之间测量的具有至少约50达因/cm2的生物粘附特性的佐剂可能是有利的(如通过美国专利号4,615,697中所述的程序测定的)。
本发明还涉及免疫对象的方法，其包括向对象施用如本文所述的任何免疫原性组合物。
术语“免疫”涉及通过向待免疫的对象施用免疫原性组合物的主动免疫，从而引起针对此类免疫原性组合物中包括的抗原的免疫学应答。
优选地，免疫导致畜群中单独或与PEDV感染组合的特定PDCoV感染的发生率降低，或导致由特定PDCoV和/或PEDV感染引起或与其相关的临床体征的严重性减少。
此外，用如本文提供的免疫原性组合物免疫有需要的对象，导致预防对象被PDCoV和/或PEDV感染的感染。甚至更优选地，免疫导致针对PDCoV和/或PEDV感染的有效、持久的免疫应答。应理解，所述时间段将持续超过2个月，优选超过3个月，更优选超过4个月，更优选超过5个月，更优选超过6个月。应理解，免疫接种可能并非在所有免疫的对象中是有效的。
优选地，在该上下文设想一群对象，其通常(即没有免疫接种)将发展通常由PDCoV感染和/或PEDV感染引起或与其相关的临床体征。本领域技术人员可以毫不费力地确定畜群的对象是否得到有效免疫。优选地，如果与未免疫或用在本发明之前可获得的免疫原性组合物免疫，但随后被特定的PDCoV和/或PEDV感染的对象相比，在至少20％、至少30％、至少40％、至少50％、至少60％、至少70％、至少80％、至少90％，再更优选至少95％且最优选100％的给定畜群的对象中的临床体征在发生率或严重性方面减少至少10％，更优选至少20％，再更优选至少30％，甚至更优选至少40％，再更优选至少50％，甚至更优选至少60％，再更优选至少70％，甚至更优选至少80％，再更优选至少90％，再更优选至少95％，且最优选100％，则免疫应该是有效的。
用于预防在有需要的对象中由PDCoV和/或PEDV引起的临床体征的方法，或保护猪不受与PDCoV和/或PEDV相关的疾病的方法，包括给猪施用含有灭活/杀死的PDCoV、和/或灭活/杀死的PEDV、和/或衍生自PDCoV和/或PEDV的刺突抗原的免疫原性组合物和/或疫苗。疫苗可以使用各种方法施用，包括鼻内、经口和/或肠胃外(例如，肌内)施用。在该方法的一个实施方案中，例如，含有灭活的PDCoV和/或灭活的PEDV的免疫原性组合物肌内施用一次或多次(例如，以2–4周的间隔)。在该方法的另一个实施方案中，例如，含有灭活的PDCoV和/或灭活的PEDV的疫苗经口施用一次或多次(例如，以2–4周的间隔)。在一个替代实施方案中，经口施用可以随后为肌内施用疫苗至少一次和/或在其之前(例如，在疫苗的肠胃外施用之后和/或之前2–4周)。理想地，给定畜群中的所有猪都按规定的间隔接种疫苗，以便保护防止疾病症状的传播。
还提供了产生灭活/杀死的PDCoV疫苗、灭活/杀死的PEDV疫苗、或组合PDCoV/PEDV免疫原性组合物的方法。对于PDCoV的生产，该方法通常包括用PDCo病毒，例如用PDCo病毒SEQ ID NO：1或2、5或6和9或10接种猪睾丸细胞。接种的猪睾丸细胞一般至少温育直到CPE为止，然后从温育的细胞中收获PED病毒。收获的含病毒的流体可以用化学灭活剂如二乙烯亚胺处理，以形成灭活/杀死的PDCo病毒。通常，灭活的病毒例如通过浓缩且与其它组分共混进一步加工，以产生商业制剂。例如，含有灭活病毒的流体可以浓缩，并且与佐剂和/或关于一种或多种其它猪病原体的抗原共混。为了产生PEDV，该方法通常包括用PED病毒，例如用PED病毒SEQ ID NO：29或SEQ ID NO：33接种猿猴细胞。使接种的猿猴细胞一般至少温育直至观察到CPE，然后从温育的细胞中收获PED病毒。收获的含病毒的流体可以用化学灭活剂如二乙烯亚胺处理，以形成灭活/杀死的PED病毒。通常，灭活的病毒例如通过浓缩且与其它组分共混进一步加工，以产生商业制剂。例如，含有灭活病毒的流体可以浓缩，并且与佐剂和/或关于一种或多种其它猪病原体的抗原共混。
还提供了产生经由表达PDCoV和/或PEDV刺突蛋白的重组杆状病毒在昆虫细胞中生成的重组表达的刺突抗原疫苗的方法。在一个示例性实施方案中的方法包括将PDCoV刺突编码序列(例如，SEQ ID NO：3、7、11、17或27)和/或PEDV刺突编码序列(SEQ ID NO：30、34、46或52)，包括例如PDCoV刺突-Fc抗原构建体(SEQ ID NO：17)、和/或PDCoV刺突Baculodisplay构建体(SEQ ID NO：27)、和/或PEDV 2a刺突构建体(SEQ ID NO：46)、和/或PEDV 2b刺突构建体(SEQ ID NO：52)，克隆到杆状病毒载体内，并且共转染Sf9昆虫细胞。
本申请还涉及试剂盒，其包括组合的(1)能够给猪施用疫苗的分配器；以及(2)化学灭活的PDCoV、和/或化学灭活的PEDV、和/或衍生自含有PDCoV和/或PEDV的疫苗的重组刺突抗原，其能够保护不受与PDCoV和/或PEDV相关的疾病。该试剂盒可以包括能够将其内容物分配为小滴例如气溶胶、雾化喷雾和/或液滴的分配器，以及例如当鼻内和/或肌内施用时，能够保护不受与PDCoV和/或PEDV相关的疾病的疫苗形式。
在本申请自始至终，该文本涉及本组合物和/或相关方法的各种实施方案。所描述的各种实施方案意欲提供各种说明性实施例，并且不应解释为对替代种类的描述。相反，应该注意，本文提供的各种实施方案的描述可以具有重叠的范围。本文讨论的实施方案仅是说明性的，并不意欲限制本发明的范围。
附图说明
下述附图构成本说明书的部分，并且被包括以进一步证实本发明的某些方面。通过参考与本文呈现的具体实施方案的详细描述组合的这些附图中的一个或多个，可以更好地理解本发明。
图1关于PDCoV分离株/毒株的完全基因组核苷酸邻近树。比例尺指示p距离。GenBank登录号连同两个字母的州或三个字母的国家代码一起列出(CHN–中国，TJK–泰国，LAO–老挝)。
图2关于PDCoV分离株/毒株的刺突氨基酸邻近树。比例尺指示p距离。GenBank登录号连同两个字母的州或三个字母的国家代码一起列出(CHN–中国，TJK–泰国，LAO–老挝)。
图3A PDCoV S1-IgG2a Fc的PDCoV S1-IgG2a Fc杆状病毒-蛋白质图的示意图。
图3B PDCoVS BD的PDCoVS BD Baculodisplay杆状病毒-蛋白质图的示意图。
图4与安慰剂(第3组)相比，用PDCoV原型(第1、2和4组，分别为BaculoS-BD-PDCoV、全细胞PDCoV和BaculoS-FC-PDCoV)的血清学研究。
图5与安慰剂(第3组)相比，如通过基于S1-IgG2a-Fc的ELISA检测的，在用PDCoV原型接种疫苗后的血清学应答：(第1、2和4组，分别为BaculoS-BD-PDCoV、全细胞PDCoV和BaculoS-FC-PDCoV)。
图6A PEDV 2a BD Baculodisplay杆状病毒蛋白质图的示意图。
图6B PEDV 2b BD Baculodisplay杆状病毒-蛋白质图的示意图。
图7在PEDV BD攻击后，通过组和天数的生存性。
图8接种疫苗的动物中的IgA水平。
具体实施方式
本发明提供了免疫原性组合物，其包括灭活/杀死形式的PDCoV和/或PEDV和/或重组表达的PDCoV和/或PEDV-刺突抗原。免疫原性组合物被设计用于保护猪不受与PDCoV和/或PEDV相关的疾病。免疫原性组合物通常包括化学灭活形式的PDCoV和/或PEDV，并且包括化学灭活/杀死的PDCoV和/或PEDV病毒的那些是特别期望的。在另一个实施方案中，免疫原性组合物包括例如经由表达PDCoV和/或PEDV刺突蛋白的重组杆状病毒，在昆虫细胞中生成的重组表达的PDCoV和/或PEDV刺突抗原。
一般地，本发明提供了包含一种或多种猪丁型冠状病毒(PDCoV)抗原和佐剂的免疫原性组合物，其中在猪丁型冠状病毒(PDCoV)中的是任何PDCoV：a.)其由SEQ ID NO：1、5或9编码，和/或包含SEQ ID NO：1、5或9的序列，和/或包含SEQ ID NO：1、5或9的RNA等价物；b.)其序列与SEQ ID NO：1、5或9至少99％相同，和/或与SEQ ID NO：1、5或9的RNA等价物至少99％相同，c.)其刺突蛋白由SEQ ID NO：3、7或11的核酸序列编码；d.)其刺突蛋白由与SEQ ID NO：3、7或11至少90％相同的核酸序列编码；e.)其由SEQ ID NO：2、6或10编码；或f.)其序列与SEQ ID NO：2、6或10至少99％相同。
在一个具体方面，根据上述句子的免疫原性组合物，其中佐剂是水包油型乳状液。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述免疫原性组合物是重组抗原或灭活的完整猪丁型冠状病毒PDCoV抗原。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述抗原是灭活的完整PDCoV抗原。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述PDCoV抗原是化学灭活的。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述PDCoV抗原通过用化学灭活剂处理而化学灭活，所述化学灭活剂包括选自乙烯亚胺、二乙烯亚胺、乙酰乙烯亚胺及其混合物的化合物。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述PDCoV通过用二乙烯亚胺处理而化学灭活。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述佐剂是基于水包油型乳状液的佐剂。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中灭活的猪丁型冠状病毒(PDCoV)包含SEQ ID NO：1、5或9，和/或包含SEQ ID NO：1、5或9的RNA等价物。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述PDCoV抗原是重组抗原。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：a.)编码猪丁型冠状病毒(PDCoV)刺突蛋白的抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：4、8、12、18或28具有至少90％的同源性；b.)包含a)的分离核酸的重组载体；c.)由a)的核酸编码的重组PDCoV刺突蛋白；和d.)其任何组合。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中此类免疫原性组合物包含药学上可接受的载体和/或赋形剂。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述水包油型乳状液是基于水包油型乳状液的佐剂。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述免疫原性组合物进一步包含一种或多种另外的抗原。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中另外的抗原是猪流行性腹泻病毒(PEDV)的抗原，其中所述猪流行性腹泻病毒(PEDV)是任何PEDV：a.)其由SEQ ID NO：29或33编码，和/或包含SEQ ID NO：29或33的序列，和/或包含SEQ ID NO：29或33的RNA等价物；b.)其序列与SEQ ID NO：29或33至少99％相同，和/或与SEQ ID NO：29或33的RNA等价物至少99％相同；c.)其刺突蛋白由SEQ ID NO：30、34、46或52的核酸序列编码；d.)其刺突蛋白由与SEQ ID NO：30、34、46或52至少90％相同的核酸序列编码；e.)其由SEQ ID NO：32或36编码；或f.)其由与SEQ ID NO：32或36至少99％相同的序列编码。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述另外的PEDV抗原是重组抗原或灭活的完整猪流行性腹泻病毒(PEDV)。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述抗原是灭活的完整猪流行性腹泻病毒(PEDV)。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述抗原是重组PEDV抗原。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：a.)编码猪流行性腹泻病毒(PEDV)刺突蛋白的抗原的分离核酸，其中所述抗原与SEQ ID NO：31、35、47或53具有至少90％的同源性；b.)包含a)的分离核酸的重组载体；c.)由a)的核酸编码的重组PEDV刺突蛋白；和d.)其任何组合。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：猪流行性腹泻病毒(PEDV)的结构蛋白M、E或N。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述免疫原性组分是所述分离的核酸。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述免疫原性组分是所述重组载体。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述免疫原性组分是所述重组猪流行性腹泻病毒(PEDV)刺突蛋白。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其中所述免疫原性组分是组合。
本发明进一步涉及用于减少与猪丁型冠状病毒(PDCoV)相关的疾病的临床症状的方法，其包括给猪施用如上述句子中任一中所述的免疫原性组合物。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其包括给猪施用包含灭活的完整PDCoV抗原的免疫原性组合物。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其包括给猪施用包含PDCoV抗原作为重组抗原的免疫原性组合物。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中此类施用的免疫原性组合物包含选自以下的一种或多种免疫原性组分：a.)其由SEQ ID NO：1编码，和/或包含SEQ ID NO：1、5或9的序列，和/或包含SEQ ID NO：1、5或9的RNA等价物；b.)其序列与SEQ ID NO：1、5或9至少99％相同，和/或与SEQ ID NO：1、5或9的RNA等价物至少99％相同；c.)其刺突蛋白由SEQ ID NO：3、7、11、17、27的核酸序列编码；d.)其刺突蛋白由与SEQID NO：3、7、11、17或27至少90％相同的核酸序列编码；e.)其由SEQ ID NO：2、6或10编码；和f.)其序列与SEQ ID NO：2、6或10至少99％相同。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中此类施用的免疫原性组合物包含选自以下的一种或多种免疫原性组分：a.)其由SEQ ID NO：1、5或9的序列编码，或者包含SEQ ID NO：1、5或9的序列；b.)其序列与SEQ ID NO：1、5或9至少99％相同；d.)其刺突蛋白由SEQ ID NO：3、7、11、17或27的核酸序列编码；和e.)其刺突蛋白由与SEQ ID NO：3、7、11、17或27至少90％相同的核酸序列编码。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中此类施用的免疫原性组合物包含重组抗原，所述重组抗原包含选自以下的一种或多种免疫原性组分：a.)编码猪流行性腹泻病毒、猪丁型冠状病毒(PDCoV)刺突蛋白抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：4、8、12、18或28具有至少90％的同源性；b.)包含a)的分离核酸的重组载体；c.)由a)的核酸编码的重组猪丁型冠状病毒(PDCoV)刺突蛋白；和d.)其任何组合。
本发明进一步涉及用于减少与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的临床症状的方法，其包括给猪施用如上述句子中任一中所述的任何免疫原性组合物，优选地，其中所述免疫原性组合物包含一种或多种PDCoV抗原和一种或多种另外的抗原，其中所述另外的抗原是猪流行性腹泻病毒(PEDV)的抗原。
在本发明的一个进一步的具体方面，该方法如上述句子中所述，其包括给此类猪施用免疫原性组合物，其中所述另外的PEDV抗原是重组抗原或灭活的完整猪流行性腹泻病毒(PEDV)。
在本发明的一个进一步的具体方面，该方法如上述句子中所述，其包括给此类猪施用如上述句子中任一中所述的免疫原性组合物。
本发明进一步涉及用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)相关的疾病的免疫原性应答的试剂盒，其包含：a.)能够给猪施用免疫原性组合物的分配器；和b.)如上述句子中任一中所述的免疫原性组合物。
本发明进一步涉及用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)相关的疾病的免疫原性应答的试剂盒，其包含：a.)能够给猪施用免疫原性的分配器；和b.)如上述句子中任一中所述的免疫原性组合物，更具体而言，其中所述抗原是灭活的完整PDCoV抗原，甚至更具体而言，其中所述PDCoV抗原是化学灭活的；更具体而言，其中所述PDCoV抗原通过用化学灭活剂处理而化学灭活，所述化学灭活剂包括选自乙烯亚胺、二乙烯亚胺、乙酰乙烯亚胺及其混合物的化合物；优选地，其中所述PDCoV通过用二乙烯亚胺处理而化学灭活。更具体而言，其中所述佐剂是基于水包油型乳状液的佐剂；且甚至更优选地，其中所述灭活的猪丁型冠状病毒(PDCoV)包含SEQ ID NO：1、5或9，和/或包含SEQ ID NO：1、5或9的RNA等价物。
本发明进一步涉及用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)相关的疾病的免疫原性应答的试剂盒，其包含：a.)能够给猪施用免疫原性组合物的分配器；和b.)如上述句子中任一中所述的免疫原性组合物，更具体而言，其中所述PDCoV抗原是重组抗原，甚至更具体而言，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：a.)编码猪丁型冠状病毒(PDCoV)刺突蛋白抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：4、8、12、18或28具有至少90％的同源性；b.)包含a)的分离核酸的重组载体；c.)由a)的核酸编码的重组PDCoV刺突蛋白；和d)其任何组合，甚至更具体而言，其中此类免疫原性组合物包含药学上可接受的载体和/或赋形剂；并且甚至更具体而言，其中所述水包油型乳状液是基于水包油型乳状液的佐剂。
本发明进一步涉及用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的免疫原性应答的试剂盒，其包含：a.)能够给猪施用免疫原性组合物的分配器；和b.)如上述句子中任一中所述的免疫原性组合物，具体地其中所述另外的抗原是猪流行性腹泻病毒(PEDV)，并且是如上述句子中任一中所述的任何PEDV抗原。
本发明进一步涉及用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的免疫原性应答的试剂盒，其包含：a.)能够给猪施用免疫原性组合物的分配器；和b.)如上述句子中任一中所述的免疫原性组合物，具体地其中所述另外的抗原是猪流行性腹泻病毒(PEDV)，是如上述句子中任一中所述的任何重组PEDV抗原或任何灭活的完整猪流行性腹泻病毒(PEDV)。
本发明进一步涉及用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的免疫原性应答的试剂盒，其包含：a.)能够给猪施用免疫原性组合物的分配器；和b.)如上述句子中任一中所述的免疫原性组合物，具体地其中所述另外的抗原是如上述句子中任一中所述的任何重组PEDV抗原。
本发明进一步涉及制备如上述句子中任一中所述的猪丁型冠状病毒(PDCoV)免疫原性组合物的方法，其进一步包括：a.)用猪丁型冠状病毒(PDCoV)接种猪睾丸细胞；b.)温育接种的猪睾丸细胞；c.)从温育的细胞中收获猪丁型冠状病毒(PDCoV)；以及d.)用化学灭活剂，优选用选自乙烯亚胺、二乙烯亚胺、乙酰乙烯亚胺或其混合物的化合物处理收获的细胞，以形成灭活的猪丁型冠状病毒(PDCoV)抗原。
在本发明的一个进一步的具体方面，该方法如上述句子中所述，其中所述猪丁型冠状病毒(PDCoV)包含另外的特征：a.)由SEQ ID NO：1、5或9的序列编码，或者包含SEQ IDNO：1、5或9的序列，b.)与SEQ ID NO：1、5或9至少99％相同的序列，c.)由SEQ ID NO：3、7、11、17或27的核酸序列编码的刺突蛋白；d.)由与SEQ ID NO：3、7、11、17或27至少90％相同的核酸序列编码的刺突蛋白。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中所述猪丁型冠状病毒(PDCoV)包含另外的特征SEQ ID NO：1、5或9，和/或SEQ ID NO：1、5或9的RNA等价物。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中所述猪睾丸细胞是AI-ST细胞。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中所述化学灭活剂包括二乙烯亚胺。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其进一步包括给猪丁型冠状病毒(PDCoV)抗原添加基于水包油型乳状液的佐剂的另外特征。
本发明进一步涉及用于制备免疫原性组合物的方法，所述免疫原性组合物包含如前述句子中任一中所述的重组猪丁型冠状病毒(PDCoV)抗原，所述方法包括：a.)在宿主细胞中表达猪丁型冠状病毒(PDCoV)的抗原，b.)收获猪丁型冠状病毒(PDCoV)细胞的抗原；以及c)将基于水包油型乳状液的佐剂加入步骤b)的猪丁型冠状病毒(PDCoV)抗原中。
在本发明的一个进一步的具体方面，该方法如上述句子中所述，其中所述猪丁型冠状病毒(PDCoV)抗原包含：a.)编码猪丁型冠状病毒(PDCoV)刺突蛋白的抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：3、7、11、17或27具有至少90％的同源性；b.)包含a)的分离核酸的重组载体；c.)由a)的核酸编码的重组猪丁型冠状病毒(PDCoV)刺突蛋白；和d.)其任何组合。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中所述猪丁型冠状病毒(PDCoV)的抗原由重组杆状病毒载体表达。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中所述猪丁型冠状病毒(PDCoV)的抗原在昆虫细胞中表达。
在本发明的一个进一步的具体方面，该方法如上述句子中任一中所述，其中作为另外的特征，基于水包油的佐剂是基于水包油型乳状液的佐剂。
本发明进一步涉及包含如上述句子中任一中所述的PDCoV的免疫原性组合物，其用于减少与猪丁型冠状病毒(PDCoV)相关的疾病的症状。
本发明进一步涉及包含PDCoV和至少一种另外的抗原的免疫原性组合物，其中至少一种另外的抗原是猪流行性腹泻病毒(PEDV)的抗原，其用于减少与猪丁型冠状病毒(PDCoV)和/或猪流行病毒(PEDV)相关的疾病的症状。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其进一步包含PEDV基因型2a的一种或多种抗原，具体地PEDV具有北美起源。
在本发明的一个进一步的具体方面，免疫原性组合物如上述句子中任一中所述，其进一步包含一种或多种PDCoV抗原，其中所述PDCoV具有北美起源。
更一般地说，本领域技术人员已知的各种化学灭活剂可以用于灭活病毒。乙烯亚胺和相关衍生物，例如二乙烯亚胺(BEI)和乙酰乙烯亚胺，是用于灭活PED病毒的合适化学灭活剂的实例。其它化学灭活剂，例如β-丙内酯、醛(例如甲醛)和/或去污剂(例如，去污剂、 X或烷基三甲基铵盐)也可以用于灭活病毒。可以使用本领域技术人员已知的标准方法进行灭活。可以以定期时间间隔获取样品并且测定残留的活病毒。用适当的特异性单克隆或多克隆抗体监测对适当细胞系的致细胞病变作用和/或荧光染色可以用于检测残留活病毒的存在。
用BEI的灭活可以通过将储备BEI溶液(例如，通过将0.1–0.2M 2-溴-乙胺氢溴酸盐加入0.1–0.2N水性NaOH中形成的溶液)与病毒流体组合至约1–5mM BEI的最终浓度来实现。通常通过将BEI-病毒混合物保持在35–40℃(例如，37℃)下，伴随24–72小时的持续混合来进行灭活。通过加入硫代硫酸钠溶液至超过BEI浓度的最终浓度(例如，加入以BEI体积17％的硫代硫酸钠以中和过量的BEI)，随后为混合，可以终止病毒灭活。
本免疫原性组合物通常包括佐剂，以及需要时的一种或多种乳化剂，例如去污剂，其掺入到灭活/杀死的PEDV。合适的佐剂包括例如乙酸维生素E增溶物、氢氧化铝、磷酸铝或氧化铝、(矿物)油乳状液、非离子型去污剂、角鲨烯和皂苷。可以使用的其它佐剂包括油基佐剂，例如弗氏完全佐剂(FCA)和弗氏不完全佐剂(FIA)。已发现交联的烯属不饱和羧酸聚合物，例如971聚合物，是用于本灭活的PEDV免疫原性组合物中的特别适合的佐剂。
关于合适的水包油型乳状液的实例是基于的佐剂，例如(水包油型乳状液)、(具有二甲基二十八烷基溴化铵(DDA)的水包油型)、(具有专利权免疫刺激剂的水包油型)、(具有交联聚合物的由水包油型组成的双重佐剂)、以及(不含动物起源组分的水包油型乳状液。(MVP Technologies，Inc.Omaha，Nebr.，USA)。包含灭活的PEDV或重组PEDV蛋白的药物/疫苗组合物已用水包油型乳状液有效地佐剂化，优选用此类基于的佐剂，更优选用(不含动物起源组分的水包油型乳状液)和(不含动物起源组分的水包油型乳状液)。
一般有利的是以剂量单位形式配制本组合物以便于施用和剂量的均匀性。如本文使用的，剂量单位形式指适合作为待治疗的哺乳动物对象的单位剂量的物理上离散的单位；每个单位含有预定数量的活性材料，其计算为与所需的药物载体结合产生所需的疗效。关于灭活/杀死的PDCoV和/或PEDV、和/或重组表达的PDCoV和/或PEDV抗原的剂量单位形式的规格由其它因素决定且取决于其它因素：(a)活性材料的独特特征和待达到的特定疗效；(b)复合此类活性材料用于治疗疾病的领域中固有的局限性；以及(c)剂量单位形式的预期施用方式。
通常配制主要活性成分，用于以有效量与如本文公开的以剂量单位形式的合适的药学上可接受的载体方便有效的施用。单位剂型可以例如含有PDCoV和/或PEDV抗原，其量范围为1至约5个相对效力单位(“RPU”)。该抗原量一般以约1至约25/ml的载体存在。在含有补充活性成分的组合物的情况下，剂量通过参考补充活性成分的通常剂量和施用方式来确定。
本发明的疫苗通常包括用药学上可接受的载体配制的灭活的PDCoV和/或PEDV。适合于注射使用的药物形式通常包括无菌水溶液(在水溶性的情况下)或分散体，以及用于临时制备无菌可注射溶液或分散体的无菌粉末。该制剂应期望地是无菌的且流动至存在容易可注射性的程度。剂型在制造和贮存条件下应该是稳定的，并且通常针对微生物如细菌和真菌的污染作用进行防腐。载体可以是溶剂或分散介质，其含有例如水、乙醇、多元醇(例如甘油、丙二醇、液体聚乙二醇等等)、其合适的混合物和植物油。一种可能的载体是生理盐溶液。例如，通过使用涂层如卵磷脂、在分散体的情况下通过维持所需的粒度和通过使用表面活性剂，可以维持溶液的适当流动性。可以通过各种抗菌剂和抗真菌剂，例如对羟基苯甲酸酯、三氯叔丁醇、苯酚、山梨酸、硫柳汞(乙基汞-硫代水杨酸钠)、新霉素(deomycin)、庆大霉素等等，来达到微生物作用的预防。在许多情况下，可能优选包括等渗剂，例如糖或氯化钠。需要时，可以通过在组合物中使用延迟吸收的试剂，例如单硬脂酸铝和明胶，来达到可注射组合物的延长吸收。
可以通过将灭活的病毒以所需的量，根据需要与如上文列举的各种其它成分一起掺入适当的溶剂中，随后为过滤灭菌来制备无菌可注射溶液。一般地，可以通过将各种活性成分掺入无菌媒介物内来制备分散体，所述无菌媒介物含有基础分散介质和来自上文列举的那些的所需其它成分。在用于制备无菌可注射溶液的无菌粉末的情况下，优选的制备方法是真空干燥和冷冻干燥技术，所述技术从其先前无菌过滤的溶液产生活性成分加上任何其它所需成分的粉末。
向本组合物中加入稳定剂以改善灭活病毒的稳定性也可能是有利的。合适的稳定剂包括例如甘油/EDTA、碳水化合物(如山梨糖醇、甘露醇、海藻糖、淀粉、蔗糖、葡聚糖或葡萄糖)、蛋白质(如白蛋白或酪蛋白)和蛋白质降解产物(如部分水解的明胶)。需要时，可以通过本领域已知的方法来缓冲制剂，使用试剂例如碱金属磷酸盐，例如磷酸氢二钠、磷酸二氢钠、磷酸氢钾和/或磷酸二氢钾。其它溶剂例如乙醇或丙二醇，可以用于增加成分在疫苗制剂中的溶解度和/或溶液的稳定性。可以用于本制剂中的进一步添加剂包括常规抗氧化剂和常规螯合剂，如乙二胺四乙酸(EDTA)。
本发明的组合物和方法可以通过下述实施例进行说明，所述实施例呈现以说明本发明，并且帮助教导普通技术人员如何制备且使用其。这些实施例决不预期缩小或以其它方式限制本发明的范围。
除非另有说明，否则本发明的实践将采用分子生物学、微生物学、重组DNA技术、蛋白质化学和免疫学的常规技术，所述技术在本领域的技术内。此类技术在文献中充分解释。参见例如Sambrook，Fritsch&Maniatis，Molecular Cloning：A Laboratory Manual，第I、II和III卷，第二版(1989)；DNA Cloning，第I和II卷(D.N.Glover编辑，1985)；Oligonucleotide Synthesis(M.J.Gait编辑1984)；Nucleic Acid Hybridization(B.D.Hames&S.J.Higgins编辑1984)；Animal Cell Culture(R.K.Freshney编辑1986)；Immobilized Cells and Enzymes(IRL press，1986)；Perbal，B.，A Practical Guide toMolecular Cloning(1984)；该系列，Methods In Enzymology(S.Colowick和N.Kaplan编辑，Academic Press，Inc.)；Protein purification methods–a practical approach(E.L.V.Harris和S.Angal，编辑，IRL Press at Oxford University Press)；和Handbookof Experimental Immunology，第I-IV卷(D.M.Weir和C.C.Blackwell编辑，1986，Blackwell Scientific Publications)。
应理解，本发明并不限于特定的DNA、RNA、多肽序列或工艺参数，因为这些当然可以改变。还应理解，本文使用的术语仅用于描述本发明的特定实施方案的目的，并不预期是限制性的。必须注意，如在本说明书和所附权利要求中使用的，单数形式“一个”、“一种”和“该/所述”包括复数指示物，除非内容另有明确说明。因此，例如，提及“抗原”包括两种或更多种抗原的混合物；提及“赋形剂”包括两种或更多种赋形剂的混合物等等。
定义：
除非另有定义，否则本文使用的所有技术和科学术语具有与本申请在提交时所属领域的技术人员通常理解相同的含义。术语的含义和范围应该是明确的；然而，在任何潜在歧义的情况下，本文提供的定义优先于任何字典或外部定义。此外，除非上下文另有要求，否则单数术语应该包括复数，并且复数术语应该包括单数。在本文中，除非另有说明，否则“或”的使用意指“和/或”。此外，术语“包括(including)”以及其它形式如“包括(includes)”和“包括(included)”的使用并非限制性的。本文提及的所有专利和出版物都引入本文作为参考。
“针对疾病的保护”、“保护性免疫”、“功能性免疫”和类似短语，意指针对通过施用本发明的一种或多种治疗组合物或其组合生成的疾病或状况的应答，其导致比已暴露于疾病或感染的非免疫对象中预期的更少的有害效应。即，感染的有害效应的严重性在接种疫苗的对象中减轻。在接种疫苗的对象中可以减少、减缓或可能完全预防感染。在本文中，当意欲完全预防感染时，它被具体陈述。如果未陈述完全预防，则该术语包括部分预防。
在本文中，“临床体征的发生率和/或严重性的减少”或“临床症状的减少”意指但不限于与野生型感染相比，减少组中受感染对象的数目、减少或消除显示出感染的临床体征的对象数目、或减少一个或多个对象中存在的任何临床体征的严重性。例如，它应该指病原体负荷的减少、病原体脱落、病原体传播中的减少、或PEDV症状的任何临床体征的减少。优选地，与未接受组合物并且变得被感染的对象相比，接受本发明的治疗组合物的一个或多个对象中这些临床体征减少至少10％。更优选地，在接受本发明的组合物的对象中，临床体征减少至少20％，优选至少30％，更优选至少40％，且甚至更优选至少50％。
术语“增加的保护”在本文中意指但不限于，在接种疫苗的对象组相对于未接种疫苗的对照对象组中，与被传染剂(优选PDCoV和/或PEDV)的感染相关的一种或多种临床症状的统计学显著减少。术语“临床症状的统计学显著减少”意指但不限于，在用传染剂攻击后，与未接种疫苗的对照组相比，接种疫苗的对象组中的至少一种临床症状的发生率的频率低至少10％，优选20％，更优选30％，甚至更优选50％，且甚至更优选70％。
“持久保护”应该指“改善的功效”，其持续至少3周，但更优选至少3个月，再更优选至少6个月。在家畜的情况下，最优选的是持久保护应该持续直到动物在其下上市用于肉类的平均年龄。
“免疫原性或免疫学组合物”指包含至少一种猪丁型冠状病毒和/或猪流行性腹泻病毒和/或其免疫原性部分的物质组合物，其在宿主中引发细胞或抗体介导的针对组合物的免疫学应答。在本发明的一个优选实施方案中，免疫原性组合物诱导免疫应答，并且更优选地，赋予针对PDCoV和/或PEDV感染的一种或多种临床体征的保护性免疫。
如本文使用的，免疫原性组合物还指包含本文所述的任何PDCoV和/或PEDV刺突多肽的组合物。根据进一步的实施方案，此类免疫原性组合物进一步包含表达所述PDCoV和/或PEDV刺突蛋白的病毒载体，优选重组杆状病毒的至少一部分。此外，免疫原性组合物可以包含i)上述任何PDCoV或PEDV蛋白，优选以上述浓度，ii)表达所述PDCoV和/或PEDV刺突蛋白的病毒载体，优选重组杆状病毒的至少一部分，以及iii)细胞培养上清液的一部分。
因此，根据一个方面，本发明涉及用于减少猪群中的PDCoV和/或PEDV感染百分比的方法，其包括给所述猪施用有效量的PDCoV和/或PEDV刺突抗原或包含PDCoV和/或PEDV抗原的免疫原性组合物的步骤，其中所述PDCoV和/或PEDV抗原是重组PDCoV和/或PEDV刺突抗原，优选杆状病毒表达的PDCoV和/或PEDV刺突蛋白。优选地，那些重组或杆状病毒表达的PDCoV和/或PEDV刺突具有如本文所述的序列。
“免疫应答”或“免疫学应答”意指但不限于，针对目的组合物或疫苗的细胞和/或抗体介导的免疫应答的发展。通常，免疫应答或免疫学应答包括但不限于下述效应中的一种或多种：抗体、B细胞、辅助性T细胞、抑制性T细胞和/或细胞毒性T细胞的产生或活化，特别针对目的组合物或疫苗中包括的一种或多种抗原。优选地，宿主将展示治疗或保护性免疫(记忆)反应，使得增强针对新感染的抗性和/或减少疾病的临床严重性。此类保护可以通过减少症状数目、症状严重性、或缺乏与病原体感染相关的一种或多种症状、延迟病毒血症发作、减少病毒持续时间、减少总体病毒负荷和/或减少病毒排泄得到证实。
抗原的“免疫保护量”或“免疫有效量”或“产生免疫应答的有效量”是有效诱导受体中的免疫原性应答的量。免疫原性应答可能足以用于诊断目的或其它测试，或者可能足以预防由疾病因子感染引起的疾病的体征或症状，包括不良健康效应或其并发症。可以诱导体液免疫或细胞介导的免疫或两者。例如，间接地通过测量抗体滴度、淋巴细胞增殖测定，或直接地通过监测用野生型毒株攻击后的体征和症状，可以评估动物对免疫原性组合物的免疫原性应答，而由疫苗赋予的保护性免疫可以通过测量例如临床体征中的减少来评估，所述临床体征例如对象的死亡率、发病率、温度数、总体身体状况以及总体健康和表现。免疫应答可以包括但不限于细胞和/或体液免疫的诱导。“免疫原性”意指诱发免疫或抗原应答。因此，免疫原性组合物是诱导免疫应答的任何组合物。
“治疗有效量”指在接受抗原或疫苗的对象中诱导免疫应答的抗原或疫苗的量，其足以预防或减少由被病原体如病毒或细菌感染引起的疾病的临床体征或症状，包括不利的健康影响或其并发症。可以诱导体液免疫或细胞介导的免疫或体液和细胞介导的免疫两者。例如，间接地通过测量抗体滴度、淋巴细胞增殖测定，或直接地通过监测用野生型毒株攻击后的体征和症状，可以评估动物对疫苗的免疫原性应答。由疫苗赋予的保护性免疫可以通过测量例如临床体征中的减少来评估，所述临床体征例如对象的死亡率、发病率、温度数、总体身体状况以及总体健康和表现。治疗有效的疫苗的量可以根据使用的特定佐剂、使用的特定抗原或对象的状况而变化，并且可以由本领域技术人员确定。
如本文使用的，“药学或兽医学上可接受的载体”包括任何和所有溶剂、分散介质、包衣、佐剂、稳定剂、稀释剂、防腐剂、抗菌剂和抗真菌剂、等渗剂、吸附延迟剂等等。在一些优选的实施方案且尤其是包括冻干的免疫原性组合物的那些中，用于本发明中的稳定剂包括用于冻干或冷冻干燥的稳定剂。
在一些实施方案中，本发明的免疫原性组合物含有佐剂。如本文使用的，“佐剂”可以包括氢氧化铝和磷酸铝、皂苷例如Quil A、QS-21(Cambridge Biotech Inc.，CambridgeMA)、GPI-0100(Galenica Pharmaceuticals，Inc.，Birmingham，AL)、油包水型乳状液、水包油型乳状液、水包油包水型乳状液。
乳状液可以特别基于轻质液体石蜡油(欧洲药典型)；类异戊二烯油如角鲨烷或角鲨烯；起因于烯烃，特别是异丁烯或癸烯的低聚反应的油；含有线性烷基的酸或醇的酯，更特别是植物油、油酸乙酯、丙二醇二-(辛酸酯/癸酸酯)、甘油基三-(辛酸酯/癸酸酯)或丙二醇二油酸酯；分支脂肪酸或醇的酯，特别是异硬脂酸酯。该油与乳化剂组合使用以形成乳状液。乳化剂优选为非离子表面活性剂，特别是脱水山梨糖醇、二缩甘露醇(例如脱水甘露醇油酸酯)、二醇、聚甘油、丙二醇和油酸、异硬脂酸、蓖麻油酸或羟基硬脂酸(其是任选乙氧基化的)的酯和聚氧丙烯-聚氧乙烯共聚物嵌段，特别是Pluronic产品，尤其是L121。参见Hunter等人The Theory and Practical Application of Adjuvants(Ed.Stewart-Tull，D.E.S.)，JohnWiley和Sons，NY，第51-94页(1995)和Todd等人Vaccine 15:564-570(1997)。示例性佐剂是在由M.Powell和M.Newman，Plenum Press，1995编辑的“Vaccine Design，TheSubunit and Adjuvant Approach”的第147页上描述的SPT乳状液，以及在同一本书的第183页上描述的乳状液MF59。
关于合适的水包油型乳状液的实例是基于的佐剂，例如(水包油型乳状液o/w)、(具有二甲基二十八烷基溴化铵(DDA)的水包油型(o/w))、(具有专利权免疫刺激剂的水包油型(o/w))、(具有交联聚合物的由水包油型(o/w)组成的双重佐剂)、以及(不含动物起源组分的水包油型乳状液)。(MVP Technologies，Inc.Omaha，Nebr.，USA)。包含灭活的PDCoV、灭活的PEDV和/或重组的PDCoV和/或PEDV蛋白的药物/疫苗组合物已用水包油型乳状液，优选用此类基于的佐剂，更优选用(水包油型乳状液o/w)和/或(不含动物起源组分的水包油型乳状液)有效地佐剂化。
用于兽医疫苗中的合适的吸附剂氢氧化铝凝胶的实例包括 REHYDRAGEL-LV；REHYDRAGEL-HPA；REHYDRAPHOS(GeneralChemical，Berkeley Heights，New Jersey，USA)。
佐剂的进一步实例是选自丙烯酸或甲基丙烯酸的聚合物和马来酸酐与烯基衍生物的共聚物的化合物。有利的佐剂化合物是尤其与糖或多元醇的聚烯基醚交联的丙烯酸或甲基丙烯酸的聚合物。这些化合物通过术语卡波姆(Phameuropa第8卷，No.2，1996年6月)已知。本领域技术人员还可以参考美国专利2,909,462，其描述了与具有至少3个羟基，优选不超过8个羟基的多羟基化的化合物交联的此类丙烯酸聚合物，至少三个羟基的氢原子被替换为具有至少2个碳原子的不饱和脂族原子团。优选的原子团是含有2至4个碳原子的那些，例如，乙烯基、烯丙基和其它烯属不饱和基团。不饱和原子团本身可以含有其它取代基，例如甲基。以名称CARBOPOL.RTM.销售的产品(也称为聚丙烯酸)；(BF Goodrich，Ohio，USA)是特别合适的。它们与烯丙基蔗糖或烯丙基季戊四醇交联。其中，可以提到CARBOPOL.RTM.974P(也称为聚丙烯酸)、CARBOPOL.RTM.934P(也称为聚丙烯酸)和CARBOPOL.RTM.971P(也称为聚丙烯酸)。最优选的是使用CARBOPOL.RTM.971P(也称为聚丙烯酸)。在马来酸酐和链烯基衍生物的共聚物中，共聚物EMA(Monsanto)是马来酸酐和乙烯的共聚物。这些聚合物在水中的溶解导致酸溶液被中和，优选至生理学pH，以便得到免疫原性、免疫学或疫苗组合物本身掺入其内的佐剂溶液。
进一步合适的佐剂尤其包括但不限于RIBI佐剂系统(Ribi Inc.)、嵌段共聚物(CytRx，Atlanta Ga.)、SAF-M(Chiron，Emeryville Calif.)、单磷酰脂质A、阿夫立定脂质-胺佐剂、来自大肠杆菌(E.coli)的热不稳定性肠毒素(重组或其它)、霍乱毒素或胞壁酰二肽。
预期佐剂可以以每剂量约100μg至约10mg的量，优选以每剂量约100μg至约10mg的量，更优选以每剂量约500μg至约5mg的量，甚至更优选以每剂量约750μg至约2.5mg的量，且最优选以每剂量约1mg的量加入。可替代地，佐剂可以以按最终产物的体积计约0.01至50％的浓度，优选以约2％至30％的浓度，更优选以约5％至25％的浓度，再更优选以约7％至22％的浓度，且最优选以10％至20％的浓度。
“稀释剂”可以包括水、盐水、右旋糖、乙醇、甘油等等。等渗剂可以尤其包括氯化钠、右旋糖、甘露醇、山梨糖醇和乳糖。稳定剂尤其包括白蛋白和乙二胺四乙酸的碱金属盐等。
“分离的”意指“通过人为”从其天然状态改变，即，如果它在自然界中出现，则它已从其原始环境改变或取出、或两者。例如，天然存在于活生物中的多核苷酸或多肽不是“分离的”，但与其天然状态的共存材料分开的相同多核苷酸或多肽是“分离的”，如该术语在本文中采用的。
“安全性”指在疫苗接种后，在接种疫苗的动物中不存在不良后果，包括但不限于：基于病毒的疫苗可能回复毒力、临床上显著的副作用如持续的全身性疾病或在疫苗施用部位处无法接受的炎症。
如本文使用的，术语“疫苗接种”或“接种疫苗”或其变体意指但不限于，包括施用本发明的免疫原性组合物的过程，当施用于动物时，所述免疫原性组合物在动物中引发、或者能够直接或间接地引发针对PDCoV和/或PEDV的免疫应答。
在本发明的上下文中，“死亡率”指通过PDCoV感染、PEDV感染和/或由PDCoV和PEDV的共感染引起的死亡，并且包括其中感染如此严重使得动物被实施安乐死以预防痛苦并且对其生命提供人道结局的情况。
在本文中，“有效剂量”意指但不限于引发或能够引发免疫应答的抗原量，所述免疫应答在抗原施用于其的动物中产生临床症状的减少。
如本领域已知的，“序列同一性”指两个或更多个多肽序列或者两个或更多个多核苷酸序列，即参考序列和待与参考序列比较的给定序列之间的关系。通过在序列已最佳比对以产生最高程度的序列相似性之后，将给定序列与参考序列进行比较来确定序列同一性，如通过此类序列的串之间的匹配所确定的。在此类比对后，在逐个位置的基础上确定序列同一性，例如，如果在该位置处，核苷酸或氨基酸残基是相同的，则序列在特定位置处是“相同的”。然后将此类位置同一性的总数目除以参考序列中的核苷酸或残基的总数目，以得到％序列同一性。序列同一性可以通过已知方法容易地计算，所述方法包括但不限于以下中描述的那些：Computational Molecular Biology，Lesk，A.N.，编辑，OxfordUniversity Press，New York(1988)，Biocomputing：Informatics and Genome Projects，Smith，D.W.，编辑，Academic Press，New York(1993)；Computer Analysis of SequenceData，Part I，Griffin，A.M.和Griffin，H.G.，编辑，Humana Press，New Jersey(1994)；Sequence Analysis in Molecular Biology，von Heinge，G.，Academic Press(1987)；Sequence Analysis Primer，Gribskov，M.和Devereux，J.，编辑，M.Stockton Press，NewYork(1991)；以及Carillo，H.和Lipman，D.，SIAM J.Applied Math.，48：1073(1988)，其教导引入本文作为参考。确定序列同一性的优选方法设计为给出测试序列之间的最大匹配。确定序列同一性的方法在可公开获得的计算机程序中编码，所述程序确定给定序列之间的序列同一性。此类程序的实例包括但不限于GCG程序包(Devereux，J.等人Nucleic AcidsResearch，12(1):387(1984))、BLASTP、BLASTN和FASTA(Altschul，S.F.等人J.Molec.Biol.，215:403-410(1990)。BLAST程序可从NCBI和其它来源公开获得(BLASTManual，Altschul，S.等人NCVI NLM NIH Bethesda，MD 20894，Altschul，S.F.等人J.Molec.Biol.，215:403-410(1990)，其教导引入本文作为参考)。这些程序使用缺省缺口权重最佳地比对序列，以便产生给定序列和参考序列之间的最高水平的序列同一性。作为说明，通过具有与参考核苷酸序列具有至少例如85％，优选90％，甚至更优选95％“序列同一性”的核苷酸序列的多核苷酸，预期给定多核苷酸的核苷酸序列与参考序列相同，除了给定多核苷酸序列可以包括参考核苷酸序列的至多15个，优选至多10个，甚至更优选至多5个点突变/100个核苷酸之外。换言之，在具有相对于参考核苷酸序列具有至少85％，优选90％，甚至更优选95％同一性的核苷酸序列的多核苷酸中，参考序列中至多15％，优选10％，甚至更优选5％的核苷酸可以被缺失或用另一种核苷酸取代，或者可以将参考序列中的总核苷酸的至多15％，优选10％，甚至更优选5％的多个核苷酸插入参考序列内。参考序列的这些突变可以出现在参考核苷酸序列的5'或3'末端位置处或者这些末端位置之间的任何位置，个别散布在参考序列中的核苷酸中或者参考序列内的一个或多个邻接组中。类似地，通过具有与参考氨基酸序列具有至少例如85％，优选90％，甚至更优选95％的序列同一性的给定氨基酸序列的多肽，预期多肽的给定氨基酸序列与参考序列相同，除了给定的多肽序列可以包括参考氨基酸序列的至多15个，优选至多10个，甚至更优选至多5个氨基酸改变/各100个氨基酸之外。换言之，为了获得与参考氨基酸序列具有至少85％，优选90％，甚至更优选95％的序列同一性的给定多肽序列，参考序列中至多15％，优选至多10％，甚至更优选至多5％的氨基酸残基可以被缺失或用另一种氨基酸取代，或者可以将参考序列中的氨基酸残基总数目的至多15％，优选至多10％，甚至更优选至多5％的多个氨基酸插入参考序列内。参考序列的这些改变可以出现在参考氨基酸序列的氨基或羧基末端位置处或者这些末端位置之间的任何位置，个别散布在参考序列中的残基中或者参考序列内的一个或多个邻接组中。优选地，并不相同的残基位置因保守氨基酸取代而不同。然而，在确定序列同源性时，保守取代不包括作为匹配。
“保守取代”指氨基酸残基由具有相似特征或特性(包括大小、疏水性等)的另一种氨基酸残基的取代，使得总体功能性不显著改变。
如本文使用的，“序列同源性”指确定扣除保守取代的两个序列的相关性的方法。为了确定序列同源性，最佳比对两个或更多个序列，并且在需要时引入缺口。换言之，为了获得与参考序列具有95％序列同源性的多肽，参考序列中85％，优选90％，甚至更优选95％的氨基酸残基必须与另一种氨基酸匹配或包含由另一种氨基酸的保守取代，或者参考序列中至多15％，优选至多10％，甚至更优选至多5％的总氨基酸残基的多个氨基酸(不包括保守取代的氨基酸)可以插入参考序列内。优选地，同源序列包含至少50个，甚至更优选100个，甚至更优选250个，甚至更优选500个氨基酸的段。
术语“序列同一性”或“百分比同一性”在本文中可互换使用。为了本发明的目的，此处定义为了确定两个氨基酸序列或两个核酸序列的百分比同一性，将序列比对用于最佳比较目的(例如，可以在第一氨基酸或核酸的序列中引入缺口，用于与第二氨基酸或核酸序列的最佳比对)。然后比较在相应的氨基酸或核苷酸位置处的氨基酸或核苷酸残基。当第一序列中的位置被与第二序列中的相应位置相同的氨基酸或核苷酸残基占据时，则该分子在该位置处是相同的。两个序列之间的百分比同一性是由序列共享的相同位置数目的函数(即，％同一性＝相同位置的数目/位置(即重叠位置)总数目x 100)。优选地，两个序列具有相同的长度。当在提及蛋白质中使用序列同一性时，认识到并非相同的残基位置经常因保守氨基酸取代而不同，其中氨基酸残基取代具有相似化学特性(例如电荷或疏水性)的其它氨基酸残基，并且因此不改变分子的功能特性。当序列在保守取代方面不同时，可以向上调整百分比序列同一性，以校正取代的保守性质。通过此类保守取代而不同的序列被称为具有“序列相似性”或“相似性”。
可以在待比较的两个序列的整个长度上或在两个序列的片段上进行序列比较。通常，比较将在待比较的两个序列的全长上进行。然而，序列同一性可以在例如二十、五十、一百或更多个邻接氨基酸残基的区域上进行。
本领域技术人员将意识到几种不同的计算机程序可用于确定两个序列之间的同源性的事实。例如，可以使用数学算法完成序列的比较和两个序列之间的百分比同一性的确定。在一个优选的实施方案中，使用Needleman和Wunsch(J.Mol.Biol.(48):444-453(1970))算法确定两个氨基酸或核酸序列之间的百分比同一性，所述算法已掺入GAP程序内的Accelrys GCG软件包中(可在http://www.accelrys.com/products/gcg/处获得)，使用Blosum 62矩阵或PAM250矩阵，以及16、14、12、10、8、6或4的缺口权重和1、2、3、4、5或6的长度权重。技术人员将了解，使用不同的算法时，所有这些不同的参数将产生略微不同的结果，但两个序列的总体百分比同一性并不显著改变。
本发明的蛋白质序列或核酸序列可以进一步用作“查询序列”，以针对公共数据库进行搜索，以例如鉴定其它家族成员或相关序列。可以使用Altschul等人(1990)J.Mol.Biol.215:403-10的BLASTN和BLASTP程序(版本2.0)进行此类搜索。BLAST蛋白质搜索可以用BLASTP程序、得分＝50、字长＝3进行，以获得与本发明的蛋白质分子同源的氨基酸序列。为了获得用于比较目的的加缺口的比对，可以利用如Altschul等人(1997)NucleicAcids Res.25(17):3389-3402中所述的Gapped BLAST。当利用BLAST和Gapped BLAST程序时，可以使用分别程序(例如，BLASTP和BLASTN)的缺省参数。参见在http://www.ncbi.nlm.nih.gov/处的美国国家生物技术信息中心(National Center forBiotechnology Information)主页。
本发明的请求保护的PEDV还应该涵盖PEDV分离株1251-125-10(“125-10”)的变体及其亚片段的变体。此类变体具有与Oklahoma毒株(SEQ ID NO 29和34)的特征基本相同的免疫学特性。术语“具有基本上相同的免疫学特性”涵盖(但不限于)所述变体基本上有效地治疗或预防由如下所述的PEDV引起的临床体征或者改善如下所述的功效参数。
本发明的请求保护的PDCoV还应该涵盖PDCoV分离株NSVL、PDCoV 2.0307和PDCoV5.0327的变体及其亚片段的变体。此类变体具有与NSVL毒株(SEQ ID NO：1和2)，以及分离株PDCoV 2.0307(SEQ ID NO：5和6)和PDCoV 5.0327(SEQ ID NO：9和10)的特征基本上相同的免疫学特性。术语“具有基本上相同的免疫学特性”涵盖(但不限于)所述变体在治疗或预防由如本文所述的PDCoV引起的临床体征或者改善如本文所述的功效参数方面基本上有效。除了可以用于疫苗中的各种PDCoV毒株外，重组刺突蛋白抗原，包括其亚片段，也可以用于疫苗中。同样地，示例性刺突蛋白序列包括但不限于具有与下文列出的PDCoV分离株或‘变体’基本上相同的免疫学特性的那些。
关于PDCoV序列(SEQ ID NO:1、2、3、4、5、6、7、8、9、10、11、12,17、18、27和28)或PEDV序列SEQ ID NO:29、30、31、32、33、34、35、36、46、47、52和53)(例如，多肽或核酸序列)的术语“变体”预期意指基本上相似的序列。对于包含开放读码框的核苷酸序列，变体包括由于遗传密码的简并性而编码天然蛋白的相同氨基酸序列的那些序列。变体核苷酸序列还包括合成衍生的核苷酸序列，例如通过使用定点诱变和开放读码框生成的那些，编码天然蛋白质，以及编码相对于天然蛋白质具有氨基酸取代用于密码子优化目的的多肽的那些。一般地，使用标准参数，使用描述的比对程序之一，与参考序列相比较，本发明的核苷酸序列变体将具有至少80％、81％、82％、83％、84％、85％、86％、87％、88％或89％，更优选至少90％、91％、92％、93％或94％，且最优选至少95％、96％、96.1％、96.2％、96.3％、96.4％、96.5％、96.6％、96.7％、96.8％、96.9％、97％、97.1％、97.2％、97.3％、97.4％、97.41％、97.42％、97.43％、97.44％、97.45％、97.46％、97.47％、97.48％、97.49％、97.5％、97.51％、97.52％、97.53％、97.54％、97.55％、97.56％、97.57％、97.58％、97.59％、97.6％、97.61％、97.62％、97.63％、97.64％、97.65％、97.66％、97.67％、97.68％、97.69％97.7％、97.71％、97.72％、97.73％、97.74％、97.75％、97.76％、97.77％、97.78％、97.79％、97.8％、97.81％、97.82％、97.83％、97.84％、97.85％、97.86％、97.87％、97.88％、97.89％、97.9％、97.91％、97.92％、97.93％、97.94％、97.95％、97.96％、97.97％、97.98％、97.99％、98％、98.01％、98.02％、98.03％、98.04％、98.05％、98.06％、98.07％、98.08％、98.09％、98.1％、98.11％、98.12％、98.13％、98.14％、98.15％、98.16％、98.17％、98.18％、98.19％、98.2％、98.21％、98.22％、98.23％、98.24％、98.25％、98.26％、98.27％、98.28％、98.29％、98.3％、98.31％、98.32％、98.33％、98.34％、98.35％、98.36％、98.37％、98.38％、98.39％、98.4％、98.41％、98.42％、98.43％、98.44％、98.45％、98.46％、98.47％、98.48％、98.49％,98.5％、98.51％、98.52％、98.53％、98.54％、98.55％、98.56％、98.57％、98.58％、98.59％、98.6％、98.61％、98.62％、98.63％、98.64％、98.65％、98.66％、98.67％、98.68％、98.69％、98.7％、98.71％、98.72％、98.73％、98.74％、98.75％、98.76％、98.77％、98.78％、98.79％、98.8％、98.81％、98.82％、98.83％、98.84％、98.85％、98.86％、98.87％、98.88％、98.89％、98.9％、98.91％、98.92％、98.93％、98.94％、98.95％、98.96％、98.97％、98.98％、98.99％、99％、99.01％、99.02％、99.03％、99.04％、99.05％、99.06％、99.07％、99.08％、99.09％、99.1％、99.11％、99.12％、99.13％、99.14％、99.15％、99.16％、99.17％、99.18％、99.19％、99.2％、99.21％、99.22％、99.23％、99.24％、99.25％、99.26％、99.27％、99.28％、99.29％、99.3％、99.31％、99.32％、99.33％、99.34％、99.35％、99.36％、99.37％、99.38％、99.39％、99.4％、99.41％、99.42％、99.43％、99.44％、99.45％、99.46％、99.47％、99.48％、99.49％、99.5％、99.51％、99.52％、99.53％、99.54％、99.55％、99.56％、99.57％、99.58％、99.59％、99.6％,99.61％、99.62％、99.63％、99.64％、99.65％、99.66％、99.67％、99.68％、99.69％、99.7％、99.71％、99.72％、99.73％、99.74％、99.75％、99.76％、99.77％、99.78％、99.79％、99.8％、99.81％、99.82％、99.83％99.84％、99.85％、99.86％、99.87％、99.88％、99.89％、99.9％、99.91％、99.92％、99.93％、99.94％、99.95％、99.96％、99.97％、99.98％和99.99％的序列同一性。
关于PDCoV序列(SEQ ID NO:1、2、3、4、5、6、7、8、9、10、11、12,17、18、27和28)(例如，多肽或核酸序列)的术语“变体”预期意指基本上相似的序列。因此，本发明的免疫原性组合物可以有效地掺入所有公认的PDCoV毒株或分离株，优选包括但不必限于与以下具有至少约90％总体核苷酸同一性的所有毒株：保藏为GenBank登录号KP981395.1的分离株NSVL-SCDV(猪丁型冠状病毒毒株USA/IL/2014/026PDV_P11)；保藏为GenBank登录号KP757890的分离株CHN-AH-2004；保藏为GenBank登录号KP757891的分离株CHN-HB-2014；保藏为GenBank登录号：KP757892的分离株CHN-JS-2014；保藏为GenBank登录号KR131621的分离株PDCoV/CHJXNI2/2015；保藏为GenBank登录号KR150443的毒株USA/Arkansas61/2015；保藏为GenBank登录号KR265847的毒株USA/Minnesota442/2014；保藏为GenBank登录号KR265848的毒株USA/Minnesota214/2014；保藏为GenBank登录号KR265849的毒株USA/Michigan447/2014；保藏为GenBank登录号KR265850的毒株USA/Michigan448/2014；保藏为GenBank登录号KR265851的毒株USA/Indiana453/2014；保藏为GenBank登录号KR265852的毒株USA/Illinois449/2014；保藏为GenBank登录号KR265853的毒株USA/Minnesota/2013；保藏为GenBank登录号KR265854的毒株USA/Minnesota454/2014；保藏为GenBank登录号KR265855的毒株USA/Minnesota455/2014；保藏为GenBank登录号KR265856的毒株USA/Illinois272/2014；保藏为GenBank登录号KR265857的毒株USA/Illinois273/2014；保藏为GenBank登录号KR265858的毒株USA/NorthCarolina452/2014；保藏为GenBank登录号KR265859的毒株USA/Minnesota159/2014；保藏为GenBank登录号KR265860的毒株USA/Nebraska209/2014；保藏为GenBank登录号KR265861的毒株USA/Nebraska210/2014；保藏为GenBank登录号KR265862的毒株USA/Ohio444/2014；保藏为GenBank登录号KR265863的毒株USA/Ohio445/2014；保藏为GenBank登录号KR265864的毒株USA/Minnesota292/2014；保藏为GenBank登录号KR265865的毒株USA/Iowa459/2014；保藏为GenBank登录号KT021234的毒株CH/SXD1/2015；保藏为GenBank登录号KT266822的毒株CH/Sichuan/S27/2012；保藏为GenBank登录号KT336560的分离株CHN-HN-2014；保藏为GenBank登录号KU051641的毒株PDCoV/Swine/Thailand/S5011/2015；保藏为GenBank登录号KU051649的毒株PDCoV/Swine/Thailand/S5015L/2015；保藏为GenBank登录号KU981059的毒株NH；保藏为GenBank登录号KU981060的毒株NH分离株第0代；保藏为GenBank登录号KU981061的NH分离株第5代；保藏为GenBank登录号KU981062的毒株NH分离株第10代；保藏为GenBank登录号KU984334的分离株TT_1115；保藏为GenBank登录号KX022602的毒株PDCoV/USA/Iowa136/2015；保藏为GenBank登录号KX022603的毒株PDCoV/USA/Minnesota140/2015；保藏为GenBank登录号KX022604的毒株PDCoV/USA/Nebraska137/2015；保藏为GenBank登录号KX022605的毒株PDCoV/USA/Nebraska145/2015；以及保藏为GenBank登录号KX118627的分离株P1_16_BTL_0115/PDCoV/2016/Lao。
如本领域已知的，术语“基因群”指属内的相关病毒；其可以进一步细分成遗传聚类。鉴定的PEDV基因群包括包含亚群G1a、G1b、R(减毒/适应)的群G1；以及包含亚群G2a和G2b的G2。G2a基因群的成员包括共享几个独特的核苷酸变化的中国毒株AH2012(GenBank登录号：KC210145)和北美毒株。分别地，毒株MN和IA2与AH2012具有99.6％的核苷酸同一性，并且毒株IA1与AH2012具有99.5％的核苷酸同一性。研究人员已推测，AH2012样病毒可能传播到中国东部地区，然后运输至美国，并且很可能是最接近北美株的祖先。基因群2a的成员与基因群1a的原型PEDV毒株CV777仅共享大约96.9％的相似性(Bridgen等人1993；Huang等人2013；GenBank：AF353511.1)。像这样，基于历史CV777衍生的G1a毒株或DR13衍生的G1b毒株的减毒PEDV疫苗可以与新近出现的中国和北美G2a PEDV毒株在抗原上较不相关。
密切相关的北美PEDV分离株US/Colorado/2013(GenBank登录号：KF272920.1)也已通过Marthaler等人，2013得到报道。如同上述北美分离株，CO/13的完全PEDV基因组与GenBank中可获得的其它完全PEDV基因组具有96.5％至99.5％的核苷酸同一性，与中国毒株AH2012(GenBank登录号KC210145)具有最高的核苷酸同一性(99.5％)。中国毒株株AH2012是2a基因群的成员。北美分离株CO/13的完全基因组与PEDV参考毒株CV777的那种的比较，显示CO/13含有1个核苷酸的插入(在位置48处)和5'UTR中5个核苷酸的缺失(在位置73和83至86处)，而刺突基因含有16个核苷酸的插入(位置20804、20810至20820、20843和21053至21055)和7个核苷酸的缺失(位置20853和21118至21124)。
已出现了PEDV的其它变体，称为'INDEL'毒株，其与较老的原型毒株相比经常是天然减毒的。这些也可以用作其中病毒是活减毒的或灭活的疫苗。在此类情况下，可能仅需要最低限度的进一步传代来提供安全的疫苗减毒。因此，本发明的示例性疫苗病毒还包括对于刺突蛋白或基于完整的病毒序列，与此类毒株具有至少95％、96％、96.1％、96.2％、96.3％、96.4％、96.5％、96.6％、96.7％、96.8％、96.9％、97％、97.1％、97.2％、97.3％、97.4％、97.41％、97.42％、97.43％、97.44％、97.45％、97.46％、97.47％、97.48％、97.49％、97.5％、97.51％、97.52％、97.53％、97.54％、97.55％、97.56％、97.57％、97.58％、97.59％、97.6％、97.61％、97.62％、97.63％、97.64％、97.65％、97.66％、97.67％、97.68％、97.69％97.7％、97.71％、97.72％、97.73％、97.74％、97.75％、97.76％、97.77％、97.78％、97.79％、97.8％、97.81％、97.82％、97.83％、97.84％、97.85％、97.86％、97.87％、97.88％、97.89％、97.9％、97.91％、97.92％、97.93％、97.94％、97.95％、97.96％、97.97％、97.98％、97.99％、98％、98.01％、98.02％、98.03％、98.04％、98.05％、98.06％、98.07％、98.08％、98.09％、98.1％、98.11％、98.12％、98.13％、98.14％、98.15％、98.16％、98.17％、98.18％、98.19％、98.2％、98.21％、98.22％、98.23％、98.24％、98.25％、98.26％、98.27％、98.28％、98.29％、98.3％、98.31％、98.32％、98.33％、98.34％、98.35％、98.36％、98.37％、98.38％、98.39％、98.4％、98.41％、98.42％、98.43％、98.44％、98.45％、98.46％、98.47％、98.48％、98.49％,98.5％、98.51％、98.52％、98.53％、98.54％、98.55％、98.56％、98.57％、98.58％、98.59％、98.6％、98.61％、98.62％、98.63％、98.64％、98.65％、98.66％、98.67％、98.68％、98.69％、98.7％、98.71％、98.72％、98.73％、98.74％、98.75％、98.76％、98.77％、98.78％、98.79％、98.8％、98.81％、98.82％、98.83％、98.84％、98.85％、98.86％、98.87％、98.88％、98.89％、98.9％、98.91％、98.92％、98.93％、98.94％、98.95％、98.96％、98.97％、98.98％、98.99％、99％、99.01％、99.02％、99.03％、99.04％、99.05％、99.06％、99.07％、99.08％、99.09％、99.1％、99.11％、99.12％、99.13％、99.14％、99.15％、99.16％、99.17％、99.18％、99.19％、99.2％、99.21％、99.22％、99.23％、99.24％、99.25％、99.26％、99.27％、99.28％、99.29％、99.3％、99.31％、99.32％、99.33％、99.34％、99.35％、99.36％、99.37％、99.38％、99.39％、99.4％、99.41％、99.42％、99.43％、99.44％、99.45％、99.46％、99.47％、99.48％、99.49％、99.5％、99.51％、99.52％、99.53％、99.54％、99.55％、99.56％、99.57％、99.58％、99.59％、99.6％,99.61％、99.62％、99.63％、99.64％、99.65％、99.66％、99.67％、99.68％、99.69％、99.7％、99.71％、99.72％、99.73％、99.74％、99.75％、99.76％、99.77％、99.78％、99.79％、99.8％、99.81％、99.82％、99.83％99.84％、99.85％、99.86％、99.87％、99.88％、99.89％、99.9％、99.91％、99.92％、99.93％、99.94％、99.95％、99.96％、99.97％、99.98％和99.99％的序列同一性或更高序列同一性的那些，无论是测量氨基酸还是编码核苷酸序列。
术语“北美起源的PEDV”意指包含SEQ ID NO：29和/或33和/或SEQ ID NO：32和/或36的PEDV分离株，和/或与SEQ ID NO：29和/或33具有至少99％序列同一性的任何PEDV分离株，和/或与SEQ ID.NO:29和/或33的RNA等价物至少99％相同的PEDV分离株，和/或其中刺突蛋白由SEQ ID NO：30、34、46或52编码的PEDV分离株，和/或其中刺突蛋白与SEQ ID：30、34、46或52具有至少98％的序列同一性的PEDV分离株，和/或其中表达的刺突蛋白与SEQ IDNO：31、35、47、53具有至少90％同源性的任何PEDV分离株。
如本领域已知的，术语“进化枝”指由祖先及其所有后代组成的组，系统发育树中的单个“分支”。作为一个例子，祖先可以是个体、种群或物种。基因群可以包括多个进化枝，例如AH2012在与北美分离株不同的进化枝中。
根据一个进一步的实施方案，本发明还涉及包含如本文所述的任何此类核酸分子的载体。换言之，本发明涉及载体，其包括任何此类刺突、M、E、N PEDV蛋白或其部分的编码序列。优选地，所述载体是表达载体，其允许表达任何此类此类刺突、M、E和/或N PEDV蛋白或蛋白质的部分。根据本发明的载体是适合于在体外或体内转染或感染细菌、酵母或动物细胞的那些。
用于制备和/或使用用于表达的载体(或重组体)的载体和方法可以通过以下中公开的方法或类似于以下中公开的方法：美国专利4,603,112、4,769,330、5,174,993、5,505,941、5,338,683、5,494,807、4,722,848、5,942,235、5,364,773、5,762,938、5,770,212、5,942,235、382,425，PCT公开WO 94/16716、WO 96/39491、WO 95/30018；Paoletti，"Applications of pox virus vectors to vaccination:An update，"PNAS USA 93:11349-11353，October 1996；Moss，"Genetically engineered poxviruses forrecombinant gene expression，vaccination，and safety,"PNAS USA 93:11341-11348，October 1996；Smith等人，美国专利号4,745,051(重组杆状病毒)；Richardson，C.D.(编辑)，Methods in Molecular Biology 39，"Baculovirus Expression Protocols"(1995Humana Press Inc.)；Smith等人，"Production of Human Beta Interferon inInsect Cells Infected with a Baculovirus Expression Vector"，Molecular andCellular Biology，December，1983，第3卷，No.12，第2156-2165页；Pennock等人，"Strongand Regulated Expression of Escherichia coli B-Galactosidase in Infect Cellswith a Baculovirus vector，"Molecular and Cellular Biology March 1984，第4卷，No.3，第406页；EPA0 370 573；于1986年10月16日提交的美国申请号920,197；EP专利公开号265785；美国专利号4,769,331(重组疱疹病毒)；Roizman，"The function of herpessimplex virus genes:A primer for genetic engineering of novel vectors,"PNASUSA 93:11307-11312，October 1996；Andreansky等人，"The application ofgenetically engineered herpes simplex viruses to the treatment ofexperimental brain tumors,"PNAS USA 93:11313-11318，October 1996；Robertson等人，"Epstein-Barr virus vectors for gene delivery to B lymphocytes"，PNAS USA93:11334-11340，October 1996；Frolov等人，"Alphavirus-based expression vectors:Strategies and applications,"PNAS USA 93:11371-11377，October 1996；Kitson等人，J.Virol.65，3068-3075，1991；美国专利号5,591,439、5,552,143；WO 98/00166；两者均于1996年7月3日提交的容许的美国申请序列号08/675,556和08/675,566(重组腺病毒)；Grunhaus等人，1992，"Adenovirus as cloning vectors,"Seminars in Virology(第3卷)第237-52页，1993；Ballay等人EMBO Journal，第4页，第3861-65页，Graham，Tibtech 8，85-87，April，1990；Prevec等人，J.Gen Virol.70，42434；PCT WO 91/11525；Felgner等人(1994)，J.Biol.Chem.269，2550-2561，Science，259:1745-49，1993；and McClements等人，"Immunization with DNA vaccines encoding glycoprotein D or glycoprotein B，alone or in combination，induces protective immunity in animal models ofherpes simplex virus-2disease"，PNAS USA 93:11414-11420，October 1996；以及美国专利号5,591,639、5,589,466和5,580,859，以及WO 90/11092、WO93/19183、WO94/21797、WO95/11307、WO95/20660；Tang等人，Nature，以及Furth等人，Analytical Biochemistry，relating to DNA expression vectors，inter alia.See also WO 98/33510；Ju等人，Diabetologia，41:736-739，1998(慢病毒表达系统)；Sanford等人，美国专利号4,945,050；Fischbach等人(Intracel)；WO 90/01543；Robinson等人，Seminars in Immunology第9卷，第271-283页(1997)(DNA载体系统)；Szoka等人，美国专利号4,394,448(将DNA插入活细胞内的方法)；McCormick等人，美国专利号5,677,178(使用致细胞病变病毒)；以及美国专利号5,928,913(用于基因递送的载体)；以及本文引用的其它文件。
优选的病毒载体包括杆状病毒，例如BaculoGold(BD Biosciences Pharmingen，San Diego，Calif.)，特别地条件是生产细胞是昆虫细胞。尽管杆状病毒表达系统是优选的，但本领域技术人员应理解，其它表达系统对于本发明的目的起作用，即将HG表达到细胞培养物的上清液内。此类其它表达系统可能需要使用信号序列以便使H5表达到培养基内。
有效剂量：
本文所述的化合物可以以治疗有效剂量施用于对象，以预防PDCoV和/或PEDV相关疾病。剂量取决于接受疫苗的宿主以及因素如宿主的大小、重量和年龄。
用于制剂中的本发明的免疫原性组合物的精确量将取决于施用途径和对象的性质(例如，年龄、大小、疾病的阶段/水平)，并且应该根据从业者的判断和根据标准临床技术的每个对象的情况来决定。有效免疫量是足以治疗或预防对象中的PDCoV和/或PEDV传染病的量。
组合物的免疫原性可以通过经由使用本领域已知的任何免疫测定监测用组合物免疫后测试对象的免疫应答来确定。体液(抗体)应答和/或细胞介导的免疫的生成可以理解为免疫应答的指示。测试对象可以包括动物，例如猪、小鼠、仓鼠、犬、猫、兔、牛、马、绵羊、家禽(例如鸡、鸭、鹅和火鸡)和人。
可以通过各种方法分析测试对象的免疫应答，所述方法例如：如通过已知技术例如酶联免疫吸附测定(ELISA)、免疫印迹、免疫沉淀、病毒中和等测定的，所得的免疫血清对免疫原性缀合物的反应性；或者，如通过本领域已知用于测定传染病因子的水平，例如病毒水平(例如，通过培养来自对象的样品)的任何方法，或本领域已知的其它技术确定的，通过保护免疫宿主免于被病原体感染和/或免疫宿主中由于被病原体感染的症状的减弱。还可以通过测量免疫球蛋白针对其的抗原水平来确定传染病因子的水平。传染病因子水平中的降低或传染病症状的改善指示该组合物是有效的。
在体内用于动物之前，可以在体外测试本发明的治疗试剂的所需治疗或预防活性。例如，可以用于确定是否指示特定治疗试剂的施用的体外测定包括体外细胞培养测定，其中来自细胞系的适当细胞或者从患有特定疾病或病症的对象培养的细胞暴露于或以其它方式施用治疗试剂，并且观察治疗试剂对细胞的作用。
可替代地，治疗剂可以通过以下来测定：使治疗剂与易受通过传染病因子的感染但未感染有传染病因子的细胞(从对象或培养的细胞系培养)接触，使细胞暴露于传染病因子，然后确定与治疗剂接触的细胞的感染率是否低于未与治疗剂接触的细胞的感染率。用传染病因子感染细胞可以通过本领域已知的任何方法测定。
另外，可以通过在治疗之前、治疗期间或治疗之后，以合适的时间间隔测量抗体在动物模型或人对象中针对的分子水平来评估治疗剂。可以鉴定分子的量中的任何变化的变化或不存在，并且将其与治疗对于对象的作用相关联。分子的水平可以通过本领域已知的任何方法确定。
在使用本发明的方法和组合物用PDCoV和/或PEDV给动物接种疫苗后，本领域已知的任何结合测定都可以用于评价所得到的抗体和特定分子之间的结合。还可以进行这些测定，以选择对于特定抗原显示出更高亲和力或特异性的抗体。
对受试者的施用：
优选的施用途径包括但不限于鼻内、经口、皮内和肌内。技术人员将认识到，本发明的组合物还可以以一个、两个或更多个剂量，以及通过其它施用途径施用。例如，此类其它途径包括皮下、皮内、静脉内、血管内、动脉内、腹膜内、鞘内、气管内、皮内、心内、叶内、髓内、肺内和阴道内。取决于所需的治疗持续时间和有效性，本发明的组合物可以一次或多次施用，也可以间歇施用，例如在每天的基础上共几天、几周或几个月，并且以不同的剂量。
包括下述实施例以证实本发明的优选实施方案。本领域技术人员应该了解，下述实施例中公开的技术代表由本发明人发现的在本发明的实践中很好地起作用的技术，并且因此可以被视为构成用于其实践的优选模式。然而，根据本公开内容，本领域技术人员应当了解，可以在所公开的具体实施方案中做出许多改变并且仍然获得相同或相似的结果，而不背离本发明的精神和范围。
实施例
实施例1：PEDV毒株的分离和生产
为了产生猪流行性腹泻病毒疫苗，杀死病毒，首先产生PEDV(分离株)的母种毒培养物。从该母种毒生长PEDV培养物，然后灭活。然后将灭活的病毒培养物与佐剂混合，以便产生猪流行性腹泻病毒疫苗。下述方法用于生产猪流行性腹泻病毒疫苗。
在2013年获得显示出极度腹泻的动物来自动物的或组织。从这些动物生成来自粘膜刮屑的匀浆物，通过0.2微米注射器式滤器过滤，并且将滤液用于接种非洲绿猴肾细胞(VERO)。病毒在PEDV维持培养基的存在下生长，所述PEDV维持培养基含有改良MEM、猪胰蛋白酶、胰蛋白胨磷酸盐肉汤、酵母提取物和HEPES缓冲液。通过检查特征性合胞体形成和细胞单层的融合来评估病毒生长且使其可视化。使用基于Illumina的测序仪技术，对CPE阳性材料进行测序。
为了产生PEDV母种毒培养物(“PEDV MSV”)，在BI VERO细胞中分离猪流行性腹泻病毒株(分离株)(PEDV分离株)，并且在BI VERO细胞中总共传代19次，然后病毒在2013 EUVERO细胞中生长至第30代。病毒的第30代被稀释并且记录为母种毒，命名为PEDV KV-1251-125-10-OK。
从母种毒中，通过用在PEDV维持培养基中的PEDV KV-1251-125-10-OK MSV感染2013 EU VERO细胞产生PEDV培养物(KV-1251-125-10-OK，在本文中称为“125-10”)，所述PEDV维持培养基含有改良的最低必需培养基、猪胰蛋白酶(10μg/ml)、胰蛋白胨磷酸盐肉汤(0.3％)、酵母提取物(0.02％)和1M HEPES缓冲液(2.5％)。2013EU VERO细胞通常以104TCID50/850cm2滚瓶的最小剂量用PEDV(125-10)MSV进行感染。此类培养物可以在无菌一次性滚瓶或微载体珠上生长。将培养物在36℃.±2℃.下温育24至48小时，直至观察到致细胞病变效应(“CPE”)。通常，在感染后12小时内可以看到特征性合胞体，合胞体扩增，细胞单层融合24-48小时，随后为细胞脱落。在温育期间，监测培养物的PEDV诱导的CPE，以确保纯PEDV毒株。如果观察到非典型CPE或者存在任何宏观或微观污染证据，则丢弃培养物。将纯病毒培养物无菌收获到无菌聚丙烯大玻璃瓶(carboy)内。将病毒冷冻解冻以释放细胞结合的病毒粒子，并且通过离心或通过经由0.45微米随后为0.2微米的滤器过滤得到澄清。测试大量病毒收获流体，以确保在灭活之前支原体的不存在。将未立即灭活的收获流体贮存于-70℃或更低温度下。
确定收获的流体的体积并且使流体的温度达到36±2℃。将0.4M的2-溴乙撑胺(BEA)溶液与0.3N NaOH的储备溶液混合，以生成二乙烯亚胺(BEI)储备溶液，然后将其加入收获的流体中，以得到最终浓度为5mM的BEI。将流体连续搅拌最少24小时。加入1.0M硫代硫酸钠溶液，以得到5mM的最终最低浓度，以中和任何残留的BEI。灭活的流体可以贮存于-70±3℃下用于长期贮存，或贮存于4±3℃下用于短期贮存。
用BEI处理后，测试培养物诱导PEDV典型的CPE的能力，以确保病毒的灭活。该任务通过将BEI处理的病毒流体经过Vero细胞，并且检查Vero细胞的任何病毒感染来完成。BEI处理的培养液通常贮存于-70℃或更低温度下，直至灭活测定已完成。
通过将灭活的PEDV培养物与佐剂以20％的包含率彻底共混以形成大量系列，将灭活的病毒配制为佐剂化的疫苗。将大量系列维持在2-8℃下，直至转移到含有一个或十个剂量(每剂量2.0ml)的小瓶内。
实施例2：猪丁型冠状病毒毒株的分离和生产
猪丁型冠状病毒(PDCoV)购自National Veterinary Service laboratory(NVSL)，Ames，IA。该病毒在NVSL从2014年来自Illinois的猪肠样品中分离。在5μg/mL TPCK胰蛋白酶的存在下，在无血清培养基中的猪睾丸(ST)细胞上分离病毒。原始病毒在NVSL进行两轮噬斑纯化，并且病毒的第十次传代用作原材料。
将第10代病毒接种并且适应在BI处的猪睾丸(ST)细胞，使用含有具有HEPES的改良的最低必需培养基、胰蛋白磷酸盐胨肉汤、酵母提取物和猪胰蛋白酶的PDCoV维持培养基。将细胞在37℃与5％CO2下温育24-48小时，并且观察PDCoV诱导的致细胞病变效应的证据。当CPE完成并且细胞已脱落时，将烧瓶冷冻解冻一次并且收获裂解产物。然后将收获的材料接种到新的单层上用于继续传代。传代以此类方式继续通过第12次。接种从第12次收获的病毒，并且使其适应命名为“AI-ST 2015”细胞的替代衍生猪睾丸(ST)细胞谱系。以与第11和12次相同的方式，在AI-ST 2015细胞中进行另外四次病毒传代，以获得P16病毒。将P16病毒接种到滚瓶内，得到最终的“P17”母种毒分离株，将其收获，通过0.2μm滤器过滤，合并以获得大量病毒原种并且贮存于-60℃以下。母种毒“P17”的平均滴度被确定为4.91log10FAID50/mL。
实施例3：PEDV分离株和PDCoV分离株的基因组序列分析
样品制备和分析：
在提取之前，用DNA酶和RNA酶的混合物预处理病毒组织培养上清液，以去除残留的宿主细胞基因组核酸。然后使用病毒RNA提取试剂盒(Qiagen，目录#52906)，从核酸酶处理的样品中提取病毒基因组RNA。在提取后，再次用DNA酶处理样品以进一步富集病毒基因组RNA。随后，通过随机引发的逆转录和Klenow片段处理，将病毒基因组RNA转换为双链cDNA(ds cDNA)。然后使用 XT文库制备试剂盒(目录#FC-131-1024)，将ds cDNA产物用于生成基于Illumina 测序仪的测序的文库。每个样品在5'和3'末端两者上均用独特标签加上条形码，以使生物信息错误框并的机会降到最低。该文库使用500循环试剂盒(目录#MS-102-2003)在测序仪上运行，并且使用NextGene(版本2.3.4)和Sequencher软件(版本5.1)的组合分析数据。选择高质量序列作为含有中值Q得分大于25的那些，并且在3'末端处以不超过3个未调用碱基或测量小于16的Q得分的3个连续碱基的切断进行修剪。然后使用在35bp段上85％或更高匹配的标准从头组装序列，以生成关于每种分离株的假定的完全基因组。然后通过基于模板的比对验证关于各自的假定的完全基因组序列，以验证单核苷酸多态性(SNP)或可变的小插入/缺失。
对于PEDV分离株1251-125-10，在修剪低质量数据后生成平均长度为136bp的总共570,253个序列。在那些序列中；484,247(84.9％)组装成长27,995bp的单个重叠群，其通过BLASTn分析揭示与单链RNA甲型冠状病毒PEDV的强烈同一性。总共11个位置显示出在单个核苷酸或小插入/缺失处的多态性，这些位置在表1中列出。
表1：分离株1251-125-10“125-10”中的多态性残基
将1251-125-10(SEQ ID NO：29)的假定的完全/接近完全的PEDV基因组与PEDV的最接近的中国AH2012(GenBank登录号：KC210145)和北美Colorado 2013分离株(GenBank登录号：AGO58924)进行比对。与两种毒株的同一性超过99.2％，指示与两者均在基因群2a中的两种毒株非常密切的关系。
接下来，检查免疫原性刺突蛋白序列与PEDV刺突蛋白的较大GenBank储库的蛋白质同一性/相似性。再次，提交的最接近的GenBank分离株衍生自由University ofMinnesota Veterinary诊断实验室保藏的北美Colorado 2013毒株(GenBank登录号：AGO58924)，其显示出超过99.5％的同一性(1380/1386个相同的氨基酸)(SEQ ID NO：X)。在6个氨基酸变化中，1个是由于在位置23,101处的多态性，其在位置838处大多数编码CGA(Arg)且小部分编码CAA(Gln)。北美Colorado 2013年毒株在该位置处含有Gln。
对于PDCoV，对12个个别分离的样品进行测序。在病毒颗粒保护的核酸消化(RNA酶+DNA酶)后提取材料，以便富集病毒RNA。在提取后，用DNA酶处理核酸以富集RNA。通过使用随机六聚体引发的逆转录和klenow处理生成双链cDNA。然后将这些产物用于文库生成。
使用 XT文库制备试剂盒(Illumina，目录#FC-131-1024)，通过文库生成处理样品用于基于测序仪的测序。每个样品在5'和3'末端两者上均用独特标签加上条形码，以使生物信息错误框并的机会降到最低。该文库使用500循环试剂盒(Illumina，目录#MS-102-2003)在测序仪上运行，并且使用测序软件(SoftGenetics，LLC，版本2.3.4.2)和软件(Gene Codes Corp.，版本5.1)的组合分析数据。选择高质量序列作为含有中值Q得分大于25的那些，并且在3'末端处以不超过3个未调用碱基或测量小于16的Q得分的3个连续碱基的切断进行修剪。然后使用在35bp段上85％或更高匹配的标准从头组装这些序列，并且与参考文件比对，使用85％或更高的标准匹配35bp的伸展。用于比对的参考序列来自从头序列组装或关于Carthage和NVSL PDCoV毒株(NSVL，猪丁型冠状病毒毒株USA/IL/2014/026PDV_P11，Genbank号KP981395.1)的测序仪衍生的参考。
总的来说，每种BI病毒分离株和其它毒株(“Carthage”原始分离株和NVSL毒株)之间的遗传相似性很高(>99.8％同一性)，并且NVSL和BI-ST细胞生长的病毒变体之间的差异显示出来甚至更少的变化(>99.95％的同一性)(表2)。还参见分别示意性地显示了关于PDCoV分离株/毒株的完全基因组核苷酸邻近树、以及关于PDCoV分离株/毒株的刺突氨基酸邻近树的图1和2。比例尺指示p距离。
NVSL和BI-ST细胞生长的病毒的变体之间的相似性很可能是由于一些样品的共享起源。一个值得注意的序列差异是在培养的PDCoV_2201-1分离株内刺突(氨基酸位置229-231)中的框内9bp缺失的存在；但在原始匀浆物或来自猪的材料中未观察到。这个位置看来是IN/DEL变异性的“热点”，因为与“Carthage”分离株没有已知连接的BI PDCoV-5.0327分离株，在“BI-ST”细胞系中也显示出相同的缺失，但在NVSL-ST细胞系中则没有。这些变化可能指示病毒的组织培养适应性。
表2：每种BI病毒分离株之间的遗传相似性。
实施例4：整个PEDV和PDCoV分离株的灭活
通过在VERO细胞中传代来测试每批PEDV病毒或库的灭活。75cm2的24小时细胞培养物用2.0mL灭活的PEDV流体接种，并且在36±3℃下维持48小时。一个VERO细胞烧瓶保持未接种。对于阳性病毒对照，用阳性对照PEDV接种VERO细胞的一个培养物。在温育期结束时，检查细胞单层的PEDV典型的CPE。将材料冷冻和解冻三次，然后将2ml每种材料接种到一天大的VERO细胞上。培养物应该在37±2℃下维持48小时。在第二次传代之后，进行第三次传代。在温育和传代后，如通过免疫荧光染色的缺乏确定的，在BEI处理的病毒液中不存在病毒感染的细胞构成令人满意的灭活测试。用阳性对照病毒接种的对照细胞应该显示PEDV典型的CPE，并且未接种的烧瓶不应显示PEDV CPE的证据。
PDCoV病毒收获物在37℃下用5mM BEI灭活24小时。灭活后，用硫代硫酸钠中和收获物。通过灭活收获物的TCID50评估来确认灭活。在灭活材料中未检测到活病毒。使用10kd中空纤维PES药液筒(GC Healthcare目录#UFP-10-C-3MA)浓缩灭活的中和的病毒收获物。用改良的最低必需培养基无菌稀释浓缩的抗原(最终浓度＝10X)。将一部分稀释的抗原与EMULSIGEN (MVP Laboratories，项目#10005408)组合，以获得12.5％的制剂。
实施例5：
编码且表达PDCoV刺突抗原的重组杆状病毒的构建
PDCoV S1-IgG2a Fc杆状病毒的制备
使用引物P2967167A(SEQ ID NO：13)和P2967167B(SEQ ID NO：14)，通过PCR从质粒DNA扩增猪丁型冠状病毒(PDCoV)刺突(aa 1-673)的S1区域。使用引物P2967167C(SEQ IDNO：15)和P2968014E(SEQ ID NO：16)，从质粒DNA扩增包含短GGS接头和铰链区的猪IgG2a的Fc区。使用引物P2967167A(SEQ ID NO：13)和P2968014E(SEQ ID NO：16)，通过重叠-延伸PCR融合两个片段，以生成含有起始密码子紧5'的Kozak共有序列的PDCoV S1-IgG2a Fc编码序列(SEQ ID NO：17)。参见图3.A中的示意图。最终的编码序列侧面为BamHI和NotI限制性位点，以促进克隆到杆状病毒转移质粒pVL1393内。一旦完成，将质粒pVL1393-PDCoV S1-IgG2a Fc与线性化的BaculoGold杆状病毒DNA一起用于转染Sf9昆虫细胞，以产生重组杆状病毒。
PDCoVS BD Baculodisplay杆状病毒的制备
将来自猪丁型冠状病毒(PDCoV)的PDCoV刺突基因克隆到来自质粒DNA的两个重叠片段(N末端和C末端)内。使用引物P2967002C(SEQ ID NO：23)和P2967002D(SEQ ID NO：24)扩增N末端片段，而使用引物P2967002E(SEQ ID NO：25)和P2967002F(SEQ ID NO：26)扩增C末端片段。扩增N末端和C末端片段，以去除天然信号序列和C末端尾部。使用引物P290194A(SEQ ID NO：19)和P2967002D(SEQ ID NO：20)，通过重叠-延伸PCR(OE-PCR)，将N末端片段与苜蓿银纹夜蛾核多角体病毒(AcNPV)gp64信号序列(O2967002A)(SEQ ID NO：21)融合。使用引物P290194B(SEQ ID NO：20)和P2967002E(SEQ ID NO：25)，通过OE-PCR将C末端片段与AcNPV gp64C末端尾部编码序列(O2967002B)(SEQ ID NO：22)融合。使用引物P290194A(SEQID NO：19)和P290194B(SEQ ID NO：20)，通过OE-PCR融合两个所得到的片段，以生成含有起始密码子紧5'的Kozak共有序列的PDCoV刺突BaculoDisplay(PDCoVS BD)编码序列(SEQ IDNO：27)。参见图3B中的示意图。最终的编码序列侧面为BamHI和NotI限制性位点，以促进克隆到杆状病毒转移质粒pVL1393内。一旦完成，将质粒pVL1393-PDCoVS BD与线性化的BaculoGold杆状病毒DNA一起用于转染Sf9昆虫细胞，以产生重组杆状病毒。
实施例6：
包含PEDV刺突抗原的药物组合物(疫苗)的制备：
对于灭活的PEDV材料，PEDV病毒收获物使用5mM BEI灭活最少24小时，澄清且0.45μm过滤。
中和后，加入各种佐剂，并且生成下述疫苗/药物组合物。
实施例7：
包含PDCoV刺突抗原的药物组合物(疫苗)的制备：
PDCoV抗原(BacluloS-BD-PDCoV和BaculoS-FC-PDCoV)的分批在3L旋转器中生长。对于感染，SF+细胞以大约1.0-2.1的MOI用病毒进行接种。将烧瓶在27℃下温育，伴随设定在100rpm下的搅动。在感染后五天起始收获。在收获时，细胞活力在24％和26％之间，并且活细胞/mL在0.31 x 106和0.36 x 106之间。通过以10,000x g离心10分钟来澄清收获流体且进行0.2μm过滤。澄清的收获流体与D组合，以获得12.5％的制剂。
实施例8：
用灭活的PDCoV和杆状病毒刺突疫苗接种猪和血清学应答的评价
研究目的是评估PDCoV疫苗原型在常规仔猪中的血清学应答。已生成了几种原型疫苗，其中PDCoV的刺突蛋白在各种主链中表达。另外，已生成了全细胞灭活的病毒制剂。
关于获得实验组的解释参见下表3。第1-4组随机化并且保存在同一房间中。另外，在当前研究中包括在大约4周龄时断奶的严格对照动物(第5组)。在D0时，将猪随机分成四组，并且施用2mL剂量的每种原型PDCoV疫苗或安慰剂。在D21时，猪接受每种原型疫苗的第二次或加强施用。在每次疫苗接种的处理施用之前和在D35时，从猪中收集血清和经口流体。测定样品的血清转换证据。每天通过房间记录一般健康观察。在疫苗施用之后，观察注射部位的反应最少三天。
表3：实验组
组 N(猪) 疫苗
1 10 BaculoS-BD-PDCoV
2 11 全细胞PDCoV
3 11 安慰剂
4 10 BaculoS-FC-PDCoV
如通过IFA检测的疫苗接种后的血清学应答
间接免疫荧光测定(IFA)
在1x磷酸盐缓冲盐水(PBS；Gibco c#10010-023)中制备每个样品从1:40到1:320的两倍连续稀释物。将100μl每种稀释的样品加入PDCoV感染的板中，并且在37℃下温育1小时。温育后，去除血清，且用1xPBS洗涤单层细胞两次。然后使用10μl/孔的1:100稀释的FITC-缀合的-山羊抗猪IgG抗体将细胞染色。与未感染的对照孔相比，滴度被报告为显示PDCoV染色的最大血清稀释度。
在D-8、D21和D35时的血清学应答在下表4中按组呈现。表5中呈现了按日和组的最小二乘平均值和检测的频率。在疫苗接种之前，没有动物具有可检测的血清学应答。(也参见图4)。
在信号疫苗接种后，20-73％的动物对用BaculoS-BD原型接种的动物具有应答。到D35时，用全细胞或BaculoS-FC原型接种疫苗的所有动物都具有可检测的应答。用BaculoS-BD原型接种疫苗的动物中仅50％具有到D35时可检测的应答。
在用原型或安慰剂接种的猪的血清学应答之间没有观察到在D-8时的显著差异。到D21和35时，血清学应答在接受原型疫苗的动物中更高(对于所有组在所有天时p<0.002)。在数字上，与BaculoSBD构建体相比，BaculoS-FC和全细胞原型疫苗看来到D35时刺激更高的IgG应答。
表4：如通过IFA测量的按天和组的血清学数据
*指示该值与安慰剂组明显不同(Dunnett方法)
如通过基于S1-IgG2a-Fc的ELISA检测的疫苗接种后的血清学应答
PDCoV IgG ELISA
将纯化的PDCoV-S1-IgG2aFc抗原(BIVI L#3091-141；0.2mg/mL)在碳酸盐缓冲液(BIVI L#3144-180)中1:6666.67稀释。ELISA板(高结合96孔板；Greiner目录#655061)用100μl/孔稀释的抗原包被，并且在4℃下温育过夜。使用自动洗板机，用300μl/孔的TBST(0.05％吐温20)将板洗涤五次。在洗涤步骤后，每孔加入150μl阻断溶液(酪蛋白阻断剂，Thermo目录#37532)，并且使板在15-30℃下温育1小时。在酪蛋白阻断剂中以1:200的稀释度制备初始血清稀释物(阴性对照和测试样品)。温育后，从板中去除酪蛋白阻断剂，并且从1:200到12800制备测试样品的连续两倍稀释物(在酪蛋白阻断剂中制备)。阴性对照的连续两倍稀释物由1:200直到1:204800制备。在板上制备稀释物，伴随100μl/孔的最终体积。将板在37℃下伴随振荡(100rpm)温育1小时。温育后，如先前所述洗涤板。制备HRP缀合的山羊抗猪IgG(H+L)(Jackson ImmunoResearch目录#114-035-003)的1:2000稀释物(在酪蛋白阻断剂中)，并且向每个孔中加入100μl。使板在37℃下伴随以100rpm的振荡温育1小时，然后如先前所述洗涤。为了检测，每孔加入100μl/孔的制备的TMB底物(KPL目录#50-76-00)。将板在15-30℃下温育10分钟。用100μl/孔的1N HCl终止反应，并且立即用自动读板仪在450nm处读数。
通过下述计算生成结果。首先，计算来自最终五个阴性血清稀释孔的平均和标准差光密度(OD)读数。然后将截断确定为平均值加上三个标准差。将每个样品光密度读数(在其初始稀释度下)除以该板的本底截断。数据报告为信号:本底比。
在D-8、D21和D35时的血清学应答按组呈现。表5中呈现了按日和组的最小二乘平均值和检测的频率(也参见图5)。在疫苗接种之前，没有动物具有可检测的血清学应答。
第二次疫苗接种后，40-45％的动物都具有应答，与组无关。到D35时，用全细胞或BaculoS-FC原型接种疫苗的所有动物都具有可检测的应答。在用原型或安慰剂接种的猪的血清学应答之间没有观察到在D8或D21时的显著差异。到D35时，血清学应答在接受原型疫苗的动物中更高(对于第1、2和4组，分别为p＝0.0039、<0.0001或<0.0001)。与IFA结果相似，与BaculoS-BD构建体相比，BaculoS-FC和全细胞原型疫苗到D35时刺激更高的IgG应答。
表5：如通过IgG ELISA测量的按天和组的血清学数据
*指示该值与安慰剂组明显不同(Dunnett方法)
实施例9：
PEDV杆状病毒疫苗的功效
下述研究评估了对用两个2-mL剂量的杀死的猪流行性腹泻病毒(PEDV)疫苗或杆状病毒构建体疫苗的疫苗接种的血清学应答，如在给以三周龄的猪施用任一疫苗后测量的。主要结果是对于在处理的猪中的疫苗接种后收集的血清样品通过荧光聚焦中和(FFN)测试的血清学。
研究组包括：T01＝PBS(n＝10)；T02＝6.93 log TCID50/ml BEI PEDV+20％(n＝20)；T03＝具有PEDV刺突Ag的杆状病毒(n＝9)；具有PEDV刺突Ag的6x浓缩杆状病毒(n＝10)；具有PEDV刺突Ag的胰蛋白酶杆状病毒(n＝10)；以及有条件批准的杀死的阳性对照疫苗(POS CON)(n＝10)。在D0时，猪在右颈部中肌内施用2-mL治疗。第二次治疗对于T01-T05在左颈部中在D14时施用，并且对于T06在D21时施用。
PEDV2a-BD的制备
在该研究中，将来自猪流行性腹泻病毒(PEDV)2a的刺突基因克隆到来自质粒DNA的两个重叠片段(N末端和C末端)内。使用引物P1360110A(SEQ ID NO：37和PEDV-S2-R(SEQID NO：38)扩增N末端片段，而使用引物PEDV-S1-F(SEQ ID NO：39)和P1360110B(SEQ IDNO：40)扩增C末端片段。扩增N末端和C末端片段，以去除天然信号序列和C末端尾部。使用引物P290194A(SEQ ID NO：45)和PEDV-S2-R(SEQ ID NO：38)，通过重叠-延伸PCR(OE-PCR)，将N末端片段与苜蓿银纹夜蛾核多角体病毒(AcNPV)gp64信号序列(O1360110C)(SEQ ID NO:41)融合。使用引物P290194B(SEQ ID NO：43)和PEDV-S1-F(SEQ ID NO：44)，通过OE-PCR将C末端片段与AcNPV gp64C末端尾部编码序列(O1360110D)(SEQ ID NO：42)融合。使用引物P290194A(SEQ ID NO：45)和P290194B(SEQ ID NO：43)，通过OE-PCR融合两个所得到的片段，以生成含有起始密码子紧5'的Kozak共有序列的PEDV刺突BaculoDisplay(PEDVS BD)(SEQ ID NO：46)。最终的编码序列侧面为BamHI和NotI限制性位点，以促进克隆到杆状病毒转移质粒pVL1393内。一旦完成，将质粒pVL1393-PEDVS BD与线性化的BaculoGold杆状病毒DNA一起用于转染Sf9昆虫细胞，以产生重组杆状病毒。参见图6A.PEDV 2a BDBaculodisplay杆状病毒的示意图。
表6：PEDV原型制剂和对照
BEI＝二乙烯亚胺
*PEDV刺突信号序列和C末端尾部替换为杆状病毒gp64等价物。在昆虫细胞中产生重组PEDv刺突-展示杆状病毒。收获受感染的培养物并且通过离心和0.2-μm过滤澄清。将澄清的收获材料用5mM BEI在37℃下灭活72小时，然后通过离心和0.2-μm过滤澄清。
血清学：
疫苗接种后的血清转换(D28和D35)在20％用由20％ BCL佐剂化的PEDv疫苗接种疫苗的猪(T02；表7)和60％用胰蛋白酶生长的PEDV SPIKE-杆状病毒接种疫苗的猪(T05；表7)中发生。对于所有处理组血清阳性的猪≥1:20的几何平均滴度呈现于下表7中。按所有猪的处理组的滴度频率分布呈现于表8中。
表7：对于血清学应答的猪，按组的血清阳性猪的比例和几何平均滴度
组 具有≥1:20应答的猪 几何平均滴度
T01 0/10(0％) 不可用
T02 4/20(20％) 1:30.7
T03 0/9(0％) 不可用
T04 0/10(0％) 不可用
T05 6/10(60％) 1:35.5
T06 7/10(70％) 1:41.9
表8：按组的滴度频率分布
结论：
在实验疫苗的两次施用后，在20％的T02猪中发生血清转换，所述实验疫苗用由BEI灭活且由20％ BCL佐剂化的6.93log10TCID50/mL PEDV配制。在用PEDV刺突糖蛋白配制的实验性重组胰蛋白酶生长的杆状病毒疫苗的两次施用后，在60％的T05猪中发生血清转换。
实施例10：
PEDV 2a BD杆状病毒疫苗的功效
该研究的主要目的是评估原型PEDV杆状病毒疫苗在常规母猪模型中的功效。将36头母猪随机分成5组。母猪在分娩前5周和2周时用实验性PEDV疫苗、安慰剂、商购可得的阳性对照进行肌内疫苗接种，或未接种疫苗(严格对照)。所使用的组和疫苗的简要描述在下表9中列出。在妊娠期、分娩期和攻击期自始至终，从母猪和猪中收集血清和粪便样品，以监测抗PEDV抗体和PEDV RNA的存在。每天记录一次临床体征。
该研究的主要结果参数是猪死亡率。基于69％的猪死亡率、临床体征的存在以及来自安慰剂组中的猪的粪便材料中PEDV RNA的鉴定，用于本研究的攻击模型被视为有效的。在母猪中使用两个剂量的BaculoS-NT构建体能够显著减少猪的死亡率。相比之下，目前市场上的PEDV疫苗也能够减少猪的死亡率；然而，减少并不显著。由于BaculoS-NT原型不在胰蛋白酶的存在下生长，并且用12.5％D佐剂化，因此预期该原型将是用于生产的优选形式并且具有21天的撤药期。
该研究的次要结果参数是疫苗原型预防猪中的临床体征的能力。由于所有猪中的大于93％对于最少连续两天具有纯液体腹泻，因此疫苗原型或商购可得的产品无一能够预防严重临床体征的发作。类似地，疫苗无一能够减少猪中的临床体征的持续时间。
表9.研究设计：
母猪在分娩前5周和2周时用实验性PEDV疫苗、安慰剂、商购可得的阳性对照进行肌内疫苗接种，或未接种疫苗(严格对照)。在妊娠期间，在每次疫苗接种前收集血液和粪便样品，且分别测定血清转换和脱落的证据。每天对每头母猪记录临床观察。
猪死亡率：
用强毒PEDV分离株攻击后的猪死亡率是用于评价疫苗功效的主要结果参数。在攻击期间按组的死亡率概括呈现于下表10中。按组和天数的生存性呈现于图7中。'按组和天数的存活猪的百分比')。在D0时，使用适当大小的无菌针头和注射器，将2mL剂量的疫苗施用于健康母猪的右颈部的肌肉组织内。在D21时，该过程是相同的，除了在颈部的左侧上给予注射之外。
攻击材料是强毒PEDV原种材料(PEDV 1251-140-4；p5；L#2842-174；滴度＝2.02 x105TCID50/mL)，其在PEDV病毒生长培养基中稀释至1.0 x 103的滴度。(具有HEPES、胰蛋白胨磷酸盐肉汤和酵母提取物的改良的最低必需培养基)。与之前的研究相似，大多数死亡率在暴露于攻击材料的8天内可见。
总之，所有组在感染后48小时内具有存在于大多数猪中的临床体征的相似趋势。临床体征在所有组中超过50％的猪存在直到DPC6。从DPC7到DPC11，临床体征得到消除。到DPC13时，在任何猪中都没有观察到临床体征。
分析：
生物统计学。所有数据都输入版本9.4内用于分析。生成按处理组的数据列表和概括统计学，包括频率分布。适当时进行掺入窝和居住效应的分析。列出了按组和窝的攻击后的猪死亡率，并且描述统计学平均值、中值、最小值和最大值用于按组的概括。使用具有皮尔森卡方作为过分散的广义线性模型将按组和窝的死亡率分析。然后，估计与安慰剂组相比，关于每个处理组的基于模型的预防分数和95％置信区间。具有皮尔森卡方作为过分散的广义线性模型与死亡率数据再一次拟合，加上年龄、处理得分和奇偶性(parity)作为固定效应，以调查这些因素是否对死亡率有任何作用。列出了研究随机化数据。列出了关于猪的粪便得分，并且使用频率分布按组和天数进行概括。概括在攻击之前和攻击期间分别完成。关于母猪的临床体征(粪便、呕吐物、注射部位病变、无乳和处理的)得分使用频率分布按组和天数分别在分娩前和分娩后列出且概括。通过窝、组和天数使用描述统计学中值、最小值和最大值，列出且概括猪重量数据。平均日增重(ADWG)通过混合模型进行分析，其中组和分娩时的重量作为固定效应；窝和用于居住的房间作为随机效应。估计最小二乘平均值及其在组间的对比；估计相应的标准误和95％置信区间。使用按通过死亡率分开的组的每个变量的计数列出且概括关于猪的尸检数据，包括器官的病理学数据以及盲肠的内容物、结肠、小肠和肺部病变，以及胃记录。是否存在任何病变也使用计数和按组的百分比进行概括。首先列出实验室数据，包括关于母猪的PED病毒血清抗体滴度(FFN)、分别关于猪和母猪的血清IgA全细胞ELISA、关于母猪的IgA和IgG baculos ELISA、关于母猪和猪的IgA和IgG IDEXX ELISA和PCR。接下来，使用几何平均值(FFN)(尽管对于FFN，未提供标准差)或平均值(ELISA、PCR)计算描述按组和天数的统计学平均值、最小值、最大值和标准差。接下来，使用具有窝和用于居住的房间作为随机效应的混合模型，来估计按组和天数的最小二乘平均值、标准误和95％置信区间。估计组对之间的最小二乘平均值的对比和相应的95％置信区间。
表10：按组的猪死亡率
该研究的主要结果参数是猪死亡率。基于该参数，在母猪中使用两个剂量的BaculoS-NT构建体能够显著减少猪的死亡率(PF＝0.48)。相比之下，前市场上的PEDV疫苗也能够减少猪的死亡率；然而，减少并不显著(第4组)。由于BaculoS-NT原型不在胰蛋白酶的存在下生长，并且用12.5％D佐剂化，因此预期该原型将是用于生产的优选形式并且具有21天的撤药期。
该研究的次要结果参数是疫苗原型预防猪和母猪中的临床体征的能力。由于所有猪中的大于93％对于最少连续两天具有纯液体腹泻，因此疫苗原型或商购可得的产品无一能够预防严重临床体征的发作。类似地，疫苗无一能够减少猪中的临床体征的持续时间。在临床体征消除后，在DPC14时12-33％的动物中鉴定出PEDV RNA。这提示疫苗接种无法保护不受定殖。在1-3头母猪/组中，至少连续两天观察到严重腹泻。这指示疫苗不能完全保护母猪不受暴露于由受攻击的猪脱落的大量病毒。另外，大多数母猪(14/16)具有在肠系膜淋巴结中存在的可检测量的病毒，指示疫苗接种不能预防定殖。
大多数接种疫苗的母猪在两个剂量的疫苗后具有可检测的IgG(和FFN)应答；与安慰剂疫苗接种的对照相比，滴度更高(数据未显示)。由于IgG ELISA和FFN结果非常相似，很可能FFN测定主要检测IgG。尽管IgG可以中和病毒，但它不太可能是PEDV疫苗功效的唯一决定因素，因为组间的明确差异并不明显。
基于如测量的IgA ELISA(IDEXX)的IgA水平，幼稚和接种疫苗的母猪到DPC13/14时具有相似的血清和乳IgA滴度。然而，在接种疫苗的动物中注意到在DPC3时收集的血清、初乳和乳中增加的IgA滴度。由于来自接种疫苗的母猪的存活猪也具有最高的IgA滴度，该数据支持以前发表的文献，强调IgA在肠道感染中的重要性。(参见图8)。
实施例11：
PEDV2b杆状病毒疫苗的制备
来自猪流行性腹泻病毒(PEDV)2b的刺突基因以两个重叠片段(N末端和C末端)制备。使用引物P290194A(SEQ ID NO：48)和P3183131B(SEQ ID NO：49)，从含有苜蓿银纹夜蛾核多角体病毒(AcNPV)gp64信号序列与PEDV 2b刺突的氨基酸22-400的融合物的合成DNA扩增N末端片段。使用引物P3183131A(SEQ ID NO：50)和P290194B(SEQ ID NO：51)，从已经含有AcNPV gp64C末端尾部编码序列的pVL1393-PEDVS BD质粒DNA扩增C末端片段。使用引物P290194A(SEQ ID NO：48)和P290194B(SEQ ID NO：51)，通过OE-PCR融合两个所得到的片段，以生成含有起始密码子紧5'的Kozak共有序列的PEDV 2b刺突BaculoDisplay(PEDVS 2bBD)编码序列(SEQ ID NO：52)。最终的编码序列侧面为BamHI和NotI限制性位点，以促进克隆到杆状病毒转移质粒pVL1393内。一旦完成，将质粒pVL1393-PEDVS 2b BD与线性化的BaculoGold杆状病毒DNA一起用于转染Sf9昆虫细胞，以产生重组杆状病毒。参见图6B.PEDV2a BD Baculodisplay杆状病毒-蛋白质图的示意图。
实施例12：PEDV和PDCoV功效研究
设计以下研究以评价杀死的PEDV疫苗和/或杀死的PDCoV疫苗或其它原型疫苗在猪中的功效。主要结果参数是用猪流行性腹泻病毒(PEDV)和/或PDCoV攻击后的仔猪死亡率。次要结果参数是母畜(dam)血清学。测量的其它参数包括：疫苗接种后母猪中的临床体征(包括ISL)；疫苗接种后母猪中的病毒脱落(经由qRT-PCR)；仔猪中的临床体征；以及仔猪中的PEDV和/或PDCoV血清学
PEDV研究：
在分娩前4周和2周(D0和D14)时，通过三种途径(肌内、鼻内和经口)给每只妊娠母畜施用2mL下述处理之一：T01(阴性对照，NC)磷酸盐缓冲液盐水；T02(BEI-VH)用20％ BCL佐剂化的；T03(严格对照，SC)充当未接种疫苗/未攻击的对照。每组使用8只动物，排除具有4只动物的T06。在D35或D36时，用1mL 2.0log10TCID50/mL PEDV病毒收获物经口攻击猪。在攻击期过程中每天观察母畜和猪中的临床体征(呕吐和腹泻)。在分娩前四周和两周(D0和D14)时、在仔猪攻击前一天(D34或D35)和脱离测试当日(D57)，从母畜中收集血清。
在D0和D14时，将PEDV原型免疫原性组合物施用于母猪。在每次疫苗接种时，母猪接受总共6mL材料，其中2mL通过肌内、鼻内和经口途径施用。对于肌内施用途径，将2mL注射剂给予在耳下的颈部肌肉组织内。用于施用的颈部侧面交替用于初始和加强疫苗接种。对于经口施用途径，使用附接到注射器的8Fr聚丙烯导管(2.7mm直径×254mm长度)，在尾部口咽上递送2ml。对于鼻内途径，使用附接到注射器的4.5英寸导管，将1ml注射到每个鼻孔内。
表11：PEDV实验性免疫原性组合物和对照产品
表12：PEDV攻击材料
疫苗功效：
猪死亡率：用强毒PEDV分离株攻击后的猪死亡率是用于评价疫苗功效的主要结果参数。下文列出了在攻击期过程中按组的死亡率概括。在T01(NC)中具有55％的死亡率和所有窝都受影响，该攻击被视为足够强毒的。与T01(NC)相比，T02(BEI-VH)证实猪死亡率中的数目减少，具有0.20(-0.550，0.586)的PF(95％CI)。由于95％CI(-0.550，0.586)包括零，因此减少并非统计学显著的。(参见表13)。
在该研究中，二项式外变化是明显的，当利用潜在的二项式分布时，导致关于T02(BEI-VH)PF的宽置信区间。死亡率在一组内的窝中变化很大，包括对于T02(BEI-VH)从0％到100％。
在尸检时获取肠样品或肠内容物，并且通过qRT-PCR测试以检测PEDV抗原。在峰值死亡率期间从动物测试的样品中，在55.5％的样品中检测到PEDV。
表13：
*基于受影响的T01(NC)比例。
**NC＝未计算的。基于研究设计，关于T02(BEI-VH)可能的置信区间
母猪血清学：
荧光聚焦中和(FFN)测定：FFN测定用于评价在疫苗接种和攻击后的母畜病毒中和应答。对于在其下从母猪收集血液的天数，按组列出的几何平均滴度呈现于下文。
在两个剂量的疫苗后，T02(BEI-VH)中的2/8(25％)母猪具有可检测水平的中和抗体。在任何其它组中未观察到可检测水平的中和抗体。
在横向暴露于PEDV后，暴露的处理组中的所有母猪都具有可检测水平的中和抗体。在测试自始至终，T03(SC)组中的动物保持血清反应阴性。在D57时的几何平均滴度(暴露后大约21天)指示，与T01(NC)相比，疫苗接种导致数目更高的滴度。T02(BEI-VH)组中的母猪具有613的GMT，与T01(NC)中母猪200的GMT相比，其是高大约3倍的滴度(p＝0.005)。由于T02(BEI-VH)组中的多个样品在测试的最高稀释度(1:640)下具有可检测的中和抗体，这些结果可能代表组间差异的保守估计。
表14：母猪血清学
当所有值都<20时，几何平均滴度呈现为<20。否则，对于GMT计算，<20的值被设为10
**关于T01(NC)和T02(BEI-VH)的D57GMT是反向变换的最小二乘平均值
基于S1的ELISA数据：
基于S1的ELISA用于评价母畜在接种疫苗和攻击后对PEDV刺突蛋白的应答。对于在其下收集样品的天数，关于初乳、乳和血清的测定结果按组列出。
在猪攻击时，与T01(NC)中的母猪相比，T02(BEI-VH)中的母猪在血清中具有显著更高的几何平均滴度(p＝0.0005)。在暴露于PEDV后，注意到两组之间更大的显著差异(p<0.0001)。
在T02(BEI-VH)和T04(NC)之间未观察到初乳和乳中的抗PEDV IgA的几何平均滴度中的显著差异。
表15：
*关于T01(NC)和T02(BEI-VH)的GMT是反向变换的最小二乘平均值
猪血清学：
在尸检时从猪中收集血清，以评估中和抗体的存在。下表16呈现了按组的阳性猪的几何平均FFN滴度。表16还包括表示为GMT大于或等于20的猪的数目超过测试动物的数目的检测频率。对于所有可用样品进行测试。由于死亡和尸检之间的时间差异，无法获得来自众多猪的样品。
下文列出了按死亡率状态(死亡：是/否)和组(总体)的FFN滴度的描述统计学。总的来说，不管尸检的时间，接种疫苗的组中相似比例的猪发生血清转换(或具有母体抗体)。然而，在T01(NC)中，与对于研究的持续时间存活的猪(43％)相比，在脱离测试前死亡的更高百分比的猪具有滴度(88％)。
当按组察看猪总体滴度时，死亡率估计值比例与关于T02(BEI-VH)的总体FFN百分比反相关。
表16：
组 猪(死亡＝是) 猪(死亡＝否) 总体 死亡率估计值比例
NC 55(28/32；88％) 33(9/21；43％) 63(37/53；59％) 0.55
BEI-VH 44(23/42；55％) 50(9/16；56％) 64(32/58；55％) 0.44
*GMT(滴度≥20的动物数目/测试的总体猪；百分比)；注意，并非从所有猪中获得血清。
攻击后的临床观察：
猪粪便得分：在下文列出了关于猪中的异常粪便观察持续时间的描述统计学，按组和死亡率状态(死亡：是/否)。总的来说，具有相同死亡率状态的猪中的异常粪便得分的中值持续时间在组中是相似的。在死亡或实施安乐死的动物中，存在异常粪便得分的数目上较短的中值持续时间。这种趋势在T01(NC)猪中最明显，并且可能继发于大多数这些动物在攻击后第一周内死亡的事实。
表17：
猪中的粪便得分的严重性概括于在下文的频率表18中。在所有处理组中，在攻击后的至少一次观察期间，大部分猪(>91％)呈现粪便得分2。
表18：
结论：
与T01(NC)组相比，在T02(BEI-VH)中观察到猪死亡率中的20％减少。在该研究中尝试了三种施用途径。尽管使用了3种途径，但不预期除IM外的途径促成基于佐剂和疫苗制剂的T02(BEI-VH)的功效。总的来说，具有6.04log TCID50/ml的最小灭活前滴度、用20％佐剂化的灭活PEDV疫苗，看来在仔猪和母猪中诱导更好的免疫应答。优选的疫苗接种方案是对于3周龄或更大的仔猪的IM施用途径，以2周的间隔的3个2ml剂量。在T02(BEI-VH)中未观察到疫苗接种后母猪中的临床体征，并且在其它处理组中受到限制。疫苗接种的使用看来不影响出生的猪存活的百分比(数据未显示)。
通过两种分开的测定(聚焦荧光中和、基于S1的ELISA)评估作为次要参数的母畜血清学。两种测定均指示，与T01(NC)相比，疫苗接种和暴露后T02(BEI-VH)中的滴度的显著增加。由于FFN测定的已知局限性，还通过基于S1的ELISA测试样品。选择该ELISA是因为刺突蛋白的S1结构域预期含有中和表位。
在横向暴露于PEDV后，暴露的处理组中的所有动物都具有可检测水平的中和抗体。与T01(NC)动物相比，T02(BEI-VH)中的母猪具有高大约三倍的滴度。这是疫苗的使用刺激了初始的初级应答并且在暴露于攻击病毒后导致更大的二次应答的证据。由于T02(BEI-VH)组中的多个样品在测试的最高稀释度(1:640)下具有可检测的中和抗体，这些结果可能代表组间差异的保守估计。
表19：猪死亡率和母猪血清学数据在下文概括。
PDCoV研究
该研究的目的是确定原型PDCoV疫苗在宿主动物中的功效。基于开发PDCoV的攻击材料和疾病模型的以下初步研究，未接种疫苗的仔猪在攻击后一周内显示出重量减轻和腹泻并且恢复。没有观察到死亡率。
表20：猪丁型冠状病毒(PDCoV)攻击材料：
为了建立关于猪丁型冠状病毒(PDCoV)的攻击毒株，在BI-ST细胞上生长两个内部攻击分离株(第3和4组)，并且与NVSL PDCoV分离株(Illinois 2014)进行比较。基于它们所具有的突变及其与NVSL(Illinois 2014)分离株的遗传距离，选择第3组和第4组中的分离株。与NSVL PDCoV分离株相比，第3组病毒(BI PDCoV-5.0327)具有在刺突糖蛋白中的3个氨基酸缺失，以及在ORF1AB基因中的两个变化。与NVSL PDCoV相比，第4组病毒(BI PDCoV2.0307)具有在刺突糖蛋白中的4个氨基酸差异，以及在ORF1AB基因中的3个差异。
攻击的仔猪就腹泻进行得分，其中0＝正常，1＝异常(非纯液体)或2＝攻击后的严重腹泻(纯液体粪便)。在阴性对照组中未观察到腹泻。另外，受NVSL PDCoV攻击的组在攻击后未显示任何临床体征、体重减轻、呕吐或腹泻。第3组和第4组的仔猪显示出严重的临床体征，其中100％经历严重腹泻(得分为2)，伴有呕吐和体重减轻。另外，第3组和第4组中的母猪患有腹泻(攻击后D6-D8)并且进行治疗。先前公布的研究描述了感染有PDCoV的仔猪中的延长感染(和病毒脱落)。他们的观察结果与我们的结果一致。
NVSL PDCoV和阴性对照组中的动物在攻击后重量增加。相比之下，与对照组相比，用两种攻击分离株BI PDCoV-5.0327(第3组)和BI PDCoV 2.0307(第4组)攻击的动物在相同时期内显示出较不稳健的重量增加。
功效研究：
表21.PDCoV研究设计(疫苗组、剂量和途径)
对于功效研究，在分娩前5周和2周通过适当的途径用如表21中详述的每种疫苗制剂给四组母猪接种疫苗。在每次疫苗接种时在施用治疗之前，从母猪收集血清用于血清转换测定。在每次疫苗接种时从母猪收集粪便样品，并且测定疫苗脱落的证据。
来自免疫母猪的仔猪在分娩时入选试验。使用经口途径，在大约5至7日龄时，用1.0mL攻击材料接种仔猪。
从攻击后天数(DPC)0到DPC14，每天一次监测仔猪和母猪两者的临床体征。攻击后粪便和血清样品取自所有仔猪和母猪。测试粪便样品中PDCoV RNA的存在，并且测试血清中抗PDCoV抗体的存在。在分娩时或接近分娩时从所有母猪中收集初乳。在DPC3时以及在DPC10到DPC14之间的一个点，从所有母猪收集乳。测试初乳和乳中抗PDCoV抗体的存在。在DPC 0、7和21时监测仔猪的体重，而终末血清收集在DPC21时。
根据本公开内容，无需过度实验可以制备且实施本文公开且请求保护的所有组合物和方法。虽然已根据优选实施方案描述了本发明的组合物和方法，但对于本领域技术人员显而易见的是，可以对本文所述的组合物和方法以及方法的步骤或步骤顺序应用变化，而不背离本发明的概念、精神和范围。更具体而言，显而易见的是，化学和生理学两者相关的某些试剂可以取代本文所述的试剂，同时实现相同或相似的结果。对于本领域技术人员显而易见的所有此类相似的取代和修改被视为在如由所附权利要求限定的本发明的精神、范围和概念内。
下述参考文献在某种程度上提供了对本文所述那些的补充性示例性程序或其它细节，具体引入本文作为参考。
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序列表
<110> 勃林格殷格翰动物保健美国有限公司
<120> 猪冠状病毒疫苗
<130> 10-0176-WO-1
<150> 62/452026
<151> 2017-01-30
<160> 53
<170> PatentIn version 3.5
<210> 1
<211> 25505
<212> DNA
<213> Artificial Sequence
<220>
<223> Porcine Deltacorona Virus Genome cDNA - NSVL
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tgacggtcaa aattggcccg tctaaacctg gggatgtcat cgtggctact gatgaggaat 1740
accttaaatg ctttgaaacc cctgaggtag gtaggctcta taaggttttc caaactcaat 1800
cttgggctat cattgagcgt gccttctcca gtttgaagat ccgcgtgtcc aaagctttat 1860
cagcatttat aagttttctg caaaaccttg cagataactt tactgcaata agtggtgttg 1920
tcactgcact cattcgtgaa ctccaggatc ttaccctgga tgtggcgaca cgtatcacta 1980
acatacaatt tgtttaccgt gccggtaagc ttattgtcga cacgacaagt gtcatagcta 2040
aacttttcca gccattttgt gattttatat cacctttcct tcggaaagtt gctggttttg 2100
caatttacac tgttggtaat cgcatgctta tgtttaccag cactggcacc tttcttctca 2160
caaaggcaac tactaagata ctcaataagg caaagtacat ctttgatgtg gagcctgagt 2220
acccagtaga tgtaacaaca tccaaagttg tagtacatga agcactccag caaaccgaca 2280
ctaagcctac tagagctctg gaggctgttg atgtcgttgt tggtaatact gtactgcaaa 2340
tggctactga tggcactgcg ttctacccat cggatggtac gcacgcctct cttccaggat 2400
tcaaagcagg ttcggatgag cttttcataa gcttcaactg cgacctcttt gatgatgaga 2460
ctaatgctca aatcaacgaa atactcgctg catatgagct taaccaacta gtggctccag 2520
gtgattctac accgcgtcaa attgcgacgt tggttgtcga tacacttgca gatgctataa 2580
cagaccactt tccggagaaa accattgatc tacctgaaga ctatcaagtc ttttctgacc 2640
atgatgacct cccactcgca caataccaca tccctgatca cctgagcctg tatattcagg 2700
ctatggaagg tgaagatgat agtggtgatg aaatatgtat tgaggacgat gattacgact 2760
gtcctcaagc cgacgaagac acagaaggag taattcccca acagtgggaa cttcctgatg 2820
ttgataaatt tttactcaag atccaggaac ggaagaccag cagcgacgaa gtacttagcg 2880
tcgacgtcta tcctaaacca gagccggtcg gcaatgttgg gattgacgac agcgcgtcgg 2940
aaaagaagcc aaatggggac tcagtaccgg atcctgaggt ccatccaaca ctagagagtg 3000
tggatgttga acgaccaacc gaaacagcaa accaggctgt tgaagacaaa ccttctgata 3060
ccacctttgt ggttgatgag gaacaattac aagaatcaac accagaacat gaactccgct 3120
cctatgaagg ggagtttgat tctgatgatg aaattattat tcctatagta ccagtaacac 3180
ctgcggattt aaaaccacag actattacta taaaggagta ctttaagtct gaaaaacttg 3240
agactattaa cgaaggatcc acagagtcag ttacacaatc tgacgattcg tttgacgagt 3300
catttgttga tgctgagtct gatgatccac aagatcctgc tgtatatgat gatacaacaa 3360
ttataacgga cagcactgat gtaggcgatg agcctgagac aactctagct accatcgtta 3420
acacacctct gacactcgat aataacttgc cacctgaagc cattaaacaa cccagcccaa 3480
ctaaggttga gttagttgtt ggtgaattgg cgagtattaa atttgacaat tctgttctag 3540
tcaaccctgc taatgcgcaa ttaacaaatg gcggtggagc tgctcgtgca attgcaaaat 3600
tagctggtcc aaaatatcaa gagtactgta atagtgtggc tcctatctca ggaccgctta 3660
ccacggactc ttttgatgcc aagaaatttg gtgtagcctg catcttgcat gtagtgccac 3720
ccaaaggttc tgaccctaat gtacaagaac tcctgtatca agcttacaag agtatcctta 3780
ctgaaccagc acactatgtt atacctatac taggtgctgg tatctttgga tgcaacccag 3840
tccactctct ggatgcgttc aggaaagcat gtccaagtga cataggtcgt gtcacccttg 3900
tcactatgaa caaaaaccat ttgcaggtgt gggatgctct caataggacc attgtacgca 3960
ccactactga ctatgatcaa gttaccacca aggcccttac accccaggga gtgttagaag 4020
ccaatctctt tgatggtgag gactttgttc aagaaccaaa acccggtcaa atctaccttg 4080
aggttactga agaagttcag aaccaagcca aggaacttga ccttaacctt cagcaatact 4140
gcgtctacct gaagacttgc caccataaat gggttgtgag tcgtacgaac gggttgatgc 4200
atctaaaaca aaaagataac aattgttttg ttagtgcagg tgtaaacctg tttcaaaaca 4260
ctgcttatca acttagacct gctattgatg ctctctatag ggagtatctt aatggtaatc 4320
caaatagatt tgttgcttgg atctacgcat ccactaaccg tcgtgttggt gagatgggtt 4380
gtccacagca agttatttct ttgctcgtta gtaactctga cgcagcattt tcagcaacta 4440
cagcctgttg taacacctac tttaaccaca caggtgttat ttcagtagct cgtgaatatg 4500
acccaataca accaaaggtc tactgcatga agtgtgatgt gtggactccc tttacacccc 4560
agagtggaaa aggtgcagtt gcaattggta tttctgcaga tgaacctacc ggtcctgcca 4620
ttaaatttgc cgcagctcac tgctggtaca ctaatggcaa gaaaacagtt aatggctatg 4680
acactaaagc taatgttgta gctacctatc ataggtttga cgtgcctaag cctcaacttg 4740
tcgaggacgt ggttgcgctg cctactaaaa atgactttga agttctcaat gttgaagaac 4800
tgccgcagga tagtgtgctc catttggacc cacctcctgt acaggcctta caacctaagg 4860
ctaaccaaca cattgagatt ctagaaaacc cagattatct ggacattttg gatctttgga 4920
ttcgtaaacc caaattcatc ctcgtaaagt cgtggagtgt tttgggtaga gcactatgta 4980
aggcaggtaa agttgtcttt gtcaatgctt cgcttttgac ccgtttctac aattaccttg 5040
tagagattgg tgctcttgac tcaacaataa ggttgtcagt cgatcttacc tgtaaatttg 5100
ttagaacggt tctcccatcg tctaacactg tacacaaaac ttgtcttggt ctgtattatt 5160
cagcccagac actttttgtt tctttagcac cattccttat gttaccagct gtagttagtc 5220
tgcttaattc aggctataca attggcacat atttgtatgc aaaaactggc tggccttgta 5280
attacaatgc cacgcaacac tttgattata attcttactg tgcaggtgac ttggtttgtc 5340
aagcctgttt tgacggtcaa gactccctac atttgtatcc gcatttacgt gttaatcagc 5400
agccccttca gaccactgac tacactgttt atgcgctttc actaatacta ctattagcta 5460
acatgactct tgtcatgggc acgctaatag ttactttctt tgtgaacttc tatggtgtgc 5520
aaataccatt ttatggtaca cttttgatag attatcaatc cgcactggtg attactttct 5580
cagtgtacta cttttataag gtaatgaagt ttttccgcca tctcacacat ggatgtaaaa 5640
ttccaacgtg tgtggtatgt gccaaacttc gtaccccacc tactataaca gttgagactg 5700
tcgttcaggg caggaaatac ccatctgtta ttgaaacaaa tggcgggttt acaatttgta 5760
aagaacacaa cttctattgc aaggactgct ctttacaaac acccggcact ttcatcccga 5820
cagaagctat tgagtcgctc tcacgagcta ccaggcttag tgtcaaacca acagcaccag 5880
cattcttact tgctagagat gttgagtgcc aaactgatgt tgtcgttgct cgcgcaatgc 5940
ataaccaaaa tgcgcatgtg tgcatttcaa aatactctga tatccgtacc gttgaccaac 6000
tacttaagcc tactccactg ttttcataca ctcccgatgt tatcatcgcg gcagactttg 6060
acaacagagg tagtcttaag acagctaaag aattagctgt ggttttgtca atggacctta 6120
aacgtactat aattatcatt gatcaggcct attctagacc tattgataat tatcaggaag 6180
ttgcttctcg tattgagaag tattacccag ttgcaaagat cacacccaca ggtgacatct 6240
ttacagacat taagcaagcg accaatggcc aagctagtga ctctgctatt aatgcagctg 6300
ttctggctgt ccagcgcggt cttgatttta caattgacaa ccctaacaac atattaccac 6360
attacgcctt tgacttttca accctcaatg cagaagacca gtctaccatt ttggagagtg 6420
gttgtgctaa aggcaatctc aagggcacta atgttggtgt tgttctttca gctagccttg 6480
ttacacgtct tagtcagcag gctatacgtg tgattgctaa tgctgcttca cgtaatggtg 6540
ttacatgcgc tgttactcct tctacacttg ttatgcgtgg gaatattgca acacagccct 6600
tgactcgcat caaagctggt gcacctccca tgcgtcaaaa aattttatgt gttatcctgg 6660
cacttgctat tgtgtacttt gctgctatgg cttttggctt tttggcaagt caacttacgc 6720
ttaatacagt gcctacgatt aaatctgata tccgcgcctc taccttctac gttgttagag 6780
atggagtctt ggatactgtt cgttcaaatg acaagtgctt tgcaaataag tttttggcat 6840
ttgatagctt cattcaagca ccttacacta attcacctga ctgtccagtt gttgtgggag 6900
ttgttgatgt aacgacgcac tctattcctg gaattccagc aggtgtcatt catagagacg 6960
gtctcatact taacatttat gaacagtctc tttatgaaac tcatcagcgt cagtctatgg 7020
ttagggatgc gttgtcactc aagacagcaa atctctttaa cctaggcaag cgtgttgtag 7080
taggatacac tcaacatgaa gttgttgtgg gtacctccta ttttaattct cctgcacttt 7140
ttaatgcaaa gtgcaccttc ttacagtatc aggacactag acaactctat tgctatgata 7200
ctgttcctac tgaacataag ctctactctg atgtgcttcc gcacgtcgag tataaggcta 7260
ttgacattaa tggtgatctt gttcctttca agataccaga gcagataatg ttctatccac 7320
atattgtgcg ctatactagc aattcctatt gccgtatggg gcattgtttt aatactaacc 7380
ctggtatttg catttcattt acggacgaat ttccgtatag tgaaaatgtc aaacctggtg 7440
tgtactgtgc tgatacctct ttgcagttgt tttcaaacct cgttttgggc actgtatctg 7500
gtattcacat ctttacatca acagctgcat tgcttggatc tactattgtg atcatactat 7560
gcgttgttgc tgttcttgca gttcagcgat tcttcaagga gtacacaact tttgttatgt 7620
acacttgtgg tcttgctctt gtcaacattg taggcattgc acttatgtac aagtgccttg 7680
tcttcgcgat tttctattat gcaatctacc tttactttgt ccttactttc ccctccttta 7740
agaggaatgt ggcattgttt tacttcgctg tagtgatcgt gccgcacgtg agtaacatgc 7800
aattgcttgc gctcattgtg tgtagcatta tctactttct ctacacctat gttcatactg 7860
tagctaagac agctgggaaa ttttcttcct tcttagacgc agctaaagct acttttgtca 7920
ttgacaatga aaagtacgtg ttgcttaaag acctcgctgg tgctgaattt gaccagtatc 7980
tggcctctta caacaagtac aaatattttt ctggtactgc ttctgataag gattatgata 8040
aggtctgtat ggcatttctt gccaaggctt tgtcatcttt tcgtgaagga ggcggttcac 8100
agttgtacac accacctaaa tttgcagttg ttcagagtct taagaccaag ctgcaagcag 8160
gtatcaaaat cctcctgcac ccttcaggtg tagttgagcg atgtatggtc tcagttgtct 8220
acaatggatc tgcattgaat ggcatctggc ttaagaatgt tgtctactgc ccacgccatg 8280
taattggaaa attccgtggt gaccagtgga ctcacatggt ctcaattgct gattgccgcg 8340
actttatagt caagtgtcca atacagggta ttcagctaaa tgtccaatca gttaagatgg 8400
taggagctct cctccagtta actgttcata ccaacaacac agccactcca gactataagt 8460
ttgaaaggct acaaccagga tcatcgatga caattgcttg tgcttatgat ggcattgtac 8520
ggcatgtcta tcacgtggtc ctccaactta ataatcttat ttatgcaagc ttccttaacg 8580
gagcttgtgg tagtgtgggt tacactctta agggtaaaac actctactta cattacatgc 8640
accacattga gtttaacaac aaaactcata gtggtacaga tcttgaaggt aacttctatg 8700
gcccctatgt ggatgaggaa gttattcagc aacaaacagc attccagtat tacactgata 8760
atgttgttgc tcaattatat gcacacttac tgactgttga tgctagacca aaatggctgg 8820
cacaatctca gataagtatc gaggatttca actcatgggc tgctaacaat tcctttgcta 8880
acttcccatg tgaacaaact aatatgtcct acattatggg actctcgcaa acagctcgag 8940
tccctgtaga acgtatcctc aataccatta tacagctaac caccaataga gatggtgctt 9000
gtattatggg atcttatgat ttcgagtgcg attggacgcc agagatggta tacaatcagg 9060
ctccaatttc attgcagtca ggagtagtta agaaaacttg tacgtggttc ttccacttct 9120
tgtttatggc tattaccatg ctactcgctg ccatgcatgt tttccctgta cacttgtatc 9180
caatagtact gccatgcttc actgtcgtgg cattcctgtt gactttaacc attaaacaca 9240
ctgttgtgtt taccactaca tacttgcttc cgtcactttt gatgatggtt gtaaatgcta 9300
acactttttg gataccgaac acatttctgc gcacctgcta cgaaactata ttcggttccc 9360
caattgctca gcgactgtat ggttacactg ttgctcttta tatgctgatc tatgctggac 9420
ttgcaatcaa ctatacgttg aaaacactcc ggtatagagc aacttcattc ttatcttttt 9480
gcatgcagtg gtttcaatat ggttatgttg cacacattgc gtacaaactg cttaataaac 9540
cctggacaga atcactactc ttcacagcct tcacaatgct aaccagtcat cccttgttgg 9600
ctgctcttag ctggtggcta gctggtcgcg taactctgcc cattatcatg cctgacttag 9660
ctattcgtgt tttggcgtat aacgtcattg gctatgtcat atgtgttcga tttggcctta 9720
tgtggcttgc aaatcggttc acaactgtac ctatgggcac ataccagtat atggtgtctg 9780
tagagcaact taagtacatg atggcagtta agatgtcccc accgcgtaat gcgtttgagg 9840
tgcttatagc caacattaga cttcttggtt tgggtggaaa ccgtaacatt gctgtttcta 9900
ctgtccaaaa caaaattctt gatgcaaaag ctactgctgt tgttgttgct aaccttcttg 9960
aaaaggctgg cgtcacaaac aagcacgcta tttgcaaaaa gattgtgaaa ctccacaatg 10020
atacccttaa agccaccact tatgaggagg ttgaggtagc acttgtgaaa cttctttctc 10080
acataattga gttcttgcca actgatcagg tagatgctta tctagctgat gcggccaatg 10140
ctcaacatgt taatacctat ttagacaact tgcttgagaa caaagctgtt gttcaggctg 10200
ttgccgatat caacattaat ctggattctt atagaattta taaggaggca gatgctattt 10260
ataaacgatc tgttgagatg aacgaatctc cgcaggagca aaagaaaaag cttaaagctg 10320
ttaacattgc aaaggcggaa tgggagcgtg aggctgcttc tcagcgtaag cttgaaaagc 10380
ttgctgatgc tgctatgaag tctatgtatc ttgcagaacg tgctgaggat cgtcgcatta 10440
agctaacctc tggacttact gcaatgcttt accatatgct tagacgtctt gactcagata 10500
gggtaaaagc tctgtttgag tgcgctaagg cacaaatctt gccaatacat gctgtagtcg 10560
gaatttctaa tgacaacctt aaagttattt ttaacgataa ggacagctac tctcattatg 10620
tagagggcaa cacacttata cataagggag ttcgctacac tattgtgaag aaactctcct 10680
tagataatgc acctattgaa ggcgtaccag aagaattccc tgtggtcgtt gagactgtta 10740
gggaaggtgt gccccagttg caaaataatg agctatgttt gcgcaatgtt ttcactgctc 10800
agaacacagc tcaggacttc aatggcaatg aatccactgt aaaatctttt tatgttacta 10860
gaaccggtaa gaagattttg gttgccatta catcaactaa agacaatctt aagactgtga 10920
cctgccttac tgagaccggt aagacagtcc ttaacttgga cccccctatg cgcttcacac 10980
ataccgtagg tggaaaacag tctgttgtct atctctattt tattcagaat attagttcac 11040
tcaacagagg tatggttatt ggccacatct ctgaaactac tatccttcag gcaagtggca 11100
ctcaaattga gtaccagcaa aatgcctctc ttttgaccta tttggctttc gctgtagacc 11160
ctaagacagc ctaccttaag catcttgctg atggtgggtc tcctatacag ggttgtattc 11220
agatgattgc tactatgggt cctggatttg cagttactac taaaccacaa cctaatgagc 11280
atcagtattc ttatggtggt gcttcaattt gtctttattg ccgtgctcat ataccacatc 11340
ctggtgttga tggacggtgc ccctacaaag gccgctttgt tcacatcgac aaagataagg 11400
aacctgtttc cttcgctttg actcatgagc catgcagttc ttgtcaacgg tgggttaatt 11460
atgactgcac ctgcggatct agtctgcaga attcggctta tttaaacgcg taacgggttc 11520
tagtgacgcc cggctagaac ccctgcagcc tggaactcaa ccagatgctg taaaaagggc 11580
cttccatgtg cataatgata ccacctctgg tatattctta agcacaaaat ctaactgcgc 11640
tcggtttaaa accacacgca gtgccctgcc tttacctaat aagggagagg ttgaattgta 11700
ctttgttact aagcagtgtg cagctaaagt cttcgaaatc gaggaggaat gctacaacgc 11760
tcttagtaca gagctttata ctactgatga tacatttggt gtccttgcca aaactgagtt 11820
ctttaagttt gacaagatac ctaatgtcaa tcgccagtat ctgactaaat atacactcct 11880
ggacttggct tatgctctac gtcatttgtc aacatctaag gatgttattc aagaaatctt 11940
gatcaccatg tgcggaaccc ctgaagattg gtttggggaa aattggtttg atccaattga 12000
gaacccatcc ttttacaagg agttccataa acttggggat attcttaacc gttgtgttct 12060
taatgccaat aagtttgcta gtgcctgtat agacgctggt cttgttggca tattaacacc 12120
cgacaaccaa gacctcctgg gtcagatcta tgactttgga gattttatta ttacacaacc 12180
aggtaatgga tgtgtggact tagcatccta ttattcttat ttaatgccca ttatgtccat 12240
gactcacatg ttaaagtgtg agtgtatgga tagtgatggc aacccacttg agtatgatgg 12300
atttcagtat gacttcacgg acttcaagct tggcttgttc gagaagtatt ttaagtactg 12360
ggaccgtcct tatcatccta acactgttga atgtccagat gaccgttgcg tattgcactg 12420
tgcgaacttc aatgtgttgt ttgctatgtg tatacctaat acggcatttg gcaatctttg 12480
ttcaagagct actgttgatg gccaccttgt ggtccagaca gtgggtgtac acttgaaaga 12540
actcggtata gtccttaacc aggacgttac cacacacatg gcaaatatta atctaaacac 12600
tctattgcga ttggttggtg atcccaccac tattgcaagt gtctcagaca agtgtgtaga 12660
tttaagaact ccttgtcaga ccttggctac tatgtctagc ggaattgcta aacagtcagt 12720
caagcccggg cattttaatc aacacttcta caagcatttg cttgatagta acctattaga 12780
ccaacttgga atagacattc gccacttcta ctatatgcag gatggtgaag cggctatcac 12840
agactacagc tactacaggt ataatacccc cacgatggta gatatcaaga tgttcttatt 12900
ttgccttgag gtggcagata agtatcttga gccctacgaa ggtggatgta ttaatgcaca 12960
gtcagttgtg gtctctaatt tggacaagtc agcgggctac ccctttaaca agctaggtaa 13020
ggctcgtaac tattacgaca tgactcatgc cgagcaaaat caactgtttg agtatacaaa 13080
acgcaatgtt ttgcctacac tcactcagat gaaccttaag tatgcaattt cagccaagga 13140
tcgtgctcgc actgtggcag gagtgtctat aattagcacc atgactaaca ggcagtacca 13200
tcaaaagatg ctgaaatcta tttcacttgc acgcaatcag accatcgtga ttggaacaac 13260
caaattctat ggtggttggg acaacatgtt acgacgactg atgtgtaata tcaacaatcc 13320
cattttagtg ggttgggatt accctaagtg tgatcgttct atgccaaaca tgctgcgcat 13380
tgccgcttcg tgcttgctag cacgaaaaca cacttgctgt aaccaaagcc agcgattcta 13440
ccgtttggct aatgaatgtt gccaagtact atctgaagtg gtagtctctg gtaacaacct 13500
ctatgtaaaa ccaggtggca ctagcagtgg tgatgcaacc acagcttatg ccaactcggt 13560
atttaacatc ttacaggtgg tttctgctaa tgtagccacc ttcttatcaa cttccaccac 13620
gacacatctt aataaggaca ttgcggactt gcatcgtagt ctttatgaag atatttatcg 13680
tggtgactct aatgatatca ccgtcatcaa tagattctac cagcatctcc aaagttactt 13740
tggacttatg atattgtctg atgatggtgt cgcatgcata gactcagccg ttgcaaaggc 13800
tggagctgtt gctgatcttg atggtttccg agacattttg ttttaccaaa acaatgttta 13860
catggcagac tcaaagtgtt ggacagaaac tgacatgaat gttggccctc atgaattttg 13920
ctcacagcat actgtgttag cagagcatga tggtaaacct tactacttac cttacccaga 13980
tgtctctcgc attctgggtg catgcatctt tgtggatgac gttaacaagg ctgaccctgt 14040
tcagaacctt gaacgttaca tctcacttgc aattgatgca tatcccctca ccaaggttga 14100
ccctattaag ggtaaagtct tttatttgtt actagactac atacgtgttc ttgytcagga 14160
gttacaggac ggtatccttg atgctttcca atcactcact gacatgtcgt atgtaaataa 14220
ctttatgaat gaggcctttt atgctcagat gtatgagcaa agtcctacac tacaggccag 14280
cggtgtttgt gtggtgtgta attcacccac tatactgcgc tgtggtgatt gcattcgtcg 14340
accactactt tgttgcgtct gtgcctacca gcatgttacg cagactacac ataaacgtat 14400
cattgctatc aacaactaca tctgtagtgt tgagaattgc aatgaggaca atgttgaaaa 14460
acttttcatt tctggcactg cgatttattg tgagaatcac aaacccacgc tgtgcatacc 14520
cattgtagct aatggttctg tttttggtat ctatcgccac actgcccgtg gtagtgatga 14580
catagacctc tttaacgagc ttgctacatc taactatgac actattgaac cttatcagaa 14640
ggccaatcgt gcacctttat cacttatgct cttcgctgct gaaaccatta aggcactcga 14700
ggagtctatc aagaagtcat atgctaccgc aaccgtcaag gatgtgtatg accaacgctt 14760
cattaaactt ctatgggaac agggtaaaaa gccgccaccc ataacgaaga accacatttt 14820
cactggctac cattttaaca agaatggaaa aacccaagtt ggtgattaca ttcttgctaa 14880
aacagatggc agtgacactt atacttacag aggaacatct acctacaaac tccaaacagg 14940
tgatgttcta gtcttaatgg cacatgttgt tacaccgctc tcagcacccc ctgtgttaac 15000
gcagacaaca tatgtcagaa aatcactttt acccgactct gttggtgcgt cttattatgt 15060
gcaacatttt aagtcatata atgagatagc tatgcagagg gttacaacag tattaggtcc 15120
accaggcaca ggtaagtcaa cctttgctat tggtttggct aagtactttc ccagtgcacg 15180
tatttgctac actgcgtctt cgcatgcagc aatcgatgca ctctgtgaaa aagctttcaa 15240
gacaatacct gtaggccaat gcagtcgtat cgtacccaca cgtacaactg ttgagtgctt 15300
tcaggagttt gtcgtaaata acacaactgc acagtatatc ttctcgacta tcaatgcctt 15360
acctgacatt aagtgtgaca ttgtagttgt agatgaggtt tctatgttga ccaattatga 15420
gctttcctct gtgaatgctc gtttggttta caatcacatt gtgtatgttg gtgatcctta 15480
tcagttacct tcacctagaa ctatgcttac gtctggccag ctttcgccag ctgactataa 15540
cgtagttact gatataatgg tacatgcagg agcggatgtt atgctcgaca tgtgctacag 15600
atgcccacgt gaaatcgttg agacagtgtc taaacttgtc tacgataaca aactaaaagc 15660
ggcgaaaccg aactcaagac agtgttacaa gaccattgtg aactttggtc ctggagacgt 15720
tgctcatgag ggacaatctg cctacaacga agcacagttg cgtttcgcac tcgcatttag 15780
acaacaaaag cggtgggata acgtgacttt catatctcca tataatgcta tgaatgtgaa 15840
agcatcctta gcaggtttct ctactcagac cgttgactct tctcaaggtt ctgagtatga 15900
ttatgttatc ttttgcgtga ccactgattc agcacacgca cttaacatgg ctcgtttgaa 15960
cgttgccctt acacgcgcaa agataggtat ccttgtggtg tttaggcagg caaacgaact 16020
ttacaatagt ttgcagtttg aatctattga ttcacagctt cagtcgagtg ctgagaaaaa 16080
cctcacacca ctgtttaagc gctgcggcta tgagtataat ggcgtccatc cagctcatgc 16140
tttgacctgg catgattgtg gtgcagagta ccgctgtgag gagccacttg ctaaattagt 16200
aggagttgcc gatggcactc ttatatcata caaaacccta gtatccacac ttgggtttct 16260
tccatcactt aaaattgatg catatcataa tatgttccta acacgtgacg cgtgtcgcac 16320
ctatgttcag agttggatcg gcatagatgt tgaagcagca cacgccataa aacctaacac 16380
cgggactaac ctgccattgc aaataggttt tagtaccgga aagaattttt cagtcactcc 16440
agagggaatt tgggtaaacg agcacggatc ttgcactgag cccgtccctg ccaaaatacc 16500
tcctggagaa caatttcgtc accttaaaaa ggacatgcgc caggcgcgtc cttggaaggt 16560
tgttcgacgt gagattgcta ctcacattgc tgaggtagct cctcacactg attatatatg 16620
ctttgtcact tgggctcacc agcttgagct agcgacaatg cgctactttg tcaaactagg 16680
tatggaagag aaatgctttt gtggcaggcg ggcttgtttc actaatggaa ctgagttcgc 16740
ttgcaaagca caccattctc tcaccattcc acaatgtgat tatgtgtaca atccattcct 16800
catcgacgtg gctacgtggg gattctcggg acggctttcc accaaccatg acgctgtgtg 16860
cacatatcat gctaatgccc atgttgcatc agctgatgca atcatgacgg tatgtttagc 16920
tatccatgaa ctgttcagta ctgttgactg gaactttgaa tttccagtaa ctgctgagca 16980
atcgcaactt aacaaggcct gtcgcttagt acaggcgaat tacttaaata tactactcac 17040
tacaaccaaa gccacggtgg ttcacgatat tggtaaccca aaaggtatcc ctatcgtgcg 17100
caaacctggt gttaaatatc acttctatga tcaagcaccc attgtcaaac acgttcaaaa 17160
actaaagtac aagccagaga tggaggcccg tttcaccgat ggtttgacta tgttttggaa 17220
ttgtaatgtt gacacatacc ctgctaacgc ccttgtgtgc cgctacgaca ctcatcggca 17280
gaagcattta attggaccta atggttcagc actatatgtt aataagcatg cttttctcac 17340
ccctgagatg catacttatg ctacacataa actcaacttg gctccactca tctactactc 17400
caccacagat tgtagtagtg aacagcctat tgttgttacc tacagagatt gtgtcacccg 17460
gtgcaatact ggaaaaactc tctgtccaaa tcatgctctt gaataccaag agtttatcaa 17520
tgcatacaat ctcatggctc gccatggatt taatgtttac ataccacgca atgtcaacgt 17580
ttacaactgt tggcttactt tcactaatct ccaaaacctt gaaaacttag cttacaactg 17640
ttattataag aactgcaatg ctcacgttga tgggcagctt gatgtagtta ttaataataa 17700
cgctgtatat gctaaggtcg acaataatct tgtcaaactt ttcgacaacc gcactaactt 17760
acctgtctca gtggcctttg aacattacac taacaggcat acccgttcac tgccaactac 17820
acagctgtta tctggtttag gcgtaaccgc caccagaaat ttcactgtgt ggttcgacaa 17880
tgatacaatt ttccaataca ctattaatgt atctacgtat actgacatcg accctagtac 17940
ccatgttgtc ctctgtgatg ataggtacgg aacagattgg agtcagttta accaacttcc 18000
taatgcagta ttcctcacca aaactaaggt gaagaaaaca gaaccgtttg tttgtacagc 18060
actgacccta aatggcgtcg ccattgacgg tgaagagcta tacatctatg tacgctataa 18120
caatcaactg accacatttg ctactacttg tacacagggt agaaatgttg agcagtttat 18180
acctaaaaca cctatggaaa gagacttcct tgagatgtct caacagtcct tcatccagca 18240
acatcaattg caggaactgg gtgttgaaca cattatctat ggtgatgatt ccagtccagt 18300
cattggcgga actcacacac ttatctcact agttaaaaac aagtttgaac atcagcttgt 18360
caaccatgtt tacaacccag tccagaactg tgttgttacc tcacctaacg caagctccaa 18420
gaacgtttgc actgttcttg atgttcttct tgatgactac attgacatca taagacaagc 18480
acatgccagt tacacaagta aatccaaagt attcactgtg tcaattgaca accaacaaat 18540
tagattcatg ctttggcatg atgagcaagt caagacttgc tacccaatct tacagtcact 18600
taccaatggt taccagatgc catctgtgta caaaacattg gttactgact tacaaccagc 18660
tgacatccct aattatcatt cctacacccc ccgggtgcct ggagtagtta agaatgttat 18720
caagtaccgc caacttttca actacatagt taaaaaggat aggttggcag taccacacaa 18780
tatgaccgta ttacaccttg gagctgcatc tgcactaggt acagcaccag gttcttcagt 18840
cataaaacaa atgtttcctg aaggaactgt tcttattgac ctcgatataa gagagttcac 18900
ttcagatgct aaccaaataa tagttacaga ctacagaact tacataccac cacaccacgt 18960
agacgtcata ttttctgacc tctactgttg tgatgacata cacttctttg acaatctaat 19020
aaggatagtt aaggagaggc tcgccctcgg tggttctatc tttgttaaga taactgaaca 19080
ttcattctca cccgaactct actcacttgc gggttggttc gatgattatc aactattttg 19140
cacagcagtc aatgcctcgt cttcagaagc atttttatgc tgttttaatt atttggggct 19200
tgctaaggaa aacattaatg gttttaactt acatgcttcc tatattcaat ggcgcaatga 19260
aatagcgttg acaccaacct attctccttt agcggacaac ccggctacgg cctgtaagct 19320
aaaagcaacg cctattatct cggctcgtga gttagagaag aagcctattc ttcgctatct 19380
cgttgcatca gggcgccttc ttgtgaggcc accagaatgc agagagctct attgattatg 19440
accttatttt gtctcgttcg agcaaagttt gctgatgatc tactcgattt gctcaccttc 19500
ccgggtgcac atcgcttctt acataaaccc acgaggaatt ccagcagtct ctactcgcgg 19560
gctaataata attttgatgt tggcgttctt cctggctacc ccactaagaa cgttaacctc 19620
ttctcaccac ttactaactc cactttgccc attaatggcc ttcatcggag ttaccaacca 19680
ctcatgctga attgtcttac taaaataact aaccacactc tcagcatgta tctcctacct 19740
agtgagatac aaacttatag ctgcggcggt gccatggtta aataccagac acatgatgca 19800
gttcgtatca ttttagacct cactgccact gaccacatct ctgttgaagt cgttggccaa 19860
catggtgaaa attatgtgtt tgtttgcagt gagcagtcta cctacaccac tgcattacac 19920
aaatctacct tcttctcact taattctgag ctttattgct ttactaataa cacctactta 19980
ggtattcttc cacctgattt aactgacttt acggtctacc gtactggtca gttctatgct 20040
aatggttacc ttttaggtac tttacctatt acggttaact atgttaggtt gtatcggggt 20100
catttgtctg ccaatagtgc ccactttgcc cttgcaaacc taaccgatac actcataaca 20160
cttaccaata ctactatatc gcaaatcact tattgtgata agtctgtagt tgattcaata 20220
gcatgccagc gctcttctca cgaagtggag gatgggtttt actccaaccc taaatctgcc 20280
gttagagcta ggcaacgtac tattgttaca ctacctaagc tccctgagct tgaagtagtg 20340
cagttaaata tttctgcaca catggatttt ggcgaagcca gacttgacag cgttaccatc 20400
aatggtaaca catcctattg tgtcactaag ccttacttca ggcttgaaac taactttatg 20460
tgtacaggtt gcactatgaa tctgcgcact gatacctgta gttttgacct gtcagcagta 20520
aacaatggca tgtcattctc tcaattctgt ctaagcactg aatctggtgc ttgtgagatg 20580
aaaattattg ttacctacgt atggaattac ttgctaaggc agcgtttgta tgttactgct 20640
gtagagggcc agactcacac tggaaccact tcagtacatg caacagacac ttctagtgta 20700
atcactgatg tctgcactga ctacactatc tatggagtct ctggtactgg cattattaag 20760
ccatcagatc tcttattgca caatggcata gtattcacct ctccaacagg tgagctttat 20820
gcatttaaaa atataaccac tggcaaaacc cttcaggtct taccgtgtga aaccccttct 20880
caactgattg tgataaacaa caccgttgtc ggtgctatca catccagtaa ttcaactgaa 20940
aataataggt ttactactac tattgtcaca cctactttct tttattccac aaatgccacc 21000
actttcaact gcactaagcc tgttttgtcc tatggaccta tcagcgtgtg tagtgatggt 21060
gcaattgtgg gaacatccac attacagaat actcgaccat ccatagtttc actatacgat 21120
ggcgaagttg aaataccatc tgcattttct ctttccgttc agacggagta cttgcaagtt 21180
caagcagagc aagttatagt tgattgtcct cagtatgtat gcaatggcaa cagccgttgt 21240
ctacaattac tggcacaata cacctcagct tgctctaaga ttgaagcagc tctgcattcc 21300
tctgcacagt tggatagcag agagattata aatatgtttc aaacatcaac acagtccttg 21360
cagttagcta atattaccaa cttcaagggt gactacaatt ttagcagcat actaaccacc 21420
agaattggtg gcagatctgc tattgaagac cttcttttta ataaagttgt tactagtggc 21480
cttggcactg ttgatcagga ctacaaatcc tgctctagag acatggccat cgctgactta 21540
gtttgttccc agtattacaa tggcatcatg gttctacctg gtgttgttga tgctgagaaa 21600
atggcaatgt atactggctc tcttactgga gctatggtat ttggaggact gactgccgca 21660
gcggcaatac catttgccac ggcagtacaa gctcgcctca attatgtcgc actgcaaaca 21720
aatgtactac aagaaaacca gaaaattctt gcagaatcat ttaaccaagc agttggcaat 21780
atatcacttg cactatcttc tgttaatgat gccatccagc aaacttctga ggctcttaac 21840
accgtagcta ttgctattaa aaagattcaa acagttgtta accagcaggg cgaggcatta 21900
tcacacctga ctgcacagct gtcaaacaat tttcaagcaa tttcgacttc tattcaagac 21960
atttacaacc gtcttgagga agtagaggct aaccagcaag ttgaccgtct catcacagga 22020
cggttggctg cacttaatgc atatgttact cagttactca atcagatgtc tcagattaga 22080
caatctcgat tgttagctca gcaaaagatt aatgagtgtg tcaaatcaca gtcgtccaga 22140
tacggtttct gtggaaatgg cacacacatc ttctcactta cacagactgc accaaatggc 22200
atatttttca tgcatgcagt gctagtaccc aacaaattca cacgtgtcaa cgcttctgcc 22260
ggcatttgtg tggataatac gagaggctac tcattgcagc ctcaacttat actctaccag 22320
tttaataact cctggagagt tacacctaga aatatgtatg aacccagact gccccggcag 22380
gctgatttca tacaattaac tgattgcagc gttacttttt acaacaccac cgctgctaat 22440
cttcccaata ttatccctga cattatagat gtcaatcaaa cagtcagtga tattattgac 22500
aatttaccta cagcaacacc tcctcagtgg gatgttggta tctataacaa cactattctc 22560
aacctcaccg ttgagattaa tgatctacaa gagcggtcta aaaacctctc acagattgca 22620
gatcgtttac aaaattatat tgacaatctt aacaatactc tagttgacct tgaatggctc 22680
aacagagtgg aaacttacct taaatggccg tggtatatat ggcttgccat tgccctggct 22740
cttattgcat ttgtgacaat cctcataaca atctttcttt gtactggttg ttgtggtggt 22800
tgctttggtt gttgtggcgg ttgttttggc cttttctcta agaagaaaag gtataccgac 22860
gaccaaccaa caccgtcctt taagtttaag gaatggtagt cgacgactgg gccgttacca 22920
tccctggaca atatattatt gctatactag ttgtcatctg cattggtgtg gcactacttt 22980
ttattaatac ttgcttagct tgtgttaaat tattttacaa gtgctaccta ggggcagcat 23040
atcttgttag gcctattata gtgtactact ccaagccgaa ccccgtacct gaggatgagt 23100
ttgtaaaagt acaccaattt cctagaaaca ctcactatgt ctgacgcaga agagtggcaa 23160
attattgttt tcattgcgat catatgggcg cttggcgtca tcctccaggg aggctatgcc 23220
acgcgtaatc gtgtgatcta tgttattaaa cttattctgc tttggctgct ccaacccttc 23280
accctagtgg tgaccatttg gaccgcagtc gacagatcat ctaagaagga cgcagttttc 23340
attgtgtcca taatttttgc cgtactgacc ttcatatcct gggccaagta ctggtatgac 23400
tcaattcgtt tattaatgaa aaccagatct gcatgggcac tctcacctga gagtagactc 23460
cttgcaggga ttatggatcc aatgggtaca tggaggtgca ttcccattga ccacatggct 23520
ccaattctca caccagtcgt taagcatggc aagctcaagc tacatgggca agagctggcc 23580
aatggcatat cagtcagaaa tccgccacag gatatggtga tagtgtcacc aagtgacacc 23640
tttcactaca cttttaagaa acctgtggaa tcaaacaacg atccagaatt tgctgttctg 23700
atataccagg gtgaccgcgc ttcaaacgct ggacttcaca ccataaccac ttcaaaggcc 23760
ggtgacgctc gcctgtataa gtatatgtaa tgtgcaactg ccatctgcag ctgcgagatt 23820
tatatagatt gtgcaataag cggcacatca gaagagagga tgttcctgag cttattgacc 23880
ctctcgttaa aactcgctgt tttgcttaca gtctcgtggt tcttgctaat gctaatccaa 23940
ttgcatttag catactacct cggaaaattc ttatcaatgg tgagccttta ctgcttgaat 24000
atggtagcat atatggtaaa gactttatca ttcgaccatc gctccaagtc attcttgaag 24060
atgaattaaa ttaaagtttt gacaccaatc tatcatggct gcaccagtag tccctactac 24120
tgacgcgtct tggtttcagg tgctcaaagc tcaaaacaaa aaggctactc atcctcagtt 24180
tcgtggcaat ggagttccgc ttaactccgc catcaaaccc gttgaaaacc atggctactg 24240
gctgcgttac accagacaaa agccaggtgg tactcccatt cctccatcct atgcctttta 24300
ttatactggc acaggtccca gaggaaatct taagtatggt gaactccctc ctaatgatac 24360
cccagcaacc actcgtgtta cttgggttaa gggttcggga gctgacactt ctattaaacc 24420
tcatgttgcc aaacgcaacc ccaacaatcc taaacatcag ctgctacctc tccgattccc 24480
aaccggagat ggcccagctc aaggtttcag agttgacccc ttcaacgcta gaggaagacc 24540
tcaggagcgt ggaagtggcc caagatctca atctgttaac tccagaggca caggcaatca 24600
gcccaggaaa cgcgaccaat ctgcaccagc tgcggtacgt cgtaagaccc agcatcaagc 24660
tcccaagcgg actttaccca agggtaaaac catttctcag gtatttggca accggtctcg 24720
tactggtgcc aatgtcggct ctgcagacac tgagaagacg ggtatggctg atcctcgcat 24780
catggctcta gccagacatg tgcctggtgt tcaggaaatg ctttttgctg gccaccttga 24840
gagcaacttt caggcggggg caattaccct taccttctcc tactcaatca cagtcaagga 24900
gggttctcct gactatgaga gacttaagga tgcgctcaat acggtcgtta accagaccta 24960
tgagccacct accaaaccaa ctaaggacaa gaagcctgac aaacaagacc agtctgctaa 25020
acccaaacag cagaagaaac ctaaaaaggt aactctgcca gcagacaaac aggattggga 25080
gtgggatgat gcttttgaga taaagcagga atcagcagcg tagacatcaa tctatgtctg 25140
ttaaacccac ccaactccac tcaaatatct ctttggttcc agagagtcgt agtgtatagc 25200
cagagagcca gtcagagggc gctatcatgc aaactagggc tggctactct agcacagaat 25260
cacatcccga taatcaacag tgctagaagg ttgattatac catttaatat gccgaggcca 25320
cgcggagtac gatcgagggt acagcataat ctcaactttt gttgagccac aattttaatc 25380
ctaattggag aaggccaaag gactgtacta cttttgtggg tgtagcagtc gcccagtggg 25440
aaagcgccaa ctaggttaca attgtggtgg ggacaaatta ggggaaatta aattggctta 25500
tagct 25505
<210> 2
<211> 25505
<212> RNA
<213> Porcine Deltacorona Virus ssRNA +
<400> 2
gcaccugaaa ccaagacgcg ucaguaguag ggacuacugg ugcagccaug auagauuggu 60
gucgguagag aacuagcgaa gcuagggaga uaaaauuaua gacuaaugcu auaauuuuua 120
ucuuuagucu auaauuuuau cucccuagcu ucgcuaguuc ucuaccgaca ccaauccagg 180
ugcgucugcc accaaguugg cuacccuucc uaggggcgcu uucgcgcuug cucaccauua 240
gauuaccugg aaaccagcca uucagguugg aguuucccca ggcucuuuug ugugggcauu 300
agcggcuugu gguuuuugca caaaaucuaa gcuacuuacc guuccucuga ccauccacca 360
cuucuauaga cagcacugau uaccguaggg uuuaagucac accggucugc accgcccguc 420
agcggacaca uuacccagca uagcacuccu ugcaccgagc cuagguagga uaaaaccccc 480
uaccggguga cucuuaaggc guuuccucca cgggauagcc acuagucacu agguguaagu 540
gaucugaucu gggcguauug uguugcgcaa gugugauacc cauaggagcg uggaauccua 600
uucugcggcu cagugccuga uauagcugug aaauggccaa gaacaagucc aagcgcgacg 660
cuauugcguu gccugaaaau guaccaccac cucugcaacu uuucauucau guugcagcug 720
cugaagaggg ucacccuaag guuacuacuu accuuggcaa cuauaaccuc uaugccacca 780
aggcuccgcc uggcgugcag guucuuagug cuaaaaccuc ucuuacugac uuugagaaug 840
ucuuuggagc ucaacccacc uugcgaucaa uucguaaucu gguuugugag gcucgcucgg 900
cugaauggac aacuuccaag aaugcuuuug cacucaaagc cacucaacuu gacuacucug 960
augccguuuu gagggcaaug auucguuucu gcccuccaaa gguguccaca cucguugccu 1020
uugcucuuuu uggccgauug guuaaaauug aggacaagga acuugcugag uuagcucgug 1080
auacugcccu ugaguuggcg uacacggcua aaauugguac aucucuugcu gacacgagau 1140
cugucucacu uauucauaag gacgcuuauc uaacucucag uaaugagguu guuggcguaa 1200
cuuuuacugc cgcacuuaug gcaaaggcua ccacuguuaa uggagcaaug caauacucaa 1260
acuuuuaccu cuacccucgu gccacuauua agguaaccga ugguaaggcu gaagcaauug 1320
caacuaagcc ucugucugcu gccacuaaag gcaagcaaau cacagaggau gucaaccuuc 1380
ucccugacua ucagcagcug cuuguugauc aagugacugg cacugagguu aagguuggag 1440
cucuaaccua uguuaagacc acugauucgc caccccuuua cuuucccaaa gucaagggug 1500
guguuauugg uauugcacuu aagcagcagg gcacugcggc uaagaagcuc aauguagucu 1560
uccaugcuca accugaugau guucugcuag ccuucauaca acuucagcaa uucuugaacc 1620
guacuucgga uucaaguguu gaaauuacug auugccagag uuaugaagua ucuccaacug 1680
ugacggucaa aauuggcccg ucuaaaccug gggaugucau cguggcuacu gaugaggaau 1740
accuuaaaug cuuugaaacc ccugagguag guaggcucua uaagguuuuc caaacucaau 1800
cuugggcuau cauugagcgu gccuucucca guuugaagau ccgcgugucc aaagcuuuau 1860
cagcauuuau aaguuuucug caaaaccuug cagauaacuu uacugcaaua agugguguug 1920
ucacugcacu cauucgugaa cuccaggauc uuacccugga uguggcgaca cguaucacua 1980
acauacaauu uguuuaccgu gccgguaagc uuauugucga cacgacaagu gucauagcua 2040
aacuuuucca gccauuuugu gauuuuauau caccuuuccu ucggaaaguu gcugguuuug 2100
caauuuacac uguugguaau cgcaugcuua uguuuaccag cacuggcacc uuucuucuca 2160
caaaggcaac uacuaagaua cucaauaagg caaaguacau cuuugaugug gagccugagu 2220
acccaguaga uguaacaaca uccaaaguug uaguacauga agcacuccag caaaccgaca 2280
cuaagccuac uagagcucug gaggcuguug augucguugu ugguaauacu guacugcaaa 2340
uggcuacuga uggcacugcg uucuacccau cggaugguac gcacgccucu cuuccaggau 2400
ucaaagcagg uucggaugag cuuuucauaa gcuucaacug cgaccucuuu gaugaugaga 2460
cuaaugcuca aaucaacgaa auacucgcug cauaugagcu uaaccaacua guggcuccag 2520
gugauucuac accgcgucaa auugcgacgu ugguugucga uacacuugca gaugcuauaa 2580
cagaccacuu uccggagaaa accauugauc uaccugaaga cuaucaaguc uuuucugacc 2640
augaugaccu cccacucgca caauaccaca ucccugauca ccugagccug uauauucagg 2700
cuauggaagg ugaagaugau aguggugaug aaauauguau ugaggacgau gauuacgacu 2760
guccucaagc cgacgaagac acagaaggag uaauucccca acagugggaa cuuccugaug 2820
uugauaaauu uuuacucaag auccaggaac ggaagaccag cagcgacgaa guacuuagcg 2880
ucgacgucua uccuaaacca gagccggucg gcaauguugg gauugacgac agcgcgucgg 2940
aaaagaagcc aaauggggac ucaguaccgg auccugaggu ccauccaaca cuagagagug 3000
uggauguuga acgaccaacc gaaacagcaa accaggcugu ugaagacaaa ccuucugaua 3060
ccaccuuugu gguugaugag gaacaauuac aagaaucaac accagaacau gaacuccgcu 3120
ccuaugaagg ggaguuugau ucugaugaug aaauuauuau uccuauagua ccaguaacac 3180
cugcggauuu aaaaccacag acuauuacua uaaaggagua cuuuaagucu gaaaaacuug 3240
agacuauuaa cgaaggaucc acagagucag uuacacaauc ugacgauucg uuugacgagu 3300
cauuuguuga ugcugagucu gaugauccac aagauccugc uguauaugau gauacaacaa 3360
uuauaacgga cagcacugau guaggcgaug agccugagac aacucuagcu accaucguua 3420
acacaccucu gacacucgau aauaacuugc caccugaagc cauuaaacaa cccagcccaa 3480
cuaagguuga guuaguuguu ggugaauugg cgaguauuaa auuugacaau ucuguucuag 3540
ucaacccugc uaaugcgcaa uuaacaaaug gcgguggagc ugcucgugca auugcaaaau 3600
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uuguaauguu gacacauacc cugcuaacgc ccuugugugc cgcuacgaca cucaucggca 17280
gaagcauuua auuggaccua augguucagc acuauauguu aauaagcaug cuuuucucac 17340
cccugagaug cauacuuaug cuacacauaa acucaacuug gcuccacuca ucuacuacuc 17400
caccacagau uguaguagug aacagccuau uguuguuacc uacagagauu gugucacccg 17460
gugcaauacu ggaaaaacuc ucuguccaaa ucaugcucuu gaauaccaag aguuuaucaa 17520
ugcauacaau cucauggcuc gccauggauu uaauguuuac auaccacgca augucaacgu 17580
uuacaacugu uggcuuacuu ucacuaaucu ccaaaaccuu gaaaacuuag cuuacaacug 17640
uuauuauaag aacugcaaug cucacguuga ugggcagcuu gauguaguua uuaauaauaa 17700
cgcuguauau gcuaaggucg acaauaaucu ugucaaacuu uucgacaacc gcacuaacuu 17760
accugucuca guggccuuug aacauuacac uaacaggcau acccguucac ugccaacuac 17820
acagcuguua ucugguuuag gcguaaccgc caccagaaau uucacugugu gguucgacaa 17880
ugauacaauu uuccaauaca cuauuaaugu aucuacguau acugacaucg acccuaguac 17940
ccauguuguc cucugugaug auagguacgg aacagauugg agucaguuua accaacuucc 18000
uaaugcagua uuccucacca aaacuaaggu gaagaaaaca gaaccguuug uuuguacagc 18060
acugacccua aauggcgucg ccauugacgg ugaagagcua uacaucuaug uacgcuauaa 18120
caaucaacug accacauuug cuacuacuug uacacagggu agaaauguug agcaguuuau 18180
accuaaaaca ccuauggaaa gagacuuccu ugagaugucu caacaguccu ucauccagca 18240
acaucaauug caggaacugg guguugaaca cauuaucuau ggugaugauu ccaguccagu 18300
cauuggcgga acucacacac uuaucucacu aguuaaaaac aaguuugaac aucagcuugu 18360
caaccauguu uacaacccag uccagaacug uguuguuacc ucaccuaacg caagcuccaa 18420
gaacguuugc acuguucuug auguucuucu ugaugacuac auugacauca uaagacaagc 18480
acaugccagu uacacaagua aauccaaagu auucacugug ucaauugaca accaacaaau 18540
uagauucaug cuuuggcaug augagcaagu caagacuugc uacccaaucu uacagucacu 18600
uaccaauggu uaccagaugc caucugugua caaaacauug guuacugacu uacaaccagc 18660
ugacaucccu aauuaucauu ccuacacccc ccgggugccu ggaguaguua agaauguuau 18720
caaguaccgc caacuuuuca acuacauagu uaaaaaggau agguuggcag uaccacacaa 18780
uaugaccgua uuacaccuug gagcugcauc ugcacuaggu acagcaccag guucuucagu 18840
cauaaaacaa auguuuccug aaggaacugu ucuuauugac cucgauauaa gagaguucac 18900
uucagaugcu aaccaaauaa uaguuacaga cuacagaacu uacauaccac cacaccacgu 18960
agacgucaua uuuucugacc ucuacuguug ugaugacaua cacuucuuug acaaucuaau 19020
aaggauaguu aaggagaggc ucgcccucgg ugguucuauc uuuguuaaga uaacugaaca 19080
uucauucuca cccgaacucu acucacuugc ggguugguuc gaugauuauc aacuauuuug 19140
cacagcaguc aaugccucgu cuucagaagc auuuuuaugc uguuuuaauu auuuggggcu 19200
ugcuaaggaa aacauuaaug guuuuaacuu acaugcuucc uauauucaau ggcgcaauga 19260
aauagcguug acaccaaccu auucuccuuu agcggacaac ccggcuacgg ccuguaagcu 19320
aaaagcaacg ccuauuaucu cggcucguga guuagagaag aagccuauuc uucgcuaucu 19380
cguugcauca gggcgccuuc uugugaggcc accagaaugc agagagcucu auugauuaug 19440
accuuauuuu gucucguucg agcaaaguuu gcugaugauc uacucgauuu gcucaccuuc 19500
ccgggugcac aucgcuucuu acauaaaccc acgaggaauu ccagcagucu cuacucgcgg 19560
gcuaauaaua auuuugaugu uggcguucuu ccuggcuacc ccacuaagaa cguuaaccuc 19620
uucucaccac uuacuaacuc cacuuugccc auuaauggcc uucaucggag uuaccaacca 19680
cucaugcuga auugucuuac uaaaauaacu aaccacacuc ucagcaugua ucuccuaccu 19740
agugagauac aaacuuauag cugcggcggu gccaugguua aauaccagac acaugaugca 19800
guucguauca uuuuagaccu cacugccacu gaccacaucu cuguugaagu cguuggccaa 19860
cauggugaaa auuauguguu uguuugcagu gagcagucua ccuacaccac ugcauuacac 19920
aaaucuaccu ucuucucacu uaauucugag cuuuauugcu uuacuaauaa caccuacuua 19980
gguauucuuc caccugauuu aacugacuuu acggucuacc guacugguca guucuaugcu 20040
aaugguuacc uuuuagguac uuuaccuauu acgguuaacu auguuagguu guaucggggu 20100
cauuugucug ccaauagugc ccacuuugcc cuugcaaacc uaaccgauac acucauaaca 20160
cuuaccaaua cuacuauauc gcaaaucacu uauugugaua agucuguagu ugauucaaua 20220
gcaugccagc gcucuucuca cgaaguggag gauggguuuu acuccaaccc uaaaucugcc 20280
guuagagcua ggcaacguac uauuguuaca cuaccuaagc ucccugagcu ugaaguagug 20340
caguuaaaua uuucugcaca cauggauuuu ggcgaagcca gacuugacag cguuaccauc 20400
aaugguaaca cauccuauug ugucacuaag ccuuacuuca ggcuugaaac uaacuuuaug 20460
uguacagguu gcacuaugaa ucugcgcacu gauaccugua guuuugaccu gucagcagua 20520
aacaauggca ugucauucuc ucaauucugu cuaagcacug aaucuggugc uugugagaug 20580
aaaauuauug uuaccuacgu auggaauuac uugcuaaggc agcguuugua uguuacugcu 20640
guagagggcc agacucacac uggaaccacu ucaguacaug caacagacac uucuagugua 20700
aucacugaug ucugcacuga cuacacuauc uauggagucu cugguacugg cauuauuaag 20760
ccaucagauc ucuuauugca caauggcaua guauucaccu cuccaacagg ugagcuuuau 20820
gcauuuaaaa auauaaccac uggcaaaacc cuucaggucu uaccguguga aaccccuucu 20880
caacugauug ugauaaacaa caccguuguc ggugcuauca cauccaguaa uucaacugaa 20940
aauaauaggu uuacuacuac uauugucaca ccuacuuucu uuuauuccac aaaugccacc 21000
acuuucaacu gcacuaagcc uguuuugucc uauggaccua ucagcgugug uagugauggu 21060
gcaauugugg gaacauccac auuacagaau acucgaccau ccauaguuuc acuauacgau 21120
ggcgaaguug aaauaccauc ugcauuuucu cuuuccguuc agacggagua cuugcaaguu 21180
caagcagagc aaguuauagu ugauuguccu caguauguau gcaauggcaa cagccguugu 21240
cuacaauuac uggcacaaua caccucagcu ugcucuaaga uugaagcagc ucugcauucc 21300
ucugcacagu uggauagcag agagauuaua aauauguuuc aaacaucaac acaguccuug 21360
caguuagcua auauuaccaa cuucaagggu gacuacaauu uuagcagcau acuaaccacc 21420
agaauuggug gcagaucugc uauugaagac cuucuuuuua auaaaguugu uacuaguggc 21480
cuuggcacug uugaucagga cuacaaaucc ugcucuagag acauggccau cgcugacuua 21540
guuuguuccc aguauuacaa uggcaucaug guucuaccug guguuguuga ugcugagaaa 21600
auggcaaugu auacuggcuc ucuuacugga gcuaugguau uuggaggacu gacugccgca 21660
gcggcaauac cauuugccac ggcaguacaa gcucgccuca auuaugucgc acugcaaaca 21720
aauguacuac aagaaaacca gaaaauucuu gcagaaucau uuaaccaagc aguuggcaau 21780
auaucacuug cacuaucuuc uguuaaugau gccauccagc aaacuucuga ggcucuuaac 21840
accguagcua uugcuauuaa aaagauucaa acaguuguua accagcaggg cgaggcauua 21900
ucacaccuga cugcacagcu gucaaacaau uuucaagcaa uuucgacuuc uauucaagac 21960
auuuacaacc gucuugagga aguagaggcu aaccagcaag uugaccgucu caucacagga 22020
cgguuggcug cacuuaaugc auauguuacu caguuacuca aucagauguc ucagauuaga 22080
caaucucgau uguuagcuca gcaaaagauu aaugagugug ucaaaucaca gucguccaga 22140
uacgguuucu guggaaaugg cacacacauc uucucacuua cacagacugc accaaauggc 22200
auauuuuuca ugcaugcagu gcuaguaccc aacaaauuca cacgugucaa cgcuucugcc 22260
ggcauuugug uggauaauac gagaggcuac ucauugcagc cucaacuuau acucuaccag 22320
uuuaauaacu ccuggagagu uacaccuaga aauauguaug aacccagacu gccccggcag 22380
gcugauuuca uacaauuaac ugauugcagc guuacuuuuu acaacaccac cgcugcuaau 22440
cuucccaaua uuaucccuga cauuauagau gucaaucaaa cagucaguga uauuauugac 22500
aauuuaccua cagcaacacc uccucagugg gauguuggua ucuauaacaa cacuauucuc 22560
aaccucaccg uugagauuaa ugaucuacaa gagcggucua aaaaccucuc acagauugca 22620
gaucguuuac aaaauuauau ugacaaucuu aacaauacuc uaguugaccu ugaauggcuc 22680
aacagagugg aaacuuaccu uaaauggccg ugguauauau ggcuugccau ugcccuggcu 22740
cuuauugcau uugugacaau ccucauaaca aucuuucuuu guacugguug uugugguggu 22800
ugcuuugguu guuguggcgg uuguuuuggc cuuuucucua agaagaaaag guauaccgac 22860
gaccaaccaa caccguccuu uaaguuuaag gaaugguagu cgacgacugg gccguuacca 22920
ucccuggaca auauauuauu gcuauacuag uugucaucug cauuggugug gcacuacuuu 22980
uuauuaauac uugcuuagcu uguguuaaau uauuuuacaa gugcuaccua ggggcagcau 23040
aucuuguuag gccuauuaua guguacuacu ccaagccgaa ccccguaccu gaggaugagu 23100
uuguaaaagu acaccaauuu ccuagaaaca cucacuaugu cugacgcaga agaguggcaa 23160
auuauuguuu ucauugcgau cauaugggcg cuuggcguca uccuccaggg aggcuaugcc 23220
acgcguaauc gugugaucua uguuauuaaa cuuauucugc uuuggcugcu ccaacccuuc 23280
acccuagugg ugaccauuug gaccgcaguc gacagaucau cuaagaagga cgcaguuuuc 23340
auugugucca uaauuuuugc cguacugacc uucauauccu gggccaagua cugguaugac 23400
ucaauucguu uauuaaugaa aaccagaucu gcaugggcac ucucaccuga gaguagacuc 23460
cuugcaggga uuauggaucc aauggguaca uggaggugca uucccauuga ccacauggcu 23520
ccaauucuca caccagucgu uaagcauggc aagcucaagc uacaugggca agagcuggcc 23580
aauggcauau cagucagaaa uccgccacag gauaugguga uagugucacc aagugacacc 23640
uuucacuaca cuuuuaagaa accuguggaa ucaaacaacg auccagaauu ugcuguucug 23700
auauaccagg gugaccgcgc uucaaacgcu ggacuucaca ccauaaccac uucaaaggcc 23760
ggugacgcuc gccuguauaa guauauguaa ugugcaacug ccaucugcag cugcgagauu 23820
uauauagauu gugcaauaag cggcacauca gaagagagga uguuccugag cuuauugacc 23880
cucucguuaa aacucgcugu uuugcuuaca gucucguggu ucuugcuaau gcuaauccaa 23940
uugcauuuag cauacuaccu cggaaaauuc uuaucaaugg ugagccuuua cugcuugaau 24000
augguagcau auaugguaaa gacuuuauca uucgaccauc gcuccaaguc auucuugaag 24060
augaauuaaa uuaaaguuuu gacaccaauc uaucauggcu gcaccaguag ucccuacuac 24120
ugacgcgucu ugguuucagg ugcucaaagc ucaaaacaaa aaggcuacuc auccucaguu 24180
ucguggcaau ggaguuccgc uuaacuccgc caucaaaccc guugaaaacc auggcuacug 24240
gcugcguuac accagacaaa agccaggugg uacucccauu ccuccauccu augccuuuua 24300
uuauacuggc acagguccca gaggaaaucu uaaguauggu gaacucccuc cuaaugauac 24360
cccagcaacc acucguguua cuuggguuaa ggguucggga gcugacacuu cuauuaaacc 24420
ucauguugcc aaacgcaacc ccaacaaucc uaaacaucag cugcuaccuc uccgauuccc 24480
aaccggagau ggcccagcuc aagguuucag aguugacccc uucaacgcua gaggaagacc 24540
ucaggagcgu ggaaguggcc caagaucuca aucuguuaac uccagaggca caggcaauca 24600
gcccaggaaa cgcgaccaau cugcaccagc ugcgguacgu cguaagaccc agcaucaagc 24660
ucccaagcgg acuuuaccca aggguaaaac cauuucucag guauuuggca accggucucg 24720
uacuggugcc aaugucggcu cugcagacac ugagaagacg gguauggcug auccucgcau 24780
cauggcucua gccagacaug ugccuggugu ucaggaaaug cuuuuugcug gccaccuuga 24840
gagcaacuuu caggcggggg caauuacccu uaccuucucc uacucaauca cagucaagga 24900
ggguucuccu gacuaugaga gacuuaagga ugcgcucaau acggucguua accagaccua 24960
ugagccaccu accaaaccaa cuaaggacaa gaagccugac aaacaagacc agucugcuaa 25020
acccaaacag cagaagaaac cuaaaaaggu aacucugcca gcagacaaac aggauuggga 25080
gugggaugau gcuuuugaga uaaagcagga aucagcagcg uagacaucaa ucuaugucug 25140
uuaaacccac ccaacuccac ucaaauaucu cuuugguucc agagagucgu aguguauagc 25200
cagagagcca gucagagggc gcuaucaugc aaacuagggc uggcuacucu agcacagaau 25260
cacaucccga uaaucaacag ugcuagaagg uugauuauac cauuuaauau gccgaggcca 25320
cgcggaguac gaucgagggu acagcauaau cucaacuuuu guugagccac aauuuuaauc 25380
cuaauuggag aaggccaaag gacuguacua cuuuuguggg uguagcaguc gcccaguggg 25440
aaagcgccaa cuagguuaca auuguggugg ggacaaauua ggggaaauua aauuggcuua 25500
uagcu 25505
<210> 3
<211> 3459
<212> DNA
<213> Artificial Sequence
<220>
<223> Deltacorona Virus SPIKE antigen CDS (NSVL) cDNA
<400> 3
atgaccttat tttgtctcgt tcgagcaaag tttgctgatg atctactcga tttgctcacc 60
ttcccgggtg cacatcgctt cttacataaa cccacgagga attccagcag tctctactcg 120
cgggctaata ataattttga tgttggcgtt cttcctggct accccactaa gaacgttaac 180
ctcttctcac cacttactaa ctccactttg cccattaatg gccttcatcg gagttaccaa 240
ccactcatgc tgaattgtct tactaaaata actaaccaca ctctcagcat gtatctccta 300
cctagtgaga tacaaactta tagctgcggc ggtgccatgg ttaaatacca gacacatgat 360
gcagttcgta tcattttaga cctcactgcc actgaccaca tctctgttga agtcgttggc 420
caacatggtg aaaattatgt gtttgtttgc agtgagcagt ctacctacac cactgcatta 480
cacaaatcta ccttcttctc acttaattct gagctttatt gctttactaa taacacctac 540
ttaggtattc ttccacctga tttaactgac tttacggtct accgtactgg tcagttctat 600
gctaatggtt accttttagg tactttacct attacggtta actatgttag gttgtatcgg 660
ggtcatttgt ctgccaatag tgcccacttt gcccttgcaa acctaaccga tacactcata 720
acacttacca atactactat atcgcaaatc acttattgtg ataagtctgt agttgattca 780
atagcatgcc agcgctcttc tcacgaagtg gaggatgggt tttactccaa ccctaaatct 840
gccgttagag ctaggcaacg tactattgtt acactaccta agctccctga gcttgaagta 900
gtgcagttaa atatttctgc acacatggat tttggcgaag ccagacttga cagcgttacc 960
atcaatggta acacatccta ttgtgtcact aagccttact tcaggcttga aactaacttt 1020
atgtgtacag gttgcactat gaatctgcgc actgatacct gtagttttga cctgtcagca 1080
gtaaacaatg gcatgtcatt ctctcaattc tgtctaagca ctgaatctgg tgcttgtgag 1140
atgaaaatta ttgttaccta cgtatggaat tacttgctaa ggcagcgttt gtatgttact 1200
gctgtagagg gccagactca cactggaacc acttcagtac atgcaacaga cacttctagt 1260
gtaatcactg atgtctgcac tgactacact atctatggag tctctggtac tggcattatt 1320
aagccatcag atctcttatt gcacaatggc atagtattca cctctccaac aggtgagctt 1380
tatgcattta aaaatataac cactggcaaa acccttcagg tcttaccgtg tgaaacccct 1440
tctcaactga ttgtgataaa caacaccgtt gtcggtgcta tcacatccag taattcaact 1500
gaaaataata ggtttactac tactattgtc acacctactt tcttttattc cacaaatgcc 1560
accactttca actgcactaa gcctgttttg tcctatggac ctatcagcgt gtgtagtgat 1620
ggtgcaattg tgggaacatc cacattacag aatactcgac catccatagt ttcactatac 1680
gatggcgaag ttgaaatacc atctgcattt tctctttccg ttcagacgga gtacttgcaa 1740
gttcaagcag agcaagttat agttgattgt cctcagtatg tatgcaatgg caacagccgt 1800
tgtctacaat tactggcaca atacacctca gcttgctcta agattgaagc agctctgcat 1860
tcctctgcac agttggatag cagagagatt ataaatatgt ttcaaacatc aacacagtcc 1920
ttgcagttag ctaatattac caacttcaag ggtgactaca attttagcag catactaacc 1980
accagaattg gtggcagatc tgctattgaa gaccttcttt ttaataaagt tgttactagt 2040
ggccttggca ctgttgatca ggactacaaa tcctgctcta gagacatggc catcgctgac 2100
ttagtttgtt cccagtatta caatggcatc atggttctac ctggtgttgt tgatgctgag 2160
aaaatggcaa tgtatactgg ctctcttact ggagctatgg tatttggagg actgactgcc 2220
gcagcggcaa taccatttgc cacggcagta caagctcgcc tcaattatgt cgcactgcaa 2280
acaaatgtac tacaagaaaa ccagaaaatt cttgcagaat catttaacca agcagttggc 2340
aatatatcac ttgcactatc ttctgttaat gatgccatcc agcaaacttc tgaggctctt 2400
aacaccgtag ctattgctat taaaaagatt caaacagttg ttaaccagca gggcgaggca 2460
ttatcacacc tgactgcaca gctgtcaaac aattttcaag caatttcgac ttctattcaa 2520
gacatttaca accgtcttga ggaagtagag gctaaccagc aagttgaccg tctcatcaca 2580
ggacggttgg ctgcacttaa tgcatatgtt actcagttac tcaatcagat gtctcagatt 2640
agacaatctc gattgttagc tcagcaaaag attaatgagt gtgtcaaatc acagtcgtcc 2700
agatacggtt tctgtggaaa tggcacacac atcttctcac ttacacagac tgcaccaaat 2760
ggcatatttt tcatgcatgc agtgctagta cccaacaaat tcacacgtgt caacgcttct 2820
gccggcattt gtgtggataa tacgagaggc tactcattgc agcctcaact tatactctac 2880
cagtttaata actcctggag agttacacct agaaatatgt atgaacccag actgccccgg 2940
caggctgatt tcatacaatt aactgattgc agcgttactt tttacaacac caccgctgct 3000
aatcttccca atattatccc tgacattata gatgtcaatc aaacagtcag tgatattatt 3060
gacaatttac ctacagcaac acctcctcag tgggatgttg gtatctataa caacactatt 3120
ctcaacctca ccgttgagat taatgatcta caagagcggt ctaaaaacct ctcacagatt 3180
gcagatcgtt tacaaaatta tattgacaat cttaacaata ctctagttga ccttgaatgg 3240
ctcaacagag tggaaactta ccttaaatgg ccgtggtata tatggcttgc cattgccctg 3300
gctcttattg catttgtgac aatcctcata acaatctttc tttgtactgg ttgttgtggt 3360
ggttgctttg gttgttgtgg cggttgtttt ggccttttct ctaagaagaa aaggtatacc 3420
gacgaccaac caacaccgtc ctttaagttt aaggaatgg 3459
<210> 4
<211> 1153
<212> PRT
<213> Porcine Deltacorona Virus
<400> 4
Met Thr Leu Phe Cys Leu Val Arg Ala Lys Phe Ala Asp Asp Leu Leu
1 5 10 15
Asp Leu Leu Thr Phe Pro Gly Ala His Arg Phe Leu His Lys Pro Thr
20 25 30
Arg Asn Ser Ser Ser Leu Tyr Ser Arg Ala Asn Asn Asn Phe Asp Val
35 40 45
Gly Val Leu Pro Gly Tyr Pro Thr Lys Asn Val Asn Leu Phe Ser Pro
50 55 60
Leu Thr Asn Ser Thr Leu Pro Ile Asn Gly Leu His Arg Ser Tyr Gln
65 70 75 80
Pro Leu Met Leu Asn Cys Leu Thr Lys Ile Thr Asn His Thr Leu Ser
85 90 95
Met Tyr Leu Leu Pro Ser Glu Ile Gln Thr Tyr Ser Cys Gly Gly Ala
100 105 110
Met Val Lys Tyr Gln Thr His Asp Ala Val Arg Ile Ile Leu Asp Leu
115 120 125
Thr Ala Thr Asp His Ile Ser Val Glu Val Val Gly Gln His Gly Glu
130 135 140
Asn Tyr Val Phe Val Cys Ser Glu Gln Ser Thr Tyr Thr Thr Ala Leu
145 150 155 160
His Lys Ser Thr Phe Phe Ser Leu Asn Ser Glu Leu Tyr Cys Phe Thr
165 170 175
Asn Asn Thr Tyr Leu Gly Ile Leu Pro Pro Asp Leu Thr Asp Phe Thr
180 185 190
Val Tyr Arg Thr Gly Gln Phe Tyr Ala Asn Gly Tyr Leu Leu Gly Thr
195 200 205
Leu Pro Ile Thr Val Asn Tyr Val Arg Leu Tyr Arg Gly His Leu Ser
210 215 220
Ala Asn Ser Ala His Phe Ala Leu Ala Asn Leu Thr Asp Thr Leu Ile
225 230 235 240
Thr Leu Thr Asn Thr Thr Ile Ser Gln Ile Thr Tyr Cys Asp Lys Ser
245 250 255
Val Val Asp Ser Ile Ala Cys Gln Arg Ser Ser His Glu Val Glu Asp
260 265 270
Gly Phe Tyr Ser Asn Pro Lys Ser Ala Val Arg Ala Arg Gln Arg Thr
275 280 285
Ile Val Thr Leu Pro Lys Leu Pro Glu Leu Glu Val Val Gln Leu Asn
290 295 300
Ile Ser Ala His Met Asp Phe Gly Glu Ala Arg Leu Asp Ser Val Thr
305 310 315 320
Ile Asn Gly Asn Thr Ser Tyr Cys Val Thr Lys Pro Tyr Phe Arg Leu
325 330 335
Glu Thr Asn Phe Met Cys Thr Gly Cys Thr Met Asn Leu Arg Thr Asp
340 345 350
Thr Cys Ser Phe Asp Leu Ser Ala Val Asn Asn Gly Met Ser Phe Ser
355 360 365
Gln Phe Cys Leu Ser Thr Glu Ser Gly Ala Cys Glu Met Lys Ile Ile
370 375 380
Val Thr Tyr Val Trp Asn Tyr Leu Leu Arg Gln Arg Leu Tyr Val Thr
385 390 395 400
Ala Val Glu Gly Gln Thr His Thr Gly Thr Thr Ser Val His Ala Thr
405 410 415
Asp Thr Ser Ser Val Ile Thr Asp Val Cys Thr Asp Tyr Thr Ile Tyr
420 425 430
Gly Val Ser Gly Thr Gly Ile Ile Lys Pro Ser Asp Leu Leu Leu His
435 440 445
Asn Gly Ile Val Phe Thr Ser Pro Thr Gly Glu Leu Tyr Ala Phe Lys
450 455 460
Asn Ile Thr Thr Gly Lys Thr Leu Gln Val Leu Pro Cys Glu Thr Pro
465 470 475 480
Ser Gln Leu Ile Val Ile Asn Asn Thr Val Val Gly Ala Ile Thr Ser
485 490 495
Ser Asn Ser Thr Glu Asn Asn Arg Phe Thr Thr Thr Ile Val Thr Pro
500 505 510
Thr Phe Phe Tyr Ser Thr Asn Ala Thr Thr Phe Asn Cys Thr Lys Pro
515 520 525
Val Leu Ser Tyr Gly Pro Ile Ser Val Cys Ser Asp Gly Ala Ile Val
530 535 540
Gly Thr Ser Thr Leu Gln Asn Thr Arg Pro Ser Ile Val Ser Leu Tyr
545 550 555 560
Asp Gly Glu Val Glu Ile Pro Ser Ala Phe Ser Leu Ser Val Gln Thr
565 570 575
Glu Tyr Leu Gln Val Gln Ala Glu Gln Val Ile Val Asp Cys Pro Gln
580 585 590
Tyr Val Cys Asn Gly Asn Ser Arg Cys Leu Gln Leu Leu Ala Gln Tyr
595 600 605
Thr Ser Ala Cys Ser Lys Ile Glu Ala Ala Leu His Ser Ser Ala Gln
610 615 620
Leu Asp Ser Arg Glu Ile Ile Asn Met Phe Gln Thr Ser Thr Gln Ser
625 630 635 640
Leu Gln Leu Ala Asn Ile Thr Asn Phe Lys Gly Asp Tyr Asn Phe Ser
645 650 655
Ser Ile Leu Thr Thr Arg Ile Gly Gly Arg Ser Ala Ile Glu Asp Leu
660 665 670
Leu Phe Asn Lys Val Val Thr Ser Gly Leu Gly Thr Val Asp Gln Asp
675 680 685
Tyr Lys Ser Cys Ser Arg Asp Met Ala Ile Ala Asp Leu Val Cys Ser
690 695 700
Gln Tyr Tyr Asn Gly Ile Met Val Leu Pro Gly Val Val Asp Ala Glu
705 710 715 720
Lys Met Ala Met Tyr Thr Gly Ser Leu Thr Gly Ala Met Val Phe Gly
725 730 735
Gly Leu Thr Ala Ala Ala Ala Ile Pro Phe Ala Thr Ala Val Gln Ala
740 745 750
Arg Leu Asn Tyr Val Ala Leu Gln Thr Asn Val Leu Gln Glu Asn Gln
755 760 765
Lys Ile Leu Ala Glu Ser Phe Asn Gln Ala Val Gly Asn Ile Ser Leu
770 775 780
Ala Leu Ser Ser Val Asn Asp Ala Ile Gln Gln Thr Ser Glu Ala Leu
785 790 795 800
Asn Thr Val Ala Ile Ala Ile Lys Lys Ile Gln Thr Val Val Asn Gln
805 810 815
Gln Gly Glu Ala Leu Ser His Leu Thr Ala Gln Leu Ser Asn Asn Phe
820 825 830
Gln Ala Ile Ser Thr Ser Ile Gln Asp Ile Tyr Asn Arg Leu Glu Glu
835 840 845
Val Glu Ala Asn Gln Gln Val Asp Arg Leu Ile Thr Gly Arg Leu Ala
850 855 860
Ala Leu Asn Ala Tyr Val Thr Gln Leu Leu Asn Gln Met Ser Gln Ile
865 870 875 880
Arg Gln Ser Arg Leu Leu Ala Gln Gln Lys Ile Asn Glu Cys Val Lys
885 890 895
Ser Gln Ser Ser Arg Tyr Gly Phe Cys Gly Asn Gly Thr His Ile Phe
900 905 910
Ser Leu Thr Gln Thr Ala Pro Asn Gly Ile Phe Phe Met His Ala Val
915 920 925
Leu Val Pro Asn Lys Phe Thr Arg Val Asn Ala Ser Ala Gly Ile Cys
930 935 940
Val Asp Asn Thr Arg Gly Tyr Ser Leu Gln Pro Gln Leu Ile Leu Tyr
945 950 955 960
Gln Phe Asn Asn Ser Trp Arg Val Thr Pro Arg Asn Met Tyr Glu Pro
965 970 975
Arg Leu Pro Arg Gln Ala Asp Phe Ile Gln Leu Thr Asp Cys Ser Val
980 985 990
Thr Phe Tyr Asn Thr Thr Ala Ala Asn Leu Pro Asn Ile Ile Pro Asp
995 1000 1005
Ile Ile Asp Val Asn Gln Thr Val Ser Asp Ile Ile Asp Asn Leu
1010 1015 1020
Pro Thr Ala Thr Pro Pro Gln Trp Asp Val Gly Ile Tyr Asn Asn
1025 1030 1035
Thr Ile Leu Asn Leu Thr Val Glu Ile Asn Asp Leu Gln Glu Arg
1040 1045 1050
Ser Lys Asn Leu Ser Gln Ile Ala Asp Arg Leu Gln Asn Tyr Ile
1055 1060 1065
Asp Asn Leu Asn Asn Thr Leu Val Asp Leu Glu Trp Leu Asn Arg
1070 1075 1080
Val Glu Thr Tyr Leu Lys Trp Pro Trp Tyr Ile Trp Leu Ala Ile
1085 1090 1095
Ala Leu Ala Leu Ile Ala Phe Val Thr Ile Leu Ile Thr Ile Phe
1100 1105 1110
Leu Cys Thr Gly Cys Cys Gly Gly Cys Phe Gly Cys Cys Gly Gly
1115 1120 1125
Cys Phe Gly Leu Phe Ser Lys Lys Lys Arg Tyr Thr Asp Asp Gln
1130 1135 1140
Pro Thr Pro Ser Phe Lys Phe Lys Glu Trp
1145 1150
<210> 5
<211> 25426
<212> DNA
<213> Artificial Sequence
<220>
<223> Porcine Deltacorona Virus BIVI-Isolate 2.0307 Genome cDNA
<400> 5
gattggtgtc ggtagagaac tagcgaagct agggagataa aattatagac taatgctata 60
atttttatct ttagtctata attttatctc cctagcttcg ctagttctct accgacacca 120
atccaggtgc gtctgccacc aagttggcta cccttcctag gggcgctttc gcgcttgctc 180
accattagat tacctggaaa ccagccattc aggttggagt ttccccaggc tcttttgtgt 240
gggcattagc ggcttgtggt ttttgcacaa aatctaagct acttaccgtt cctctgacca 300
tccaccactt ctatagacag cactgattac cgtagggttt aagtcacacc ggtctgcacc 360
gcccgtcagc ggacacatta cccagcatag cactccttgc accgagccta ggtaggataa 420
aaccccctac cgggtgactc ttaaggcgtt tcctccacgg gatagccact agtcactagg 480
tgtaagtgat ctgatctggg cgtattgtgt tgcgcaagtg tgatacccat aggagcgtgg 540
aatcctattc tgcggctcag tgcctgatat agctgtgaaa tggccaagaa caagtccaag 600
cgcgacgcta ttgcgttgcc tgaaaatgta ccaccacctc tgcaactttt cattcatgtt 660
gcagctgctg aagagggtca ccctaaggtt actacttacc ttggcaacta taacctctat 720
gccaccaagg ctccgcctgg cgtgcaggtt cttagtgcta aaacctctct tactgacttt 780
gagaatgtct ttggagctca acccaccttg cgatcaattc gtaatctggt ttgtgaggct 840
cgctcggctg aatggacaac ttccaagaat gcttttgcac tcaaagccac tcaacttgac 900
tactctgatg ccgttttgag ggcaatgatt cgtttctgcc ctccaaaggt gtccacactc 960
gctgcctttg ctctttttgg ccgattggtt aaaattgagg acaaggaact tgctgagtta 1020
gctcgtgata ctgcccttga gttggcgtac acggctaaaa ttggtacatc tcttgctgac 1080
acgagatctg tctcacttat tcataaggac gcttatctaa ctctcagtaa tgaggttgtt 1140
ggcgtaactt ttactgccgc acttatggca aaggctacca ctgttaatgg agcaatgcaa 1200
tactcaaact tttacctcta ccctcgtgcc actattaagg taaccgatgg taaggctgaa 1260
gcaattgcaa ctaagcctct gtctgctgcc actaaaggca agcaaatcac agaggatgtc 1320
aacctcctcc ctgactatca gcagctgctt gttgatcaag tgactggcac tgaggttaag 1380
gttggagctc taacctatgt taagaccact gattcgccac ccctttactt tcccaaagtc 1440
aagggtggtg ttattggtat tgcacttaag cagcagggca ctgcggctaa gaagctcaat 1500
gtagtcttcc atgctcaacc tgatgatgtt ctgctagcct tcatacaact tcagcaattc 1560
ttgaaccgta cttcggattc aagtgttgaa attactgatt gccagagtta tgaagtatct 1620
ccaactgtga cggtcaaaat tggcccgtct aaacctgggg atgtcatcgt ggctactgat 1680
gaggaatacc ttaaatgctt tgaaacccct gaggtaggta ggctctataa ggttttccaa 1740
actcaatctt gggctatcat tgagcgtgcc ttctccagtt tgaagatccg cgtgtccaaa 1800
gctttatcag catttataag ttttctgcaa aaccttgcag ataactttac tgcaataagt 1860
ggtgttgtca ctgcactcat tcgtgaactc caggatctta ccctggatgt ggcgacacgt 1920
atcactaaca tacaatttgt ttaccgtgcc ggtaagctta ttgtcgacac gacaagtgtc 1980
atagctaaac ttttccagcc attttgtgat tttatatcac ctttccttcg gaaagttgct 2040
ggttttgcaa tttacactgt tggtaatcgc atgcttatgt ttaccagcac tggcaccttt 2100
cttctcacaa aggcaactac taagatactc aataaggcaa agtacatctt tgatgtggag 2160
cctgagtacc cagtagatgt aacaacatcc aaagttgtag tacatgaagc actccagcaa 2220
accgacacta agcctactag agctctggag gctgttgatg tcgttgttgg taatactgta 2280
ctgcaaatgg ctactgatgg cactgcgttc tacccatcgg atggtacgca cgcctctctt 2340
ccaggattca aagcaggttc ggatgagctt ttcataagct tcaactgcga cctctttgat 2400
gatgagacta atgctcaaat caacgaaaca ctcgctgcat atgagcttaa ccaactagtg 2460
gctccaggtg attctacacc gcgtcaaatt gcgacgttgg ttgtcgatac acttgcagat 2520
gctataacag accactttcc ggagaaaacc attgatctac ctgaagacta tcaagtcttt 2580
tctgaccatg atgacctccc actcgcacaa taccacatcc ctgatcacct gagcctgtat 2640
attcaggcta tggaaggtga agatgatagt ggtgatgaaa tatgtattga ggacgatgat 2700
tacgactgtc ctcaagccga cgaagacaca gaaggagtaa ttccccaaca gtgggaactt 2760
cctgatgttg ataaattttt actcaagatc caggaacgga agaccagcag cgacgaagta 2820
cttagcgtcg acgtctatcc taaaccagag ccggtcggca atgttgggat tgacgacagc 2880
gcgtcggaaa agaagccaaa tggggactca gtaccggatc ctgaggtcca tccaacacta 2940
gagagtgtgg atgttgaacg accaaccgaa acagcaaacc aggctgttga agacaaacct 3000
tctgatacca cctttgtggt tgatgaggaa caattacaag aatcaacacc agaacatgaa 3060
ctccgctcct atgaagggga gtttgattct gatgatgaaa ttattattcc tatagtacca 3120
gtaacacctg cggatttaaa accacagact attactataa aggagtactt taagtctgaa 3180
aaacttgaga ctattaacga aggatccaca gagtcagtta cacaatctga cgattcgttt 3240
gacgagtcat ttgttgatgc tgagtctgat gatccacaag atcctgctgt atatgatgat 3300
acaacaatta taacggacag cactgatgta ggcgatgagc ctgagacaac tctagctacc 3360
atcgttaaca cacctctgac actcgataat aacttgccac ctgaagccat taaacaaccc 3420
agcccaacta aggttgagtt agttgttggt gaattggcga gtattaaatt tgacaattct 3480
gttctagtca accctgctaa tgcgcaatta acaaatggcg gtggagctgc tcgtgcaatt 3540
gcaaaattag ctggtccaaa atatcaagag tactgtaata gtgtggctcc tatctcagga 3600
ccgcttacca cggactcttt tgatgccaag aaatttggtg tagcctgcat cttgcatgta 3660
gtgccaccca aaggttctga ccctaatgta caagaactcc tgtatcaagc ttacaagagt 3720
atccttactg aaccagcaca ctatgttata cctatactag gtgctggtat ctttggatgc 3780
aatccagtcc actctctgga tgcgttcagg aaagcatgtc caagtgacat aggtcgtgtc 3840
acccttgtca ctatgaacaa aaaccatttg caggtgtggg atgctctcaa taggaccatt 3900
gtacgcacca ctactgacta tgatcaagtt accaccaagg cccttacacc ccagggagtg 3960
ttagaagcca atctctttga tggtgaggac tttgttcaag aaccaaaacc cggtcaaatc 4020
taccttgagg ttactgaaga agttcagaac caagccaagg aacttgacct taaccttcag 4080
caatactgcg tctacctgaa gacttgccac cataaatggg ttgtgagtcg tacgaacggg 4140
ttgatgcatc taaaacaaaa agataacaat tgttttgtta gtgcaggtgt aaacctgttt 4200
caaaacactg cttatcaact tagacctgct attgatgctc tctataggga gtatcttaat 4260
ggtaatccaa atagatttgt tgcttggatc tacgcatcca ctaaccgtcg tgttggtgag 4320
atgggttgtc cacagcaagt tatttctttg ctcgttagta actctgacgc agcattttca 4380
gcaactacag cctgttgtaa cacctacttt aaccacacag gtgttatttc agtagctcgt 4440
gaatatgacc caatacaacc aaaggtctac tgcatgaagt gtgatgtgtg gactcccttt 4500
acaccccaga gtggaaaagg tgcagttgca attggtattt ctgcagatga acctaccggt 4560
cctgccatta aatttgccgc agctcactgc tggtacacta atggcaagaa aacagttaat 4620
ggctatgaca ctaaagctaa tgttgtagct acctatcata ggtttgacgt gcctaagcct 4680
caacttgtcg aggacgtggt tgcgctgcct actaaaaatg actttgaagt tctcaatgtt 4740
gaagaactgc cgcaggatag tgtgctccat ttggacccac ctcctgtaca ggccttacaa 4800
cctaaggcta accaacacat tgagattcta gaaaacccag attatctgga cattttggat 4860
ctttggattc gtaaacccaa attcatcctc gtaaagtcgt ggagtgtttt gggtagagca 4920
ctatgtaagg caggtaaagt tgtctttgtc agtgcttcgc ttttgacccg tttctacaat 4980
taccttgtag agattggtgc tcttgactca acaataaggt tgtcagtcga tcttacctgt 5040
aaatttgtta gaacggttct cccatcgtct aacactgtac acaaaacttg tcttggtctg 5100
tattattcag cccagacact ttttgtttct ttagcaccat tccttatgtt accagctgta 5160
gttagtctgc ttaattcagg ctatacaatt ggcacatatt tgtatgcaaa aactggctgg 5220
ccttgtaatt acaatgccac gcaacacttt gattataatt cttactgtgc aggtgacttg 5280
gtttgtcaag cctgttttga cggtcaagac tccctacatt tgtatccgca tttacgtgtt 5340
aatcagcagc cccttcagac cactgactac actgtttatg cgctttcact aatactacta 5400
ttagctaaca tgactcttgt catgggcacg ctaatagtta ctttctttgt gaacttctat 5460
ggtgtgcaaa taccatttta tggtacactt ttgatagatt atcaatccgc actggtgatt 5520
actttctcag tgtactactt ttataaggta atgaagtttt tccgccatct cacacatgga 5580
tgtaaaattc caacgtgtgt ggtatgtgcc aaacttcgta ccccacctac tataacagtt 5640
gagactgtcg ttcagggcag gaaataccca tctgttattg aaacaaatgg cgggtttaca 5700
atttgtaaag aacacaactt ctattgcaag gactgctctt tacaaacacc cggcactttc 5760
atcccgacag aagctattga gtcgctctca cgagctacca ggcttagtgt caaaccaaca 5820
gcaccagcat tcttacttgc tagagatgtt gagtgccaaa ctgatgttgt cgttgctcgc 5880
gcaatgcata accaaaatgc gcatgtgtgc atttcaaaat actctgatat ccgtaccgtt 5940
gaccaactac ttaagcctac tccactgttt tcatacactc ccgatgttat catcgcggca 6000
gactttgaca acagaggtag tcttaagaca gctaaagaat tagctgtggt tttgtcaatg 6060
gaccttaaac gtactataat tatcattgat caggcctatt ctagacctat tgataattat 6120
caggaagttg cttctcgtat tgagaagtat tacccagttg caaagatcac acccacaggt 6180
gacatcttta cagacattaa gcaagcgacc aatggccaag ctagtgactc tgctattaat 6240
gcagctgttc tggctgtcca gcgcggtctt gattttacaa ttgacaaccc taacaacata 6300
ttaccacatt acgcctttga cttttcgacc ctcaatgcag aagaccagtc taccattttg 6360
gagagtggtt gtgctaaagg caatctcaag ggcactaatg ttggtgttgt tctttcagct 6420
agccttgtta cacgtcttag tcagcaggct atacgtgtga ttgctaatgc tgcttcacgt 6480
aatggtgtta catgcgctgt tactccttct acacttgtta tgcgtgggaa tattgcaaca 6540
cagcccttga ctcgcatcaa agctggtgca cctcccatgc gtcaaaaaat tttatgtgtt 6600
atcctggcac ttgctattgt gtactttgct gctatggctt ttggcttttt ggcaagtcaa 6660
attacgctta atacagtgcc tacgattaaa tctgatatcc gcgcctctac cttctacgtt 6720
gttagagatg gagtcttgga tactgttcgt tcaaatgaca agtgctttgc aaataagttt 6780
ttggcatttg atagcttcat tcaagcacct tacactaatt cacctgactg tccagttgtt 6840
gtgggagttg ttgatgtaac gacgcactct attcctggaa ttccagcagg tgtcattcat 6900
agagacggtc tcatacttaa catttatgaa cagtctcttt atgaaactca tcagcgtcag 6960
tctatggtta gggatgcgtt gtcactcaag acagcaaatc tctttaacct aggcaagcgt 7020
gttgtagtag gatacactca acatgaagtt gttgtgggta cctcctattt taattctcct 7080
gcacttttta atgcaaagtg caccttctta cagtatcagg acactagaca actctattgc 7140
tatgatactg ttcctactga acataagctc tactctgatg tgcttccgca cgtcgagtat 7200
aaggctattg acattaatgg tgatcttgtt cctttcaaga taccagagca gataatgttc 7260
tatccacata ttgtgcgcta tactagcaat tcctattgcc gtatggggca ttgttttaat 7320
actaaccctg gtatttgcat ttcatttacg gacgaatttc cgtatagtga aaatgtcaaa 7380
cctggtgtgt actgtgctga tacctctttg cagttgtttt caaacctcgt tttgggcact 7440
gtatctggta ttcacatctt tacatcaaca gctgcattgc ttggatctac tattgtgatc 7500
atactatgcg ttgttgctgt tcttgcagtt cagcgattct tcaaggagta cacaactttt 7560
gttatgtaca cttgtggtct tgctcttgtc aacattgtag gcattgcact tatgtacaag 7620
tgccttgtct tcgcgatttt ctattatgca atctaccttt actttgtcct tactttcccc 7680
tcctttaaga ggaatgtggc attgttttac ttcgctgtag tgatcgtgcc gcacgtgagt 7740
aacatgcaat tgcttgcgct cattgtgtgt agcattatct actttctcta cacctatgtt 7800
catactgtag ctaagacagc tgggaaattt tcttccttct tagacgcagc taaagctact 7860
tttgtcattg acaatgaaaa gtacgtgttg cttaaagacc tcgctggtgc tgaatttgac 7920
cagtatctgg cctcttacaa caagtacaaa tatttttctg gtactgcttc tgataaggat 7980
tatgataagg tctgtatggc atttcttgcc aaggctttgt catcttttcg tgaaggaggc 8040
ggttcacagt tgtacacacc acctaaattt gcagttgttc agagtcttaa gaccaagctg 8100
caagcaggta tcaaaatcct cctgcaccct tcaggtgtag ttgagcgatg tatggtctca 8160
gttgtctaca atggatctgc attgaatggc atctggctta agaatgttgt ctactgccca 8220
cgccatgtaa ttggaaaatt ccgtggtgac cagtggactc acatggtctc aattgctgat 8280
tgccgcgact ttatagtcaa gtgtccaata cagggtattc agctaaatgt ccaatcagtt 8340
aagatggtag gagctctcct ccagttaact gttcatacca acaacacagc cactccagac 8400
tataagtttg aaaggctcca accaggatca tcgatgacaa ttgcttgtgc ttatgatggc 8460
attgtacggc atgtctatca cgtggtcctc caacttaata atcttattta tgcaagcttc 8520
cttaacggag cttgtggtag tgtgggttac actcttaagg gtaaaacact ctacttacat 8580
tacatgcacc acattgagtt taacaacaaa actcatagtg gtacagatct tgaaggtaac 8640
ttctatggcc cctatgtgga tgaggaagtt attcagcaac aaacagcatt ccagtattac 8700
actgataatg ttgttgctca attatatgca cacttactga ctgttgatgc tagaccaaaa 8760
tggctggcac aatctcagat aagtatcgag gatttcaact catgggctgc taacaattcc 8820
tttgctaact tcccatgtga acaaactaat atgtcctaca ttatgggact ctcgcaaaca 8880
gctcgagtcc ctgtagaacg tatcctcaat accattatac agctaaccac caatagagat 8940
ggtgcttgta ttatgggatc ttatgatttc gagtgcgatt ggacgccaga gatggtatac 9000
aatcaggctc caatttcatt gcagtcagga gtagttaaga aaacttgtac gtggttcttc 9060
cacttcttgt ttatggctat taccatgcta ctcgctgcca tgcatgtttt ccctgtacac 9120
ttgtatccaa tagtactgcc atgcttcact gtcgtggcat tcctgttgac tttaaccatt 9180
aaacacactg ttgtgtttac cactacatac ttgcttccgt cacttttgat gatggttgta 9240
aatgctaaca ctttttggat accgaacaca tttctgcgca cctgctacga aactatattc 9300
ggttccccaa ttgctcagcg actgtatggt tacactgttg ctctttatat gctgatctat 9360
gctggacttg caatcaacta tacgttgaaa acactccggt atagagcaac ttcattctta 9420
tctttttgca tgcagtggtt tcaatatggt tatgttgcac acattgcgta caaactgctt 9480
aataaaccct ggacagaatc actactcttc acagccttca caatgctaac cagtcatcct 9540
ttgttggctg ctcttagctg gtggctagct ggtcgcgtaa ctctgcccat tatcatgcct 9600
gacttagcta ttcgtgtttt ggcgtataac gtcattggct atgtcatatg tgttcgattt 9660
ggccttatgt ggcttgcaaa tcggttcaca actgtaccta tgggcacata ccagtatatg 9720
gtgtctgtag agcaacttaa gtacatgatg gcagttaaga tgtccccacc gcgtaatgcg 9780
tttgaggtgc ttatagccaa cattagactt cttggtttgg gtggaaaccg taacattgct 9840
gtttctactg tccaaaacaa aattcttgat gcaaaagcta ctgctgttgt tgttgctaac 9900
cttcttgaaa aggctggcgt cacaaacaag cacgctattt gcaaaaagat tgtgaaactc 9960
cacaatgata cccttaaagc caccacttat gaggaggttg aggtagcact tgtgaaactt 10020
ctttctcaca taattgagtt cttgccaact gatcaggtag atgcttatct agctgatgcg 10080
gccaatgctc aacatgttaa tacctatttt gacaacttgc ttgagaacaa agctgttgtt 10140
caggctgttg ccgatatcaa cattaatctg gattcttata gaatttataa ggaggcagat 10200
gctatttata aacgatctgt tgagatgaac gaatctccgc aggagcaaaa gaaaaagctt 10260
aaagctgtta acattgcaaa ggcggaatgg gagcgtgagg ctgcttctca gcgtaagctt 10320
gaaaagcttg ctgatgctgc tatgaagtct atgtatcttg cagaacgtgc tgaggatcgt 10380
cgcattaagc taacctctgg acttactgca atgctttacc atatgcttag acgtcttgac 10440
tcagataggg taaaagctct gtttgagtgc gctaaggcac aaatcttgcc aatacatgct 10500
gtagtcggaa tttctaatga caaccttaaa gttattttta acgataagga cagctactct 10560
cattatgtag agggcaacac acttatacat aagggagttc gctacactat tgtgaagaaa 10620
ctctccttag ataatgcacc tattgaaggc gtaccagaag aattccctgt ggtcgttgag 10680
actgttaggg aaggtgtgcc ccagttgcaa aataatgagc tatgtttgcg caatgttttc 10740
actgctcaga acacagctca ggacttcaat ggcaatgaat ccactgtaaa atctttttat 10800
gttactagaa ccggtaagaa gattttggtt gccattacat caactaaaga caatcttaag 10860
actgtgacct gccttactga gaccggtaag acagtcctta acttggaccc ccctatgcgc 10920
ttcgcacata ccgtaggtgg aaaacagtct gttgtctatc tctattttat tcagaatatt 10980
agttcactca acagaggtat ggttattggc cacatctctg aaactactat ccttcaggca 11040
agtggcactc aaattgagta ccagcaaaat gcctctcttt tgacctattt ggctttcgct 11100
gtagacccta agacagccta ccttaagcat cttgctgatg gtgggtctcc tatacagggt 11160
tgtattcaga tgattgctac tatgggtcct ggatttgcag ttactactaa accacaacct 11220
aatgagcatc agtattctta tggtggtgct tcaatttgtc tttattgccg tgctcatata 11280
ccacatcctg gtgttgatgg acggtgcccc tacaaaggcc gctttgttca catcgacaaa 11340
gataaggaac ctgtttcctt cgccttgact catgagccat gcagttcttg tcaacggtgg 11400
gttaattatg actgcacctg cggatctagt ctgcagaatt cggcttattt aaacgagtaa 11460
cgggttctag tgacgcccgg ctagaacccc tgcagcctgg aactcaacca gatgctgtaa 11520
aaagggcctt ccatgtgcat aatgatacca cctctggtat attcttaagc acaaaatcta 11580
actgcgctcg gtttaaaacc acacgcagtg ccctgccttt acctaataag ggagaggttg 11640
aattgtactt tgttactaag cagtgtgcag ctaaagtctt cgaaatcgag gaggaatgct 11700
acaacgctct tagtacagag ctttatacta ctgatgatac atttggtgtc cttgccaaaa 11760
ctgagttctt taagtttgac aagataccta atgtcaatcg ccagtatctg actaaatata 11820
cactcctgga cttggcttat gctctacgtc atttgtcaac atctaaggat gttattcaag 11880
aaatcttgat caccatgtgc ggaacccctg aagattggtt tggggaaaat tggtttgatc 11940
caattgagaa cccatccttt tacaaggagt tccataaact tggggatatt cttaaccgtt 12000
gtgttcttaa tgccaataag tttgctagtg cctgtataga cgctggtctt gttggcatat 12060
taacacccga caaccaagac ctcctgggtc agatctatga ctttggagat tttattatta 12120
cacaaccagg taatggatgt gtggacttag catcctatta ttcttattta atgcccatta 12180
tgtccatgac tcacatgtta aagtgtgagt gtatggatag tgatggcaac ccacttgagt 12240
atgatggatt tcagtatgac ttcacggact tcaagcttgg cttgttcgag aagtatttta 12300
agtactggga ccgtccttac catcctaaca ctgttgaatg tccagatgac cgttgcgtat 12360
tgcactgtgc gaacttcaat gtgttgtttg ctatgtgtat acctaatacg gcatttggca 12420
atctttgttc aagagctact gttgatggcc accttgtggt ccagacagtg ggtgtacact 12480
tgaaagaact cggtatagtc cttaaccagg acgttaccac acacatggca aatattaatc 12540
taaacactct attgcgattg gttggtgatc ccaccactat tgcaagtgtc tcagacaagt 12600
gtgtagattt aagaactcct tgtcagacct tggctactat gtctagcgga attgctaaac 12660
agtcagtcaa gcccgggcat tttaatcaac acttctacaa gcatttgctt gatagtaacc 12720
tattagacca acttggaata gacattcgcc acttctacta tatgcaggat ggtgaagcgg 12780
ctatcacaga ctacagctac tacaggtata atacccccac gatggtagat atcaagatgt 12840
tcttattttg ccttgaggtg gcagataagt atcttgagcc ctacgaaggt ggatgtatta 12900
atgcacagtc agttgtggtc tctaatttgg acaagtcagc gggctacccc tttaacaagc 12960
taggtaaggc tcgtaactat tacgacatga ctcatgccga gcaaaatcaa ctgtttgagt 13020
atacaaaacg caatgttttg cctacactca ctcagatgaa ccttaagtat gcaatttcag 13080
ccaaggatcg tgctcgcact gtggcaggag tgtctataat tagcaccatg actaacaggc 13140
agtaccatca aaagatgctg aaatctattt cacttgcacg caatcagacc atcgtgattg 13200
gaacaaccaa attctatggt ggttgggaca acatgttacg acgactgatg tgtaatatca 13260
acaatcccat tttagtgggt tgggattacc ctaagtgtga tcgttctatg ccaaacatgc 13320
tgcgcattgc cgcttcgtgc ttgctagcac gaaaacacac ttgctgtaac caaagccagc 13380
gattctaccg tttggctaat gaatgttgcc aagtactatc tgaagtggta gtctctggta 13440
acaacctcta tgtaaaacca ggtggcacta gcagtggtga tgcaaccaca gcttatgcca 13500
attcggtatt taacatctta caggtggttt ctgctaatgt agccaccttc ttatcaactt 13560
ccaccacgac acatcttaat aaggacattg cggacttgca tcatagtctt tatgaagata 13620
tttatcgtgg tgactctaat gatatcactg tcatcaatag attctaccag catctccaaa 13680
gttactttgg acttatgata ttgtctgatg atggtgtcgc atgcatagac tcagccgttg 13740
caaaggctgg agctgttgct gatcttgatg gtttccgaga cattttgttt taccaaaaca 13800
atgtttacat ggcagactca aagtgttgga cagaaactga catgaatgtt ggccctcatg 13860
aattttgctc acagcatact gtgttagcag agcatgatgg taaaccttac tacttacctt 13920
acccagatgt ctctcgcatt ctgggtgcat gcatctttgt ggatgacgtt aacaaggctg 13980
accctgttca gaaccttgaa cgttacatct cacttgcaat tgatgcatat cccctcacca 14040
aggttgaccc tattaagggt aaagtctttt atttgttact agactacata cgtgttcttg 14100
ctcaggagtt acaggatggt atccttgatg ctttccaatc actcactgac atgtcgtatg 14160
taaataactt tatgaatgag gccttttatg ctcagatgta tgagcaaagt cctacactac 14220
aggccagcgg tgtttgtgtg gtgtgtaatt cacccactat actgcgctgt ggtgattgca 14280
ttcgtcgacc actactttgt tgcgtctgtg cctaccagca tgttacgcag actacacata 14340
aacgtatcat tgctatcaac aactacatct gtagtgttga gaattgcaat gaggacaatg 14400
ttgaaaaact tttcatttct ggcactgcga tttattgtga gaatcacaaa cctacgctgt 14460
gcatacccat tgtagctaac ggttctgttt ttggtatcta tcgccacact gcccgtggta 14520
gtgatgacat agacctcttt aacgagcttg ctacatctaa ctatgacact attgaacctt 14580
atcagaaggc caatcgtgca cctttatcac ttatgctctt cgctgctgaa accattaagg 14640
cactcgagga gtctatcaag aagtcatatg ctaccgcaac cgtcaaggat gtgtatgacc 14700
aacgcttcat taaacttcta tgggaacagg gtaaaaagcc gccacccata acgaagaacc 14760
acattttcac tggctaccat tttaacaaga atggaaaaac ccaagttggt gattacattc 14820
ttgctaaaac agatggcagt gacacttata cttacagagg aacatctacc tacaaactcc 14880
aaacaggtga tgttctagtc ttaatggcac atgttgttac accgctctca gcaccccctg 14940
tgctaacgca gacaacatat gtcagaaaat cacttttacc cgactctgtt ggtgcgtctt 15000
attatgtgca acattttaag tcatataatg agatagctat gcagagggtt acaacagtat 15060
taggtccacc aggcacaggt aagtcaacct ttgctattgg tttggctaag tactttccca 15120
gtgcacgtat ttgctacact gcgtcttcgc atgcagcaat cgatgcactc tgtgaaaaag 15180
ctttcaagac aatacctgta ggccaatgca gtcgtatcgt acccacacgt acaactgttg 15240
agtgctttca ggagtttgtc gtaaataaca caactgcaca gtatatcttc tcgactatca 15300
atgccttacc tgacattaag tgtgacattg tagttgtaga tgaggtttct atgttgacca 15360
attatgagct ttcctctgtg aatgctcgtt tggtttacaa tcacattgtg tatgttggtg 15420
atccttatca gttaccttca cctagaacta tgcttacgtc tggccagctt tcgccagctg 15480
actataacgt agttactgat ataatggtac atgcaggagc ggatgttatg ctcgacatgt 15540
gctacagatg cccacgtgaa atcgttgaga cagtgtctaa acttgtctac gataacaaac 15600
taaaagcggc gaaaccgaac tcaagacagt gttacaagac cattgtgaac tttggtcctg 15660
gagacgttgc tcatgaggga caatctgcct acaacgaagc acagttgcgt ttcgcactcg 15720
catttagaca acaaaagcgg tgggataacg tgactttcat atctccatat aatgctatga 15780
atgtgaaagc atccttagca ggtttctcta ctcagaccgt tgactcttct caaggttctg 15840
agtatgatta tgttatcttt tgcgtgacca ctgattcagc acacgcactt aacatggctc 15900
gtttgaacgt tgcccttaca cgtgcaaaga taggtatcct tgtggtgttt aggcaggcaa 15960
acgaacttta caatagtttg cagtttgaat ctattgattc acagcttcag tcgagtgctg 16020
agaaaaacct cacaccactg tttaagcgct gcggctatga gtataatggc gtccatccag 16080
ctcatgcttt gacctggcat gattgtggtg cagagtaccg ctgtgaggag ccacttgcta 16140
aattagtagg agttgccgat ggcactctta tatcatacaa aaccctagta tccacacttg 16200
ggtttcttcc atcacttaaa attgatgcat atcataatat gttcctaaca cgtgatgcgt 16260
gtcgcaccta tgttcagagt tggatcggca tagatgttga agcagcacac gccataaaac 16320
ctaacaccgg gactaacctg ccattgcaaa taggttttag taccggaaag aatttttcag 16380
tcactccaga gggaatttgg gtaaacgagc acggatcttg cactgagccc gtccctgcca 16440
aaatacctcc tggagaacaa tttcgtcacc ttaaaaagga catgcgccag gcgcgtcctt 16500
ggatggttgt tcgacgtgag attgctactc acattgctga ggtagctcct cacactgatt 16560
atatatgctt tgtcacttgg gctcaccagc ttgagctagc gacaatgcgc tactttgtca 16620
aactaggtat ggaagagaga tgcttttgtg gcaggcgggc ttgtttcact aatggaactg 16680
agttcgcttg caaagcacac cattctctca ccattccaca atgtgattat gtgtacaatc 16740
cattcctcat cgacgtggct acgtggggat tctcgggacg gctttccacc aaccatgacg 16800
cggtgtgcac atatcatgct aatgcccatg ttgcatcagc tgatgcaatc atgacggtat 16860
gtttagctat ccatgaactg ttcagtactg ttgactggaa ctttgaattt ccagtaactg 16920
ctgagcaatc gcaacttaac aaggcctgtc gcttagtaca ggcgaattac ttaaatatac 16980
tactcactac aaccaaagcc acggtggttc acgatattgg taacccaaaa ggtatcccta 17040
tcgtgcgcaa acctggtgtt aaatatcact tctatgatca agcacccatt gtcaaacacg 17100
ttcaaaaact aaagtacaag ccagagatgg aggcccgttt caccgatggt ttgactatgt 17160
tttggaattg taatgttgac acataccctg ctaacgccct tgtgtgccgc tacgacactc 17220
atcggcagaa gcatttaatt ggacctaatg gttcagcact atatgttaat aagcatgctt 17280
ttctcacccc tgagatgcat acttatgcta cacataaact caacttggct ccactcatct 17340
actactccac cacagattgt agtagtgaac agcctattgt tgttacctac agagattgtg 17400
tcacccggtg caatactgga aaaactctct gtccaaatca tgctcttgaa tatcaagagt 17460
ttatcaatgc atacaatctc atggctcgcc atggatttaa tgtttacata ccacgcaatg 17520
tcaacgttta caactgttgg cttactttca ctaatctcca aaaccttgaa aacttagctt 17580
acaactgtta ttataagaac tgcaatgctc acgttgatgg gcagcttgat gtagttatta 17640
ataataacgc tgtatatgct aaggtcgaca ataatcttgt caaacttttc gacaaccgca 17700
ctaacttacc tgtctcagtg gcctttgaac attacactaa caggcatacc cgttcactgc 17760
caactacaca gctgttatct ggtttaggcg taaccgccac cagaaatttc actgtgtggt 17820
tcgacaatga tacaattttc caatacacta ttaatgtatc tacgtatact gacatcgacc 17880
ctagtaccca tgttgtcctc tgtgatgata ggtacggaac agattggagt cagtttaacc 17940
aacttcctaa tgcagtattc ctcaccaaaa ctaaggtgaa gaaaacagaa ccgtttgttt 18000
gtacagcact gaccctaaat ggcgtcgcca ttgacggtga agagctatac atctatgtac 18060
gctataacaa tcaactgacc acatttgcta ctacttgtac acagggtaga aatgttgagc 18120
agtttatacc taaaacacct atggaaagag acttccttga gatgtctcaa cagtccttca 18180
tccagcaaca tcaattgcag gaactgggtg ttgaacacat tatctatggt gatgattcca 18240
gtccagtcat tggcggaact cacacactta tctcactagt taaaaacaag tttgaacatc 18300
agcttgtcaa ccatgtttac aacccagtcc agaactgtgt tgttacctca cctaacgcaa 18360
gctccaagaa cgtttgcact gttcttgatg ttctacttga tgactacatt gacatcataa 18420
gacaagcaca tgccagttac acaagtaaat ccaaagtatt cactgtgtca attgacaacc 18480
aacaaattag attcatgctt tggcatgatg agcaagtcaa gacttgctac ccaatcttac 18540
agtcacttac caatggttac cagatgccat ctgtgtacaa aacattggtt actgacttac 18600
aaccagctga catccctaat tatcattcct acaccccccg ggtgcctgga gtagttaaga 18660
atgttatcaa gtaccgccaa cttttcaact acatagttaa aaaggatagg ttggcagtac 18720
cacacaatat gaccgtatta caccttggag ctgcatctgc actaggtaca gcaccaggtt 18780
cttcagtcat aaaacaaatg tttcctgaag gaactgttct tattgacctc gatataagag 18840
agttcacttc agatgctaac caaataatag ttacagacta cagaacttac ataccaccac 18900
accacgtaga cgtcatattt tctgacctct actgttgtga tgacatacac ttctttgaca 18960
atctaataag gatagttaag gagaggctcg ccctcggtgg ttctatcttt gttaagataa 19020
ctgaacattc attctcaccc gaactctact cacttgcggg ttggttcgat gattatcaac 19080
tattttgcac agcagtcaat gcctcgtctt cagaagcatt tttatgctgt tttaattatt 19140
tggggcttgc taaggaaaac attaatggtt ttaacttaca tgcttcctat attcaatggc 19200
gcaatgaaat agcgttgaca ccaacctatt ctcctttagc ggacaacccg gctacggcct 19260
gtaagctaaa agcaacgcct attatctcgg ctcgtgagtt agagaagaag cctattcttc 19320
gctatctcgt tgcatcaggg cgccttcttg tgaggccacc agaatgcaga gagctctatt 19380
gattatgacc ttactttgtc tcgttcgagc aaagtttgct gatgatctac tcgacttgct 19440
caccttcccg ggtgcacatc gcttcttaca taaacccacg aggaattcca gcagtctcta 19500
ctcgcgggct aataataatt ttgatgttgg cgttcttcct ggctacccca ctaagaacgt 19560
taacctcttc tcaccactta ctaactccac tttgcccatt aatggccttc atcggagtta 19620
ccaaccactc atgctgaatt gtcttactaa aataactaac cacactctca gcatgtatct 19680
cctacctagt gagatacaaa cttatagctg cggcggtgcc atggttaaat accagacaca 19740
tgatgcagtt cgtatcattt tagacctcac tgccactgac cacatctctg ttgaagtcgt 19800
tgaccaacat ggtgaaaatt atgtgtttgt ttgcagtgag cagtttaact acaccactgc 19860
attacacaat tctaccttct tctcacttaa ttctgcgctt tattgcttta ctaataacac 19920
ctacttaggt attcttccac ctgatttaac tgactttacg gtctaccgta ctggtcagtt 19980
ctatgctaat ggttaccttt taggtacttt acctattacg gttaactatg ttaggttgta 20040
tcggggtcat ttgtctgcca atagtgccca ctttgccctt gcaaacctaa ccgatacact 20100
cataacactt accaatacta ctatatcgca aatcacttat tgtgataagt cagtagttga 20160
ttcaatagca tgccagcgct cttctcacga agtggaggat gggttttact ccgaccctaa 20220
atctgccgtt agagctaggc aacgtactat tgttacacta cctaagctcc ctgagcttga 20280
agtagtgcag ttaaatattt ctgcacacat ggattttggc gaagccagac ttgacagcgt 20340
taccatcaat ggtaacacat cctattgtgt cactaagcct tacttcaggc ttgaaactaa 20400
ctttatgtgt acaggttgca ctatgaatct gcgcactgat acctgtagtt ttgacctgtc 20460
agcagtaaac aatggcatgt cattctctca attctgtcta agcactgaat ctggtgcttg 20520
tgagatgaaa attattgtta cctacgtatg gaattacttg ctaaggcagc gtttgtatgt 20580
tactgctgta gagggccaga ctcacactgg aaccacttca gtacatgcaa cagacacttc 20640
tagtgtaatc actgatgtct gcactgacta cactatctat ggagtctctg gtactggcat 20700
tattaagcca tcagatctct tattgcacaa tggcatagca ttcacctctc caacaggtga 20760
gctttatgca tttaaaaata taaccactgg caaaaccctt caggtcttac cgtgtgaaac 20820
cccttctcaa ctgattgtga taaacaacac cgttgtcggt gctatcacat ccagtaattc 20880
aactgaaaat aataggttta ctactactat tgtcacacct actttctttt attccacaaa 20940
tgccaccact ttcaactgca ctaagcctgt tttgtcctat ggacctatca gcgtgtgtag 21000
tgatggtgca attgtgggaa catccacatt acagaatact cgaccatcca tagttttact 21060
atacgatggc gaagttgaaa taccatctgc attttctctt tccgttcaga cggagtactt 21120
gcaagttcaa gcagagcaag ttatagttga ttgtcctcag tatgtatgca atggcaacag 21180
ccgttgtcta caattactgg cacaatacac ctcagcttgc tctaacattg aagcagctct 21240
gcattcctct gcacagttgg atagcagaga gattataaat atgtttcaaa catcaacaca 21300
gtccttgcag ttagctaata ttaccaactt caagggtgac tacaatttta gcagcatact 21360
aaccaccaga attggtggca gatctgctat tgaagacctt ctttttaata aagttgttac 21420
tagtggcctt ggcactgttg atcaggacta caaatcctgc tctagagaca tggccatcgc 21480
tgacttagtt tgttcccagt attacaatgg catcatggtt ctacctggtg ttgttgatgc 21540
tgagaaaatg gcaatgtata ctggctctct tactggagct atggtatttg gaggactgac 21600
tgccgcagcg gcaataccat ttgccacggc agtacaagct cgcctcaatt atgtcgcact 21660
gcaaacaaat gtactacaag aaaaccagaa aattcttgca gaatcattta accaagcagt 21720
tggcaatata tcacttgcac tatcttctgt taatgatgcc atccagcaaa cttctgaggc 21780
tcttaacacc gtagctattg ctattaaaaa gattcaaaca gttgttaacc agcagggcga 21840
ggcattatca cacctgactg cacagctgtc aaacaatttt caagcaattt cgacttctat 21900
tcaagacatt tacaaccgtc ttgaggaagt agaggctaac cagcaagttg accgtctcat 21960
cacaggacgg ttggctgcac ttaatgcata tgttactcag ttactcaatc agatgtctca 22020
gattagacaa tctcgattgt tagctcagca aaagattaat gagtgtgtca aatcacagtc 22080
gtccagatac ggtttctgtg gaaatggcac acacatcttc tcacttacac agactgcacc 22140
aaatggcata tttttcatgc atgcagtgct agtacccaac aaattcacac gtgtcaacgc 22200
ttctgccggc atttgtgtgg ataatacgag aggctactca ttgcagcctc aacttatact 22260
ctaccagttt aataactcct ggagagttac acctagaaat atgtatgaac ccagactgcc 22320
ccggcaggct gatttcatac aattaactga ttgcagcgtt actttttaca acaccaccgc 22380
tgctaatctt cccaatatta tccctgacat tatagatgtc aatcaaacag tcagtgatat 22440
tattgacaat ttacctacag caacacctcc tcagtgggat gttggtatct ataacaacac 22500
tattctcaac ctcaccgttg agattaatga tctacaagag cggtctaaaa acctctcaca 22560
gattgcagat cgtttacaaa attatattga caatcttaac aatactctag ttgaccttga 22620
atggctcaac agagtggaaa cttaccttaa atggccgtgg tatatatggc ttgccattgc 22680
cctggctctt attgcatttg tgacaatcct cataacaatc tttctttgta ctggttgttg 22740
tggtggttgc tttggttgtt gtggcggttg ttttggcctt ttctctaaga agaaaaggta 22800
taccgacgac caaccaacac cgtcctttaa gtttaaggaa tggtagtcga cgactgggcc 22860
gttaccatcc ctggacaata tattattgct atactagttg tcatctgcat tggtgtggca 22920
ctacttttta ttaatacttg cttagcttgt gttaaattat tttacaagtg ctacctaggg 22980
gcagcatatc ttgttaggcc tattatagtg tactactcca agccgaaccc cgtacctgag 23040
gatgagtttg taaaagtaca ccaatttcct agaaacactc actatgtctg acgcagaaga 23100
gtggcaaatt attgttttca ttgcgatcat atgggcgctt ggcgtcatcc tccagggagg 23160
ctatgccacg cgtaatcgtg tgatctatgt tattaaactt attctgcttt ggctgctcca 23220
acccttcacc ctagtggtga ccatttggac cgcagtcgac agatcatcta agaaggacgc 23280
agttttcatt gtgtccataa tttttgccgt actgaccttc atatcctggg ccaagtactg 23340
gtatgactca attcgtttat taatgaaaac cagatctgca tgggcactct cacctgagag 23400
tagactcctt gcagggatta tggatccaat gggtacatgg aggtgcattc ccatcgacca 23460
catggctcca attctcacac cagtcgttaa gcatggcaag ctcaagctac atgggcaaga 23520
gctggccaat ggcatatcag tcagaaatcc gccacaggat atggtgatag tgtcaccaag 23580
tgacaccttt cactacactt ttaagaaacc tgtggaatca aacaacgatc cagaatttgc 23640
tgttctgata taccagggtg accgcgcttc aaacgctgga cttcacacca taaccacttc 23700
aaaggccggt gacgctcgcc tgtataagta tatgtaatgt gcaactgcca tctgcagctg 23760
cgagatttat atagattgtg caataagcgg cacatcagaa gagaggatgt tcctgagctt 23820
attgaccctc tcgttaaaac tcgctgtttt gcttacagtc tcgtggttct tgctaatgct 23880
aatccaattg catttagcat actacctcgg aaaattctta tcaatggtga gcctttactg 23940
cttgaatatg gtagcatata tggtaaagac tttatcattc gaccatcgct ccaagtcatt 24000
cttgaagatg aattaaatta aagttttgac accaatctat catggctgca ccagtagtcc 24060
ctactactga cgcgtcttgg tttcaggtgc tcaaagctca aaacaaaaag gctactcatc 24120
ctcagtttcg tggcaatgga gttccgctta actccgccat caaacccgtt gaaaaccatg 24180
gctactggct gcgttacacc agacaaaagc caggtggtac tcccattcct ccatcctatg 24240
ccttttatta tactggcaca ggtcccagag gaaatcttaa gtatggtgaa ctccctccta 24300
atgatacccc agcaaccact cgtgttactt gggttaaggg ttcgggagct gacacttcta 24360
ttaaacctca tgttgccaaa cgcaacccca acaatcctaa acatcagctg ctacctctcc 24420
gattcccaac cggagatggc ccagctcaag gtttcagagt tgaccccttc aacgctagag 24480
gaagacctca ggagcgtgga agtggcccaa gatctcaatc tgttaactcc agaggcacag 24540
gcaatcagcc caggaaacgc gaccaatctg caccagctgc ggtacgtcgt aagacccagc 24600
atcaagctcc caagcggact ttacccaagg gtaaaaccat ttctcaggta tttggcaacc 24660
ggtctcgtac tggtgccaat gtcggctctg cagacactga gaagacgggt atggctgatc 24720
ctcgcatcat ggctctagcc agacatgtgc ctggtgttca ggaaatgctt tttgctggcc 24780
accttgagag caactttcag gcgggggcaa ttacccttac cttctcctac tcaatcacag 24840
tcaaggaggg ttctcctgac tatgagagac ttaaggatgc gctcaatacg gtcgttaacc 24900
agacctatga gccacctacc aaaccaacta aggacaagaa gcctgacaaa caagaccagt 24960
ctgctaaacc caaacagcag aagaaaccta aaaaggtaac tctgccagca gacaaacagg 25020
attgggagtg ggatgatgct tttgagataa agcaggaatc agcagcgtag acatcaatct 25080
atgtctgtta aacccaccca actccactca aatatctctt tggttccaga gagtcgtagt 25140
gtatagccag agagccagtc agagggcgct atcatgcaaa ctagggctgg ctactctagc 25200
acagaatcac atcccgataa tcaacagtgc tagaaggttg attataccat ttaatatgcc 25260
gaggccacgc ggagtacgat cgagggtaca gcataatctc aacttttgtt gagccacaat 25320
tttaatccta attggagaag gccaaaggac tgtactactt ttgtgggtgt agcagtcgcc 25380
cagtgggaaa gcgccaacta ggttacaatt gtggtgggga caaatt 25426
<210> 6
<211> 25426
<212> RNA
<213> Artificial Sequence
<220>
<223> Porcine Deltacorona Virus BIV-Isolate 2.0307 ssRNA + Genome
<400> 6
gauugguguc gguagagaac uagcgaagcu agggagauaa aauuauagac uaaugcuaua 60
auuuuuaucu uuagucuaua auuuuaucuc ccuagcuucg cuaguucucu accgacacca 120
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caaaggcugg agcuguugcu gaucuugaug guuuccgaga cauuuuguuu uaccaaaaca 13800
auguuuacau ggcagacuca aaguguugga cagaaacuga caugaauguu ggcccucaug 13860
aauuuugcuc acagcauacu guguuagcag agcaugaugg uaaaccuuac uacuuaccuu 13920
acccagaugu cucucgcauu cugggugcau gcaucuuugu ggaugacguu aacaaggcug 13980
acccuguuca gaaccuugaa cguuacaucu cacuugcaau ugaugcauau ccccucacca 14040
agguugaccc uauuaagggu aaagucuuuu auuuguuacu agacuacaua cguguucuug 14100
cucaggaguu acaggauggu auccuugaug cuuuccaauc acucacugac augucguaug 14160
uaaauaacuu uaugaaugag gccuuuuaug cucagaugua ugagcaaagu ccuacacuac 14220
aggccagcgg uguuugugug guguguaauu cacccacuau acugcgcugu ggugauugca 14280
uucgucgacc acuacuuugu ugcgucugug ccuaccagca uguuacgcag acuacacaua 14340
aacguaucau ugcuaucaac aacuacaucu guaguguuga gaauugcaau gaggacaaug 14400
uugaaaaacu uuucauuucu ggcacugcga uuuauuguga gaaucacaaa ccuacgcugu 14460
gcauacccau uguagcuaac gguucuguuu uugguaucua ucgccacacu gcccguggua 14520
gugaugacau agaccucuuu aacgagcuug cuacaucuaa cuaugacacu auugaaccuu 14580
aucagaaggc caaucgugca ccuuuaucac uuaugcucuu cgcugcugaa accauuaagg 14640
cacucgagga gucuaucaag aagucauaug cuaccgcaac cgucaaggau guguaugacc 14700
aacgcuucau uaaacuucua ugggaacagg guaaaaagcc gccacccaua acgaagaacc 14760
acauuuucac uggcuaccau uuuaacaaga auggaaaaac ccaaguuggu gauuacauuc 14820
uugcuaaaac agauggcagu gacacuuaua cuuacagagg aacaucuacc uacaaacucc 14880
aaacagguga uguucuaguc uuaauggcac auguuguuac accgcucuca gcacccccug 14940
ugcuaacgca gacaacauau gucagaaaau cacuuuuacc cgacucuguu ggugcgucuu 15000
auuaugugca acauuuuaag ucauauaaug agauagcuau gcagaggguu acaacaguau 15060
uagguccacc aggcacaggu aagucaaccu uugcuauugg uuuggcuaag uacuuuccca 15120
gugcacguau uugcuacacu gcgucuucgc augcagcaau cgaugcacuc ugugaaaaag 15180
cuuucaagac aauaccugua ggccaaugca gucguaucgu acccacacgu acaacuguug 15240
agugcuuuca ggaguuuguc guaaauaaca caacugcaca guauaucuuc ucgacuauca 15300
augccuuacc ugacauuaag ugugacauug uaguuguaga ugagguuucu auguugacca 15360
auuaugagcu uuccucugug aaugcucguu ugguuuacaa ucacauugug uauguuggug 15420
auccuuauca guuaccuuca ccuagaacua ugcuuacguc uggccagcuu ucgccagcug 15480
acuauaacgu aguuacugau auaaugguac augcaggagc ggauguuaug cucgacaugu 15540
gcuacagaug cccacgugaa aucguugaga cagugucuaa acuugucuac gauaacaaac 15600
uaaaagcggc gaaaccgaac ucaagacagu guuacaagac cauugugaac uuugguccug 15660
gagacguugc ucaugaggga caaucugccu acaacgaagc acaguugcgu uucgcacucg 15720
cauuuagaca acaaaagcgg ugggauaacg ugacuuucau aucuccauau aaugcuauga 15780
augugaaagc auccuuagca gguuucucua cucagaccgu ugacucuucu caagguucug 15840
aguaugauua uguuaucuuu ugcgugacca cugauucagc acacgcacuu aacauggcuc 15900
guuugaacgu ugcccuuaca cgugcaaaga uagguauccu ugugguguuu aggcaggcaa 15960
acgaacuuua caauaguuug caguuugaau cuauugauuc acagcuucag ucgagugcug 16020
agaaaaaccu cacaccacug uuuaagcgcu gcggcuauga guauaauggc guccauccag 16080
cucaugcuuu gaccuggcau gauuguggug cagaguaccg cugugaggag ccacuugcua 16140
aauuaguagg aguugccgau ggcacucuua uaucauacaa aacccuagua uccacacuug 16200
gguuucuucc aucacuuaaa auugaugcau aucauaauau guuccuaaca cgugaugcgu 16260
gucgcaccua uguucagagu uggaucggca uagauguuga agcagcacac gccauaaaac 16320
cuaacaccgg gacuaaccug ccauugcaaa uagguuuuag uaccggaaag aauuuuucag 16380
ucacuccaga gggaauuugg guaaacgagc acggaucuug cacugagccc gucccugcca 16440
aaauaccucc uggagaacaa uuucgucacc uuaaaaagga caugcgccag gcgcguccuu 16500
ggaugguugu ucgacgugag auugcuacuc acauugcuga gguagcuccu cacacugauu 16560
auauaugcuu ugucacuugg gcucaccagc uugagcuagc gacaaugcgc uacuuuguca 16620
aacuagguau ggaagagaga ugcuuuugug gcaggcgggc uuguuucacu aauggaacug 16680
aguucgcuug caaagcacac cauucucuca ccauuccaca augugauuau guguacaauc 16740
cauuccucau cgacguggcu acguggggau ucucgggacg gcuuuccacc aaccaugacg 16800
cggugugcac auaucaugcu aaugcccaug uugcaucagc ugaugcaauc augacgguau 16860
guuuagcuau ccaugaacug uucaguacug uugacuggaa cuuugaauuu ccaguaacug 16920
cugagcaauc gcaacuuaac aaggccuguc gcuuaguaca ggcgaauuac uuaaauauac 16980
uacucacuac aaccaaagcc acggugguuc acgauauugg uaacccaaaa gguaucccua 17040
ucgugcgcaa accugguguu aaauaucacu ucuaugauca agcacccauu gucaaacacg 17100
uucaaaaacu aaaguacaag ccagagaugg aggcccguuu caccgauggu uugacuaugu 17160
uuuggaauug uaauguugac acauacccug cuaacgcccu ugugugccgc uacgacacuc 17220
aucggcagaa gcauuuaauu ggaccuaaug guucagcacu auauguuaau aagcaugcuu 17280
uucucacccc ugagaugcau acuuaugcua cacauaaacu caacuuggcu ccacucaucu 17340
acuacuccac cacagauugu aguagugaac agccuauugu uguuaccuac agagauugug 17400
ucacccggug caauacugga aaaacucucu guccaaauca ugcucuugaa uaucaagagu 17460
uuaucaaugc auacaaucuc auggcucgcc auggauuuaa uguuuacaua ccacgcaaug 17520
ucaacguuua caacuguugg cuuacuuuca cuaaucucca aaaccuugaa aacuuagcuu 17580
acaacuguua uuauaagaac ugcaaugcuc acguugaugg gcagcuugau guaguuauua 17640
auaauaacgc uguauaugcu aaggucgaca auaaucuugu caaacuuuuc gacaaccgca 17700
cuaacuuacc ugucucagug gccuuugaac auuacacuaa caggcauacc cguucacugc 17760
caacuacaca gcuguuaucu gguuuaggcg uaaccgccac cagaaauuuc acuguguggu 17820
ucgacaauga uacaauuuuc caauacacua uuaauguauc uacguauacu gacaucgacc 17880
cuaguaccca uguuguccuc ugugaugaua gguacggaac agauuggagu caguuuaacc 17940
aacuuccuaa ugcaguauuc cucaccaaaa cuaaggugaa gaaaacagaa ccguuuguuu 18000
guacagcacu gacccuaaau ggcgucgcca uugacgguga agagcuauac aucuauguac 18060
gcuauaacaa ucaacugacc acauuugcua cuacuuguac acaggguaga aauguugagc 18120
aguuuauacc uaaaacaccu auggaaagag acuuccuuga gaugucucaa caguccuuca 18180
uccagcaaca ucaauugcag gaacugggug uugaacacau uaucuauggu gaugauucca 18240
guccagucau uggcggaacu cacacacuua ucucacuagu uaaaaacaag uuugaacauc 18300
agcuugucaa ccauguuuac aacccagucc agaacugugu uguuaccuca ccuaacgcaa 18360
gcuccaagaa cguuugcacu guucuugaug uucuacuuga ugacuacauu gacaucauaa 18420
gacaagcaca ugccaguuac acaaguaaau ccaaaguauu cacuguguca auugacaacc 18480
aacaaauuag auucaugcuu uggcaugaug agcaagucaa gacuugcuac ccaaucuuac 18540
agucacuuac caaugguuac cagaugccau cuguguacaa aacauugguu acugacuuac 18600
aaccagcuga caucccuaau uaucauuccu acaccccccg ggugccugga guaguuaaga 18660
auguuaucaa guaccgccaa cuuuucaacu acauaguuaa aaaggauagg uuggcaguac 18720
cacacaauau gaccguauua caccuuggag cugcaucugc acuagguaca gcaccagguu 18780
cuucagucau aaaacaaaug uuuccugaag gaacuguucu uauugaccuc gauauaagag 18840
aguucacuuc agaugcuaac caaauaauag uuacagacua cagaacuuac auaccaccac 18900
accacguaga cgucauauuu ucugaccucu acuguuguga ugacauacac uucuuugaca 18960
aucuaauaag gauaguuaag gagaggcucg cccucggugg uucuaucuuu guuaagauaa 19020
cugaacauuc auucucaccc gaacucuacu cacuugcggg uugguucgau gauuaucaac 19080
uauuuugcac agcagucaau gccucgucuu cagaagcauu uuuaugcugu uuuaauuauu 19140
uggggcuugc uaaggaaaac auuaaugguu uuaacuuaca ugcuuccuau auucaauggc 19200
gcaaugaaau agcguugaca ccaaccuauu cuccuuuagc ggacaacccg gcuacggccu 19260
guaagcuaaa agcaacgccu auuaucucgg cucgugaguu agagaagaag ccuauucuuc 19320
gcuaucucgu ugcaucaggg cgccuucuug ugaggccacc agaaugcaga gagcucuauu 19380
gauuaugacc uuacuuuguc ucguucgagc aaaguuugcu gaugaucuac ucgacuugcu 19440
caccuucccg ggugcacauc gcuucuuaca uaaacccacg aggaauucca gcagucucua 19500
cucgcgggcu aauaauaauu uugauguugg cguucuuccu ggcuacccca cuaagaacgu 19560
uaaccucuuc ucaccacuua cuaacuccac uuugcccauu aauggccuuc aucggaguua 19620
ccaaccacuc augcugaauu gucuuacuaa aauaacuaac cacacucuca gcauguaucu 19680
ccuaccuagu gagauacaaa cuuauagcug cggcggugcc augguuaaau accagacaca 19740
ugaugcaguu cguaucauuu uagaccucac ugccacugac cacaucucug uugaagucgu 19800
ugaccaacau ggugaaaauu auguguuugu uugcagugag caguuuaacu acaccacugc 19860
auuacacaau ucuaccuucu ucucacuuaa uucugcgcuu uauugcuuua cuaauaacac 19920
cuacuuaggu auucuuccac cugauuuaac ugacuuuacg gucuaccgua cuggucaguu 19980
cuaugcuaau gguuaccuuu uagguacuuu accuauuacg guuaacuaug uuagguugua 20040
ucggggucau uugucugcca auagugccca cuuugcccuu gcaaaccuaa ccgauacacu 20100
cauaacacuu accaauacua cuauaucgca aaucacuuau ugugauaagu caguaguuga 20160
uucaauagca ugccagcgcu cuucucacga aguggaggau ggguuuuacu ccgacccuaa 20220
aucugccguu agagcuaggc aacguacuau uguuacacua ccuaagcucc cugagcuuga 20280
aguagugcag uuaaauauuu cugcacacau ggauuuuggc gaagccagac uugacagcgu 20340
uaccaucaau gguaacacau ccuauugugu cacuaagccu uacuucaggc uugaaacuaa 20400
cuuuaugugu acagguugca cuaugaaucu gcgcacugau accuguaguu uugaccuguc 20460
agcaguaaac aauggcaugu cauucucuca auucugucua agcacugaau cuggugcuug 20520
ugagaugaaa auuauuguua ccuacguaug gaauuacuug cuaaggcagc guuuguaugu 20580
uacugcugua gagggccaga cucacacugg aaccacuuca guacaugcaa cagacacuuc 20640
uaguguaauc acugaugucu gcacugacua cacuaucuau ggagucucug guacuggcau 20700
uauuaagcca ucagaucucu uauugcacaa uggcauagca uucaccucuc caacagguga 20760
gcuuuaugca uuuaaaaaua uaaccacugg caaaacccuu caggucuuac cgugugaaac 20820
cccuucucaa cugauuguga uaaacaacac cguugucggu gcuaucacau ccaguaauuc 20880
aacugaaaau aauagguuua cuacuacuau ugucacaccu acuuucuuuu auuccacaaa 20940
ugccaccacu uucaacugca cuaagccugu uuuguccuau ggaccuauca gcguguguag 21000
ugauggugca auugugggaa cauccacauu acagaauacu cgaccaucca uaguuuuacu 21060
auacgauggc gaaguugaaa uaccaucugc auuuucucuu uccguucaga cggaguacuu 21120
gcaaguucaa gcagagcaag uuauaguuga uuguccucag uauguaugca auggcaacag 21180
ccguugucua caauuacugg cacaauacac cucagcuugc ucuaacauug aagcagcucu 21240
gcauuccucu gcacaguugg auagcagaga gauuauaaau auguuucaaa caucaacaca 21300
guccuugcag uuagcuaaua uuaccaacuu caagggugac uacaauuuua gcagcauacu 21360
aaccaccaga auugguggca gaucugcuau ugaagaccuu cuuuuuaaua aaguuguuac 21420
uaguggccuu ggcacuguug aucaggacua caaauccugc ucuagagaca uggccaucgc 21480
ugacuuaguu uguucccagu auuacaaugg caucaugguu cuaccuggug uuguugaugc 21540
ugagaaaaug gcaauguaua cuggcucucu uacuggagcu augguauuug gaggacugac 21600
ugccgcagcg gcaauaccau uugccacggc aguacaagcu cgccucaauu augucgcacu 21660
gcaaacaaau guacuacaag aaaaccagaa aauucuugca gaaucauuua accaagcagu 21720
uggcaauaua ucacuugcac uaucuucugu uaaugaugcc auccagcaaa cuucugaggc 21780
ucuuaacacc guagcuauug cuauuaaaaa gauucaaaca guuguuaacc agcagggcga 21840
ggcauuauca caccugacug cacagcuguc aaacaauuuu caagcaauuu cgacuucuau 21900
ucaagacauu uacaaccguc uugaggaagu agaggcuaac cagcaaguug accgucucau 21960
cacaggacgg uuggcugcac uuaaugcaua uguuacucag uuacucaauc agaugucuca 22020
gauuagacaa ucucgauugu uagcucagca aaagauuaau gaguguguca aaucacaguc 22080
guccagauac gguuucugug gaaauggcac acacaucuuc ucacuuacac agacugcacc 22140
aaauggcaua uuuuucaugc augcagugcu aguacccaac aaauucacac gugucaacgc 22200
uucugccggc auuugugugg auaauacgag aggcuacuca uugcagccuc aacuuauacu 22260
cuaccaguuu aauaacuccu ggagaguuac accuagaaau auguaugaac ccagacugcc 22320
ccggcaggcu gauuucauac aauuaacuga uugcagcguu acuuuuuaca acaccaccgc 22380
ugcuaaucuu cccaauauua ucccugacau uauagauguc aaucaaacag ucagugauau 22440
uauugacaau uuaccuacag caacaccucc ucagugggau guugguaucu auaacaacac 22500
uauucucaac cucaccguug agauuaauga ucuacaagag cggucuaaaa accucucaca 22560
gauugcagau cguuuacaaa auuauauuga caaucuuaac aauacucuag uugaccuuga 22620
auggcucaac agaguggaaa cuuaccuuaa auggccgugg uauauauggc uugccauugc 22680
ccuggcucuu auugcauuug ugacaauccu cauaacaauc uuucuuugua cugguuguug 22740
uggugguugc uuugguuguu guggcgguug uuuuggccuu uucucuaaga agaaaaggua 22800
uaccgacgac caaccaacac cguccuuuaa guuuaaggaa ugguagucga cgacugggcc 22860
guuaccaucc cuggacaaua uauuauugcu auacuaguug ucaucugcau ugguguggca 22920
cuacuuuuua uuaauacuug cuuagcuugu guuaaauuau uuuacaagug cuaccuaggg 22980
gcagcauauc uuguuaggcc uauuauagug uacuacucca agccgaaccc cguaccugag 23040
gaugaguuug uaaaaguaca ccaauuuccu agaaacacuc acuaugucug acgcagaaga 23100
guggcaaauu auuguuuuca uugcgaucau augggcgcuu ggcgucaucc uccagggagg 23160
cuaugccacg cguaaucgug ugaucuaugu uauuaaacuu auucugcuuu ggcugcucca 23220
acccuucacc cuagugguga ccauuuggac cgcagucgac agaucaucua agaaggacgc 23280
aguuuucauu guguccauaa uuuuugccgu acugaccuuc auauccuggg ccaaguacug 23340
guaugacuca auucguuuau uaaugaaaac cagaucugca ugggcacucu caccugagag 23400
uagacuccuu gcagggauua uggauccaau ggguacaugg aggugcauuc ccaucgacca 23460
cauggcucca auucucacac cagucguuaa gcauggcaag cucaagcuac augggcaaga 23520
gcuggccaau ggcauaucag ucagaaaucc gccacaggau auggugauag ugucaccaag 23580
ugacaccuuu cacuacacuu uuaagaaacc uguggaauca aacaacgauc cagaauuugc 23640
uguucugaua uaccagggug accgcgcuuc aaacgcugga cuucacacca uaaccacuuc 23700
aaaggccggu gacgcucgcc uguauaagua uauguaaugu gcaacugcca ucugcagcug 23760
cgagauuuau auagauugug caauaagcgg cacaucagaa gagaggaugu uccugagcuu 23820
auugacccuc ucguuaaaac ucgcuguuuu gcuuacaguc ucgugguucu ugcuaaugcu 23880
aauccaauug cauuuagcau acuaccucgg aaaauucuua ucaaugguga gccuuuacug 23940
cuugaauaug guagcauaua ugguaaagac uuuaucauuc gaccaucgcu ccaagucauu 24000
cuugaagaug aauuaaauua aaguuuugac accaaucuau cauggcugca ccaguagucc 24060
cuacuacuga cgcgucuugg uuucaggugc ucaaagcuca aaacaaaaag gcuacucauc 24120
cucaguuucg uggcaaugga guuccgcuua acuccgccau caaacccguu gaaaaccaug 24180
gcuacuggcu gcguuacacc agacaaaagc caggugguac ucccauuccu ccauccuaug 24240
ccuuuuauua uacuggcaca ggucccagag gaaaucuuaa guauggugaa cucccuccua 24300
augauacccc agcaaccacu cguguuacuu ggguuaaggg uucgggagcu gacacuucua 24360
uuaaaccuca uguugccaaa cgcaacccca acaauccuaa acaucagcug cuaccucucc 24420
gauucccaac cggagauggc ccagcucaag guuucagagu ugaccccuuc aacgcuagag 24480
gaagaccuca ggagcgugga aguggcccaa gaucucaauc uguuaacucc agaggcacag 24540
gcaaucagcc caggaaacgc gaccaaucug caccagcugc gguacgucgu aagacccagc 24600
aucaagcucc caagcggacu uuacccaagg guaaaaccau uucucaggua uuuggcaacc 24660
ggucucguac uggugccaau gucggcucug cagacacuga gaagacgggu auggcugauc 24720
cucgcaucau ggcucuagcc agacaugugc cugguguuca ggaaaugcuu uuugcuggcc 24780
accuugagag caacuuucag gcgggggcaa uuacccuuac cuucuccuac ucaaucacag 24840
ucaaggaggg uucuccugac uaugagagac uuaaggaugc gcucaauacg gucguuaacc 24900
agaccuauga gccaccuacc aaaccaacua aggacaagaa gccugacaaa caagaccagu 24960
cugcuaaacc caaacagcag aagaaaccua aaaagguaac ucugccagca gacaaacagg 25020
auugggagug ggaugaugcu uuugagauaa agcaggaauc agcagcguag acaucaaucu 25080
augucuguua aacccaccca acuccacuca aauaucucuu ugguuccaga gagucguagu 25140
guauagccag agagccaguc agagggcgcu aucaugcaaa cuagggcugg cuacucuagc 25200
acagaaucac aucccgauaa ucaacagugc uagaagguug auuauaccau uuaauaugcc 25260
gaggccacgc ggaguacgau cgaggguaca gcauaaucuc aacuuuuguu gagccacaau 25320
uuuaauccua auuggagaag gccaaaggac uguacuacuu uugugggugu agcagucgcc 25380
cagugggaaa gcgccaacua gguuacaauu guggugggga caaauu 25426
<210> 7
<211> 3480
<212> DNA
<213> Artificial Sequence
<220>
<223> Porcine Deltacorona Virus BIV-Isolate 2.0307 Spike Antigen cDNA
<400> 7
atgcagagag ctctattgat tatgacctta ctttgtctcg ttcgagcaaa gtttgctgat 60
gatctactcg acttgctcac cttcccgggt gcacatcgct tcttacataa acccacgagg 120
aattccagca gtctctactc gcgggctaat aataattttg atgttggcgt tcttcctggc 180
taccccacta agaacgttaa cctcttctca ccacttacta actccacttt gcccattaat 240
ggccttcatc ggagttacca accactcatg ctgaattgtc ttactaaaat aactaaccac 300
actctcagca tgtatctcct acctagtgag atacaaactt atagctgcgg cggtgccatg 360
gttaaatacc agacacatga tgcagttcgt atcattttag acctcactgc cactgaccac 420
atctctgttg aagtcgttga ccaacatggt gaaaattatg tgtttgtttg cagtgagcag 480
tttaactaca ccactgcatt acacaattct accttcttct cacttaattc tgcgctttat 540
tgctttacta ataacaccta cttaggtatt cttccacctg atttaactga ctttacggtc 600
taccgtactg gtcagttcta tgctaatggt taccttttag gtactttacc tattacggtt 660
aactatgtta ggttgtatcg gggtcatttg tctgccaata gtgcccactt tgcccttgca 720
aacctaaccg atacactcat aacacttacc aatactacta tatcgcaaat cacttattgt 780
gataagtcag tagttgattc aatagcatgc cagcgctctt ctcacgaagt ggaggatggg 840
ttttactccg accctaaatc tgccgttaga gctaggcaac gtactattgt tacactacct 900
aagctccctg agcttgaagt agtgcagtta aatatttctg cacacatgga ttttggcgaa 960
gccagacttg acagcgttac catcaatggt aacacatcct attgtgtcac taagccttac 1020
ttcaggcttg aaactaactt tatgtgtaca ggttgcacta tgaatctgcg cactgatacc 1080
tgtagttttg acctgtcagc agtaaacaat ggcatgtcat tctctcaatt ctgtctaagc 1140
actgaatctg gtgcttgtga gatgaaaatt attgttacct acgtatggaa ttacttgcta 1200
aggcagcgtt tgtatgttac tgctgtagag ggccagactc acactggaac cacttcagta 1260
catgcaacag acacttctag tgtaatcact gatgtctgca ctgactacac tatctatgga 1320
gtctctggta ctggcattat taagccatca gatctcttat tgcacaatgg catagcattc 1380
acctctccaa caggtgagct ttatgcattt aaaaatataa ccactggcaa aacccttcag 1440
gtcttaccgt gtgaaacccc ttctcaactg attgtgataa acaacaccgt tgtcggtgct 1500
atcacatcca gtaattcaac tgaaaataat aggtttacta ctactattgt cacacctact 1560
ttcttttatt ccacaaatgc caccactttc aactgcacta agcctgtttt gtcctatgga 1620
cctatcagcg tgtgtagtga tggtgcaatt gtgggaacat ccacattaca gaatactcga 1680
ccatccatag ttttactata cgatggcgaa gttgaaatac catctgcatt ttctctttcc 1740
gttcagacgg agtacttgca agttcaagca gagcaagtta tagttgattg tcctcagtat 1800
gtatgcaatg gcaacagccg ttgtctacaa ttactggcac aatacacctc agcttgctct 1860
aacattgaag cagctctgca ttcctctgca cagttggata gcagagagat tataaatatg 1920
tttcaaacat caacacagtc cttgcagtta gctaatatta ccaacttcaa gggtgactac 1980
aattttagca gcatactaac caccagaatt ggtggcagat ctgctattga agaccttctt 2040
tttaataaag ttgttactag tggccttggc actgttgatc aggactacaa atcctgctct 2100
agagacatgg ccatcgctga cttagtttgt tcccagtatt acaatggcat catggttcta 2160
cctggtgttg ttgatgctga gaaaatggca atgtatactg gctctcttac tggagctatg 2220
gtatttggag gactgactgc cgcagcggca ataccatttg ccacggcagt acaagctcgc 2280
ctcaattatg tcgcactgca aacaaatgta ctacaagaaa accagaaaat tcttgcagaa 2340
tcatttaacc aagcagttgg caatatatca cttgcactat cttctgttaa tgatgccatc 2400
cagcaaactt ctgaggctct taacaccgta gctattgcta ttaaaaagat tcaaacagtt 2460
gttaaccagc agggcgaggc attatcacac ctgactgcac agctgtcaaa caattttcaa 2520
gcaatttcga cttctattca agacatttac aaccgtcttg aggaagtaga ggctaaccag 2580
caagttgacc gtctcatcac aggacggttg gctgcactta atgcatatgt tactcagtta 2640
ctcaatcaga tgtctcagat tagacaatct cgattgttag ctcagcaaaa gattaatgag 2700
tgtgtcaaat cacagtcgtc cagatacggt ttctgtggaa atggcacaca catcttctca 2760
cttacacaga ctgcaccaaa tggcatattt ttcatgcatg cagtgctagt acccaacaaa 2820
ttcacacgtg tcaacgcttc tgccggcatt tgtgtggata atacgagagg ctactcattg 2880
cagcctcaac ttatactcta ccagtttaat aactcctgga gagttacacc tagaaatatg 2940
tatgaaccca gactgccccg gcaggctgat ttcatacaat taactgattg cagcgttact 3000
ttttacaaca ccaccgctgc taatcttccc aatattatcc ctgacattat agatgtcaat 3060
caaacagtca gtgatattat tgacaattta cctacagcaa cacctcctca gtgggatgtt 3120
ggtatctata acaacactat tctcaacctc accgttgaga ttaatgatct acaagagcgg 3180
tctaaaaacc tctcacagat tgcagatcgt ttacaaaatt atattgacaa tcttaacaat 3240
actctagttg accttgaatg gctcaacaga gtggaaactt accttaaatg gccgtggtat 3300
atatggcttg ccattgccct ggctcttatt gcatttgtga caatcctcat aacaatcttt 3360
ctttgtactg gttgttgtgg tggttgcttt ggttgttgtg gcggttgttt tggccttttc 3420
tctaagaaga aaaggtatac cgacgaccaa ccaacaccgt cctttaagtt taaggaatgg 3480
<210> 8
<211> 1158
<212> PRT
<213> Porcine Deltacorona Virus
<400> 8
Met Gln Arg Ala Leu Leu Ile Met Thr Leu Leu Cys Leu Val Arg Ala
1 5 10 15
Lys Phe Ala Asp Asp Leu Leu Asp Leu Leu Thr Phe Pro Gly Ala His
20 25 30
Arg Phe Leu His Lys Pro Thr Arg Asn Ser Ser Ser Leu Tyr Ser Arg
35 40 45
Ala Asn Asn Asn Phe Asp Val Gly Val Leu Pro Gly Tyr Pro Thr Lys
50 55 60
Asn Val Asn Leu Phe Ser Pro Leu Thr Asn Ser Thr Leu Pro Ile Asn
65 70 75 80
Gly Leu His Arg Ser Tyr Gln Pro Leu Met Leu Asn Cys Leu Thr Lys
85 90 95
Ile Thr Asn His Thr Leu Ser Met Tyr Leu Leu Pro Ser Glu Ile Gln
100 105 110
Thr Tyr Ser Cys Gly Gly Ala Met Val Lys Tyr Gln Thr His Asp Ala
115 120 125
Val Arg Ile Ile Leu Asp Leu Thr Ala Thr Asp His Ile Ser Val Glu
130 135 140
Val Val Asp Gln His Gly Glu Asn Tyr Val Phe Val Cys Ser Glu Gln
145 150 155 160
Phe Asn Tyr Thr Thr Ala Leu His Asn Ser Thr Phe Phe Ser Leu Asn
165 170 175
Ser Ala Leu Tyr Cys Phe Thr Asn Asn Thr Tyr Leu Gly Ile Leu Pro
180 185 190
Pro Asp Leu Thr Asp Phe Thr Val Tyr Arg Thr Gly Gln Phe Tyr Ala
195 200 205
Asn Gly Tyr Leu Leu Gly Thr Leu Pro Ile Thr Val Asn Tyr Val Arg
210 215 220
Leu Tyr Arg Gly His Leu Ser Ala Asn Ser Ala His Phe Ala Leu Ala
225 230 235 240
Asn Leu Thr Asp Thr Leu Ile Thr Leu Thr Asn Thr Thr Ile Ser Gln
245 250 255
Ile Thr Tyr Cys Asp Lys Ser Val Val Asp Ser Ile Ala Cys Gln Arg
260 265 270
Ser Ser His Glu Val Glu Asp Gly Phe Tyr Ser Asp Pro Lys Ser Ala
275 280 285
Val Arg Ala Arg Gln Arg Thr Ile Val Thr Leu Pro Lys Leu Pro Glu
290 295 300
Leu Glu Val Val Gln Leu Asn Ile Ser Ala His Met Asp Phe Gly Glu
305 310 315 320
Ala Arg Leu Asp Ser Val Thr Ile Asn Gly Asn Thr Ser Tyr Cys Val
325 330 335
Thr Lys Pro Tyr Phe Arg Leu Glu Thr Asn Phe Met Cys Thr Gly Cys
340 345 350
Thr Met Asn Leu Arg Thr Asp Thr Cys Ser Phe Asp Leu Ser Ala Val
355 360 365
Asn Asn Gly Met Ser Phe Ser Gln Phe Cys Leu Ser Thr Glu Ser Gly
370 375 380
Ala Cys Glu Met Lys Ile Ile Val Thr Tyr Val Trp Asn Tyr Leu Leu
385 390 395 400
Arg Gln Arg Leu Tyr Val Thr Ala Val Glu Gly Gln Thr His Thr Gly
405 410 415
Thr Thr Ser Val His Ala Thr Asp Thr Ser Ser Val Ile Thr Asp Val
420 425 430
Cys Thr Asp Tyr Thr Ile Tyr Gly Val Ser Gly Thr Gly Ile Ile Lys
435 440 445
Pro Ser Asp Leu Leu Leu His Asn Gly Ile Ala Phe Thr Ser Pro Thr
450 455 460
Gly Glu Leu Tyr Ala Phe Lys Asn Ile Thr Thr Gly Lys Thr Leu Gln
465 470 475 480
Val Leu Pro Cys Glu Thr Pro Ser Gln Leu Ile Val Ile Asn Asn Thr
485 490 495
Val Val Gly Ala Ile Thr Ser Ser Asn Ser Thr Glu Asn Asn Arg Phe
500 505 510
Thr Thr Thr Ile Val Thr Pro Thr Phe Phe Tyr Ser Thr Asn Ala Thr
515 520 525
Thr Phe Asn Cys Thr Lys Pro Val Leu Ser Tyr Gly Pro Ile Ser Val
530 535 540
Cys Ser Asp Gly Ala Ile Val Gly Thr Ser Thr Leu Gln Asn Thr Arg
545 550 555 560
Pro Ser Ile Val Leu Leu Tyr Asp Gly Glu Val Glu Ile Pro Ser Ala
565 570 575
Phe Ser Leu Ser Val Gln Thr Glu Tyr Leu Gln Val Gln Ala Glu Gln
580 585 590
Val Ile Val Asp Cys Pro Gln Tyr Val Cys Asn Gly Asn Ser Arg Cys
595 600 605
Leu Gln Leu Leu Ala Gln Tyr Thr Ser Ala Cys Ser Asn Ile Glu Ala
610 615 620
Ala Leu His Ser Ser Ala Gln Leu Asp Ser Arg Glu Ile Ile Asn Met
625 630 635 640
Phe Gln Thr Ser Thr Gln Ser Leu Gln Leu Ala Asn Ile Thr Asn Phe
645 650 655
Lys Gly Asp Tyr Asn Phe Ser Ser Ile Leu Thr Thr Arg Ile Gly Gly
660 665 670
Arg Ser Ala Ile Glu Asp Leu Leu Phe Asn Lys Val Val Thr Ser Gly
675 680 685
Leu Gly Thr Val Asp Gln Asp Tyr Lys Ser Cys Ser Arg Asp Met Ala
690 695 700
Ile Ala Asp Leu Val Cys Ser Gln Tyr Tyr Asn Gly Ile Met Val Leu
705 710 715 720
Pro Gly Val Val Asp Ala Glu Lys Met Ala Met Tyr Thr Gly Ser Leu
725 730 735
Thr Gly Ala Met Val Phe Gly Gly Leu Thr Ala Ala Ala Ala Ile Pro
740 745 750
Phe Ala Thr Ala Val Gln Ala Arg Leu Asn Tyr Val Ala Leu Gln Thr
755 760 765
Asn Val Leu Gln Glu Asn Gln Lys Ile Leu Ala Glu Ser Phe Asn Gln
770 775 780
Ala Val Gly Asn Ile Ser Leu Ala Leu Ser Ser Val Asn Asp Ala Ile
785 790 795 800
Gln Gln Thr Ser Glu Ala Leu Asn Thr Val Ala Ile Ala Ile Lys Lys
805 810 815
Ile Gln Thr Val Val Asn Gln Gln Gly Glu Ala Leu Ser His Leu Thr
820 825 830
Ala Gln Leu Ser Asn Asn Phe Gln Ala Ile Ser Thr Ser Ile Gln Asp
835 840 845
Ile Tyr Asn Arg Leu Glu Glu Val Glu Ala Asn Gln Gln Val Asp Arg
850 855 860
Leu Ile Thr Gly Arg Leu Ala Ala Leu Asn Ala Tyr Val Thr Gln Leu
865 870 875 880
Leu Asn Gln Met Ser Gln Ile Arg Gln Ser Arg Leu Leu Ala Gln Gln
885 890 895
Lys Ile Asn Glu Cys Val Lys Ser Gln Ser Ser Arg Tyr Gly Phe Cys
900 905 910
Gly Asn Gly Thr His Ile Phe Ser Leu Thr Gln Thr Ala Pro Asn Gly
915 920 925
Ile Phe Phe Met His Ala Val Leu Val Pro Asn Lys Phe Thr Arg Val
930 935 940
Asn Ala Ser Ala Gly Ile Cys Val Asp Asn Thr Arg Gly Tyr Ser Leu
945 950 955 960
Gln Pro Gln Leu Ile Leu Tyr Gln Phe Asn Asn Ser Trp Arg Val Thr
965 970 975
Pro Arg Asn Met Tyr Glu Pro Arg Leu Pro Arg Gln Ala Asp Phe Ile
980 985 990
Gln Leu Thr Asp Cys Ser Val Thr Phe Tyr Asn Thr Thr Ala Ala Asn
995 1000 1005
Leu Pro Asn Ile Ile Pro Asp Ile Ile Asp Val Asn Gln Thr Val
1010 1015 1020
Ser Asp Ile Ile Asp Asn Leu Pro Thr Ala Thr Pro Pro Gln Trp
1025 1030 1035
Asp Val Gly Ile Tyr Asn Asn Thr Ile Leu Asn Leu Thr Val Glu
1040 1045 1050
Ile Asn Asp Leu Gln Glu Arg Ser Lys Asn Leu Ser Gln Ile Ala
1055 1060 1065
Asp Arg Leu Gln Asn Tyr Ile Asp Asn Leu Asn Asn Thr Leu Val
1070 1075 1080
Asp Leu Glu Trp Leu Asn Arg Val Glu Thr Tyr Leu Lys Trp Pro
1085 1090 1095
Trp Tyr Ile Trp Leu Ala Ile Ala Leu Ala Leu Ile Ala Phe Val
1100 1105 1110
Thr Ile Leu Ile Thr Ile Phe Leu Cys Thr Gly Cys Cys Gly Gly
1115 1120 1125
Cys Phe Gly Cys Cys Gly Gly Cys Phe Gly Leu Phe Ser Lys Lys
1130 1135 1140
Lys Arg Tyr Thr Asp Asp Gln Pro Thr Pro Ser Phe Lys Phe Lys
1145 1150 1155
<210> 9
<211> 25498
<212> DNA
<213> Artificial Sequence
<220>
<223> Porcine Deltacorona Virus BIV-Isolate 5.0327 Genome cDNA
<400> 9
ctttgagcac ctgaaaccaa gacgcgtcag tagtagggac tactggtgca gccatgatag 60
attggtgtcg gtagagaact agcgaagcta gggagataaa attatagact aatgctataa 120
tttttatctt tagtctataa ttttatctcc ctagcttcgc tagttctcta ccgacaccaa 180
tccaggtgcg tctgccacca agttggctac ccttcctagg ggcgctttcg cgcttgctca 240
ccattagatt acctggaaac cagccattca ggttggagtt tccccaggct cttttgtgtg 300
ggcattagcg gcttgtggtt tttgcacaaa atctaagcta cttaccgttc ctctgaccat 360
ccaccacttc tatagacagc actgattacc gtagggttta agtcacaccg gtctgcaccg 420
cccgtcagcg gacacattac ccagcatagc actccttgca ccgcgcctag gtaggataaa 480
accccctacc gggtgactct taaggcgttt cctccacggg atagccacta gtcactaggt 540
gtaagtgatc tgatctgggc gtattgtgtt gcgcaagtgt gatacccata ggagcgtgga 600
atcctattct gcggctcagt gcctgatata gctgtgaaat ggccaagaac aagtccaagc 660
gcgacgctat tgcgttgcct gaaaatgtac caccacctct gcaacttttc attcatgttg 720
cagctgctga agagggtcac cctaaggtta ctacttacct tggcaactat aacctctatg 780
ccaccaaggc tccgcctggc gtgcaggttc ttagtgctaa aacctctctt actgactttg 840
agaatgtctt tggagctcaa cccaccttgc gatcaattcg taatctggtt tgtgaggctc 900
gctcggctga atggacaact tccaagaatg cttttgcact caaagccact caacttgact 960
actctgatgc cgttttgagg gcaatgattc gtttctgccc tccaaaggtg tccacactcg 1020
ctgcctttgc tctttttggc cgattggtta aaattgagga caaggaactt gctgagttag 1080
ctcgtgatac tgcccttgag ttggcgtaca cggctaaaat tggtacatct cttgctgaca 1140
cgagatctgt ctcacttatt cataaggacg cttatctaac tctcagtaat gaggttgttg 1200
gcgtaacttt tactgccgca cttatggcaa aggctaccac tgttaatgga gcaatgcaat 1260
actcaaactt ttacctctac cctcgtgcca ctattaaggt aaccgatggt aaggctgaag 1320
caattgcaac taagcctctg tctgctgcca ctaaaggcaa gcaaatcaca gaggatgtca 1380
accttctccc tgactatcag cagctgcttg ttgatcaagt gactggcact gaggttaagg 1440
ttggagctct aacctatgtt aagaccactg attcgccacc cctttacttt cccaaagtca 1500
agggtggtgt tattggtatt gcacttaagc agcagggcac tgtggctaag aagctcaatg 1560
tagtcttcca tgctcaacct gatgatgttc tgctagcctt catacaactt cagcaattct 1620
tgaaccgtac ttcggattca agtgttgaaa ttactgattg ccagagttat gaagtatctc 1680
caactgtgac ggtcaaaatt ggcccgtcta aacctgggga tgtcatcgtg gctactgatg 1740
aggaatacct taaatgcttt gaaacccctg aggtaggtag gctctataag gttttccaaa 1800
ctcaatcttg ggctatcatt gagcgtgcct tctccagttt gaagatccgc gtgtccaaag 1860
ctttatcagc atttataagt tttctgcaaa accttgcaga taactttact gcaataagtg 1920
gtgttgtcac tgcactcatt cgtgaactcc aggatcttac cctggatgtg gcgacacgta 1980
tcactaacat acaatttgtt taccgtgccg gtaagcttat tgtcgacacg acaagtgtca 2040
tagctaaact tttccagcca ttttgtgatt ttatatcacc tttccttcgg aaagttgctg 2100
gttttgcaat ttacactgtt ggtaatcgca tgcttatgtt taccagcact ggcacctttc 2160
ttctcacaaa ggcaactact aagatactca ataaggcaaa gtacatcttt gatgtggagc 2220
ctgagtaccc agtagatgta acaacatcca aagttgtagt acatgaagca ctccagcaaa 2280
ccgacactaa gcctactaga gctctggagg ctgttgatgt cgttgttggt aatactgtac 2340
tgcaaatggc tactgatggc actgcgttct acccatcgga tggtacgcac gcctctcttc 2400
caggattcaa agcaggttcg gatgagcttt tcataagctt caactgcgac ctctttgatg 2460
atgagactaa tgctcaaatc aacgaaacac tcgctgcata tgagcttaac caactagtgg 2520
ctccaggtga ttctacaccg cgtcaaattg cgacgttggt tgtcgataca cttgcagatg 2580
ctataacaga ccactttccg gagaaaacca ttgatctacc tgaagactat caagtctttt 2640
ctgaccatga tgacctccca ctcgcacaat accacatccc tgatcacctg agcctgtata 2700
ttcaggctat ggaaggtgaa gatgatagtg gtgatgaaat atgtattgag gacgatgatt 2760
acgactgtcc tcaagccgac gaagacacag aaggagtaat tccccaacag tgggaacttc 2820
ctgatgttga taaattttta ctcaagatcc aggaacggaa gaccagcagc gacgaagtac 2880
ttagcgtcga cgtctatcct aaaccagagc cggtcggcaa tgttgggatt gacgacagcg 2940
cgtcggaaaa gaagccaaat ggggactcag taccggatcc tgaggtccat ccaacactag 3000
agagtgtgga tgttgaacga ccaaccgaaa cagcaaacca ggctgttgaa gacaaacctt 3060
ctgataccac ctttgtggtt gatgaggaac aattacaaga atcaacacca gaacatgaac 3120
tccgctccta tgaaggggag tttgattctg atgatgaaat tattattcct atagtaccag 3180
taacacctgc ggatttaaaa ccacagacta ttactataaa ggagtacttt aagtctgaaa 3240
aacttgagac tattaacgaa ggatccacag agtcagttac acaatctgac gattcgtttg 3300
acgagtcatt tgttgatgct gagtctgatg atccacaaga tcctgctgta tatgatgata 3360
caacaattat aacggacagc actgatgtag gcgatgagcc tgagacaact ctagctacca 3420
tcgttaacac acctctgaca ctcgataata acttgccacc tgaagccatt aaacaaccca 3480
gcccaactaa ggttgagtta gttgttggtg aattggcgag tattaaattt gacaattctg 3540
ttctagtcaa ccctgctaat gcgcaattaa caaatggcgg tggagctgct cgtgcaattg 3600
caaaattagc tggtccaaaa tatcaagagt actgtaatag tgtggctcct atctcaggac 3660
cgcttaccac ggactctttt gatgccaaga aatttggtgt agcctgcatc ttgcatgtag 3720
tgccacccaa aggttctgac cctaatgtac aagaactcct gtatcaagct tacaagagta 3780
tccttactga accagcacac tatgttatac ctatactagg tgctggtatc tttggatgca 3840
acccagtcca ctctctggat gcgttcagga aagcatgtcc aagtgacata ggtcgtgtca 3900
cccttgtcac tatgaacaaa aaccatttgc aggtgtggga tgctctcaat aggaccattg 3960
tacgcaccac tactgactat gatcaagtta ccaccaaggc ccttacaccc cagggagtgt 4020
tagaagccaa tctctttgat ggtgaggact ttgttcaaga accaaaaccc ggtcaaatct 4080
accttgaggt tactgaagaa gttcagaacc aagccaagga acttgacctt aaccttcagc 4140
aatactgcgt ctacctgaag acttgccacc ataaatgggt tgtgagtcgt acgaacgggt 4200
tgatgcatct aaaacaaaaa gataacaatt gttttgttag tgcaggtgta aacctgtttc 4260
aaaacactgc ttatcaactt agacctgcta ttgatgctct ctatagggag tatcttaatg 4320
gtaatccaaa tagatttgtt gcttggatct acgcatccac taaccgtcgt gttggtgaga 4380
tgggttgtcc acagcaagtt atttctttgc tcgttagtaa ctctgacgca gcattttcag 4440
caactacagc ctgttgtaac acctacttta accacacagg tgttatttca gtagctcgtg 4500
aatatgaccc aatacaacca aaggtctact gcatgaagtg tgatgtgtgg actcccttta 4560
caccccagag tggaaaaggt gcagttgcaa ttggtatttc tgcagatgaa cctaccggtc 4620
ctgccattaa atttgccgca gctcactgct ggtacactaa tggcaagaaa acagttaatg 4680
gctatgacac taaagctaat gttgtagcta cctatcatag gtttgacgtg cctaagcctc 4740
aacttgtcga ggacgtggtt gcgctgccta ctaaaaatga ctttgaagtt ctcaatgttg 4800
aagaactgcc gcaggatagt gtgctccatt tggacccacc tcctgtacag gccttacaac 4860
ctaaggctaa ccaacacatt gagattctag aaaacccaga ttatctggac attttggatc 4920
tttggattcg taaacccaaa ttcatcctcg taaagtcgtg gagtgttttg ggtagagcac 4980
tatgtaaggc aggtaaagtt gtctttgtca gtgcttcgct tttgacccgt ttctacaatt 5040
accttgtaga gattggtgct cttgactcaa caataaggtt gtcagtcgat cttacctgta 5100
aatttgttag aacggttctc ccatcgtcta acactgtaca caaaacttgt cttggtctgt 5160
attattcagc ccagacactt tttgtttctt tagcaccatt ccttatgtta ccagctgtag 5220
ttagtctgct taattcaggc tatacaattg gcacatattt gtatgcaaaa actggctggc 5280
cttgtaatta caatgccacg caacactttg attataattc ttactgtgca ggtgacttgg 5340
tttgtcaagc ctgttttgac ggtcaagact ccctacattt gtatccgcat ttacgtgtta 5400
atcagcagcc ccttcagacc actgactaca ctgtttatgc gctttcacta atactactat 5460
tagctaacat gactcttgtc atgggcacgc taatagttac tttctttgtg aacttctatg 5520
gtgtgcaaat accattttat ggtacacttt tgatagatta tcaatccgca ctggtgatta 5580
ctttctcagt gtactacttt tataaggtaa tgaagttttt ccgccatctc acacatggat 5640
gtaaaattcc aacgtgtgtg gtatgtgcca aacttcgtac cccacctact ataacagttg 5700
agactgtcgt tcagggcagg aaatacccat ctgttattga aacaaatggc gggtttacaa 5760
tttgtaaaga acacaacttc tattgcaagg actgctcttt acaaacaccc ggcactttca 5820
tcccgacaga agctattgag tcgctctcac gagctaccag gcttagtgtc aaaccaacag 5880
caccagcatt cttacttgct agagatgttg agtgccaaac tgatgttgtc gttgctcgcg 5940
caatgcataa ccaaaatgcg catgtgtgca tttcaaaata ctctgatatc cgtaccgttg 6000
accaactact taagcctact ccactgtttt catacactcc cgatgttatc atcgcggcag 6060
actttgacaa cagaggtagt cttaagacag ctaaagaatt agctgtggtt ttgtcaatgg 6120
accttaaacg tactataatt atcattgatc aggcctattc tagacctatt gataattatc 6180
aggaagttgc ttctcgtatt gagaagtatt acccagttgc aaagatcaca cccacaggtg 6240
acatctttac agacattaag caagcgacca atggccaagc tagtgactct gctattaatg 6300
cagctgttct ggctgtccag cgcggtcttg attttacaat tgacaaccct aacaacatat 6360
taccacatta cgcctttgac ttttcaaccc tcaatgcaga agaccagtct accattttgg 6420
agagtggttg tgctaaaggc aatctcaagg gcactaatgt tggtgttgtt ctttcagcta 6480
gccttgttac acgtcttagt cagcaggcta tacgtgtgat tgctaatgct gcttcacgta 6540
atggtgttac atgcgctgtt actccttcta cacttgttat gcgtgggaat attgcaacac 6600
agcccttgac tcgcatcaaa gctggtgcac ctcccatgcg tcaaaaaatt ttatgtgtta 6660
tcctggcact tgctattgtg tactttgctg ctatggcttt tggctttttg gcaagtcaaa 6720
ttacgcttaa tacagtgcct acgattaaat ctgatatccg cgcctctacc ttctacgttg 6780
ttagagatgg agtcttggat actgttcgtt caaatgacaa gtgctttgca aataagtttt 6840
tggcatttga tagcttcatt caagcacctt acactaattc acctgactgt ccagttgttg 6900
tgggagttgt tgatgtaacg acgcactcta ttcctggaat tccagcaggt gtcattcata 6960
gagacggtct catacttaac atttatgaac agtctcttta tgaaactcat cagcgtcagt 7020
ctatggttag ggatgcgttg tcactcaaga cagcaaatct ctttaaccta ggcaagcgtg 7080
ttgtagtagg atacactcaa catgaagttg ttgtgggtac ctcctatttt aattctcctg 7140
cactttttaa tgcaaagtgc accttcttac agtatcagga cactagacaa ctctattgct 7200
atgatactgt tcctactgaa cataagctct actctgatgt gcttccgcac gtcgagtata 7260
aggctattga cattaatggt gatcttgttc ctttcaagat accagagcag ataatgttct 7320
atccacatat tgtgcgctat actagcaatt cctattgccg tatggggcat tgttttaata 7380
ctaaccctgg tatttgcatt tcatttacgg acgaatttcc gtatagtgaa aatgtcaaac 7440
ctggtgtgta ctgtgctgat acctctttgc agttgttttc aaacctcgtt ttgggcactg 7500
tatctggtat tcacatcttt acatcaacag ctgcattgct tggatctact attgtgatca 7560
tactatgcgt tgttgctgtt cttgcagttc agcgattctt caaggagtac acaacttttg 7620
ttatgtacac ttgtggtctt gctcttgtca acattgtagg cattgcactt atgtacaagt 7680
gccttgtctt cgcgattttc tattatgcaa tctaccttta ctttgtcctt actttcccct 7740
cctttaagag gaatgtggca ttgttttact tcgctgtagt gatcgtgccg cacgtgagta 7800
acatgcaatt gcttgcgctc attgtgtgta gcattatcta ctttctctac acctatgttc 7860
atactgtagc taagacagct gggaaatttt cttccttctt agacgcagct aaagctactt 7920
ttgtcattga caatgaaaag tacgtgttgc ttaaagacct cgctggtgct gaatttgacc 7980
agtatctggc ctcttacaac aagtacaaat atttttctgg tactgcttct gataaggatt 8040
atgataaggt ctgtatggca tttcttgcca aggctttgtc atcttttcgt gaaggaggcg 8100
gttcacagtt gtacacacca cctaaatttg cagttgttca gagtcttaag accaagctgc 8160
aagcaggtat caaaatcctc ctgcaccctt caggtgtagt tgagcgatgt atggtctcag 8220
ttgtctacaa tggatctgca ttgaatggca tctggcttaa gaatgttgtc tactgcccac 8280
gccatgtaat tggaaaattc cgtggtgacc agtggactca catggtctca attgctgatt 8340
gccgcgactt tatagtcaag tgtccaatac agggtattca gctaaatgtc caatcagtta 8400
agatggtagg agctctcctc cagttaactg ttcataccaa caacacagcc actccagact 8460
ataagtttga aaggctacaa ccaggatcat cgatgacaat tgcttgtgct tatgatggca 8520
ttgtacggca tgtctatcac gtggtcctcc aacttaataa tcttatttat gcaagcttcc 8580
ttaacggagc ttgtggtagt gtgggttaca ctcttaaggg taaaacactc tacttacatt 8640
acatgcacca cattgagttt aacaacaaaa ctcatagtgg tacagatctt gaaggtaact 8700
tctatggccc ctatgtggat gaggaagtta ttcagcaaca aacagcattc cagtattaca 8760
ctgataatgt tgttgctcaa ttatatgcac acttactgac tgttgatgct agaccaaaat 8820
ggctggcaca atctcagata agtatcgagg atttcaactc atgggctgct aacaattcct 8880
ttgctaactt cccatgtgaa caaactaata tgtcctacat tatgggactc tcgcaaacag 8940
ctcgagtccc tgtagaacgt atcctcaata ccattataca gctaaccacc aatagagatg 9000
gtgcttgtat tatgggatct tatgatttcg agtgcgattg gacgccagag atggtataca 9060
atcaggctcc aatttcattg cagtcaggag tagttaagaa aacttgtacg tggttcttcc 9120
acttcttgtt tatggctatt accatgctac tcgctgccat gcatgttttc cctgtacact 9180
tgtatccaat agtactgcca tgcttcactg tcgtggcatt cctgttgact ttaaccatta 9240
aacacactgt tgtgtttacc actacatact tgcttccgtc acttttgatg atggttgtaa 9300
atgctaacac tttttggata ccgaacacat ttctgcgcac ctgctacgaa actatattcg 9360
gttccccaat tgctcagcga ctgtatggtt acactgttgc tctttatatg ctgatctatg 9420
ctggacttgc aatcaactat acgttgaaaa cactccggta tagagcaact tcattcttat 9480
ctttttgcat gcagtggttt caatatggtt atgttgcaca cattgcgtac aaactgctta 9540
ataaaccctg gacagaatca ctactcttca cagccttcac aatgctaacc agtcatccct 9600
tgttggctgc tcttagctgg tggctagctg gtcgcgtaac tctgcccatt atcatgcctg 9660
acttagctat tcgtgttttg gcgtataacg tcattggcta tgtcatatgt gttcgatttg 9720
gccttatgtg gcttgcaaat cggttcacaa ctgtacctat gggcacatac cagtatatgg 9780
tgtctgtaga gcaacttaag tacatgatgg cagttaagat gtccccaccg cgtaatgcgt 9840
ttgaggtgct tatagccaac attagacttc ttggtttggg tggaaaccgt aacattgctg 9900
tttctactgt ccaaaacaaa attcttgatg caaaagctac tgctgttgtt gttgctaacc 9960
ttcttgaaaa ggctggcgtc acaaacaagc acgctatttg caaaaagatt gtgaaactcc 10020
acaatgatac ccttaaagcc accacttatg aggaggttga ggtagcactt gtgaaacttc 10080
tttctcacat aattgagttc ttgccaactg atcaggtaga tgcttatcta gctgatgcgg 10140
ccaatgctca acatgttaat acctatttag acaacttgct tgagaacaaa gctgttgttc 10200
aggctgttgc cgatatcaac attaatctgg attcttatag aatttataag gaggcagatg 10260
ctatttataa acgatctgtt gagatgaacg aatctccgca ggagcaaaag aaaaagctta 10320
aagctgttaa cattgcaaag gcggaatggg agcgtgaggc tgcttctcag cgtaagcttg 10380
aaaagcttgc tgatgctgct atgaagtcta tgtatcttgc agaacgtgct gaggatcgtc 10440
gcattaagct aacctctgga cttactgcaa tgctttacca tatgcttaga cgtcttgact 10500
cagatagggt aaaagctctg tttgagtgcg ctaaggcaca aatcttgcca atacatgctg 10560
tagtcggaat ttctaatgac aaccttaaag ttatttttaa cgataaggac agctactctc 10620
attatgtaga gggcaacaca cttatacata agggagttcg ctacactatt gtgaagaaac 10680
tctccttaga taatgcacct attgaaggcg taccagaaga attccctgtg gtcgttgaga 10740
ctgttaggga aggtgtgccc cagttgcaaa ataatgagct atgtttgcgc aatgttttca 10800
ctgctcagaa cacagctcag gacttcaatg gcaatgaatc cactgtaaaa tctttttatg 10860
ttactagaac cggtaagaag attttggttg ccattacatc aactaaagac aatcttaaga 10920
ctgtgacctg ccttactgag accggtaaga cagtccttaa cttggacccc cctatgcgct 10980
tcacacatac cgtaggtgga aaacagtctg ttgtctatct ctattttatt cagaatatta 11040
gttcactcaa cagaggtatg gttattggcc acatctctga aactactatc cttcaggcaa 11100
gtggcactca aattgagtac cagcaaaatg cctctctttt gacctatttg gctttcgctg 11160
tagaccctaa gacagcctac cttaagcatc ttgctgatgg tgggtctcct atacagggtt 11220
gtattcagat gattgctact atgggtcctg gatttgcagt tactactaaa ccacaaccta 11280
atgagcatca gtattcttat ggtggtgctt caatttgtct ttattgccgt gctcatatac 11340
cacatcctgg tgttgatgga cggtgcccct acaaaggccg ctttgttcac atcgacaaag 11400
ataaggaacc tgtttccttc gctttgactc atgagccatg cagttcttgt caacggtggg 11460
ttaattatga ctgcacctgc ggatctagtc tgcagaattc ggcttattta aacgcgtaac 11520
gggttctagt gacgcccggc tagaacccct gcagcctgga actcaaccag atgctgtaaa 11580
aagggccttc catgtgcata atgataccac ctctggtata ttcttaagca caaaatctaa 11640
ctgcgctcgg tttaaaacca cacgcagtgc cctgccttta cctaataagg gagaggttga 11700
attgtacttt gttactaagc agtgtgcagc taaagtcttc gaaatcgagg aggaatgcta 11760
caacgctctt agtacagagc tttatactac tgatgataca tttggtgtcc ttgccaaaac 11820
tgagttcttt aagtttgaca agatacctaa tgtcaatcgc cagtatctga ctaaatatac 11880
actcctggac ttggcttatg ctctacgtca tttgtcaaca tctaaggatg ttattcaaga 11940
aatcttgatc accatgtgcg gaacccctga agattggttt ggggaaaatt ggtttgatcc 12000
aattgagaac ccatcctttt acaaggagtt ccataaactt ggggatattc ttaaccgttg 12060
tgttcttaat gccaataagt ttgctagtgc ctgtatagac gctggtcttg ttggcatatt 12120
aacacccgac aaccaagacc tcctgggtca gatctatgac tttggagatt ttattattac 12180
acaaccaggt aatggatgtg tggacttagc atcctattat tcttatttaa tgcccattat 12240
gtccatgact cacatgttaa agtgtgagtg tatggatagt gatggcaacc cacttgagta 12300
tgatggattt cagtatgact tcacggactt caagcttggc ttgttcgaga agtattttaa 12360
gtactgggac cgtccttacc atcctaacac tgttgaatgt ccagatgacc gttgcgtatt 12420
gcactgtgcg aacttcaatg tgttgtttgc tatgtgtata cctaatacgg catttggcaa 12480
tctttgttca agagctactg ttgatggcca ccttgtggtc cagacagtgg gtgtacactt 12540
gaaagaacty ggtatagtcc ttaaccagga cgttaccaca cacatggcaa atattaatct 12600
aaacactcta ttgcgattgg ttggtgatcc caccactatt gcaagtgtct cagacaagtg 12660
tgtagattta agaactcctt gtcagacctt ggctactatg tctagcggaa ttgctaaaca 12720
gtcagtcaag cccgggcatt ttaatcaaca cttctacaag catttgcttg atagtaacct 12780
attagaccaa cttggaatag acattcgcca cttctactat atgcaggatg gtgaagcggc 12840
tatcacagac tacagctact acaggtataa tacccccacg atggtagata tcaagatgtt 12900
cttattttgc cttgaggtgg cagataagta tcttgagccc tacgaaggtg gatgtattaa 12960
tgcacagtca gttgtggtct ctaatttgga caagtcagcg ggctacccct ttaacaagct 13020
aggtaaggct cgtaactatt acgacatgac tcatgccgag caaaatcaac tgtttgagta 13080
tacaaaacgc aatgttttgc ctacactcac tcagatgaac cttaagtatg caatttcagc 13140
caaggatcgt gctcgcactg tggcaggagt gtctataatt agcaccatga ctaacaggca 13200
gtaccatcaa aagatgctga aatctatttc acttgcacgc aatcagacca tcgtgattgg 13260
aacaaccaaa ttctatggtg gttgggacaa catgttacga cgactgatgt gtaatatcaa 13320
caatcccatt ttagtgggtt gggattaccc taagtgtgat cgttctatgc caaacatgct 13380
gcgcattgcc gcttcgtgct tgctagcacg aaaacacact tgctgtaacc aaagccagcg 13440
attctaccgt ttggctaatg aatgttgcca agtactatct gaagtggtag tctctggtaa 13500
caacctctat gtaaaaccag gtggcactag cagtggtgat gcaaccacag cttatgccaa 13560
ctcggtattt aacatcttac aggtggtttc tgctaatgta gccaccttct tatcaacttc 13620
caccacgaca catcttaata aggacattgc ggacttgcat cgtagtcttt atgaagatat 13680
ttatcgtggt gactctaatg atatcaccgt catcaataga ttctaccagc atctccaaag 13740
ttactttgga cttatgatat tgtctgatga tggtgtcgca tgcatagact cagccgttgc 13800
aaaggctgga gctgttgctg atcttgatgg tttccgagac attttgtttt accaaaacaa 13860
tgtttacatg gcagactcaa agtgttggac agaaactgac atgaatgttg gccctcatga 13920
attttgctca cagcatactg tgttagcaga gcatgatggt aaaccttact acttacctta 13980
cccagatgtc tctcgcattc tgggtgcatg catctttgtg gatgacgtta acaaggctga 14040
ccctgttcag aaccttgaac gttacatctc acttgcaatt gatgcatatc ccctcaccaa 14100
ggttgaccct attaagggta aagtctttta tttgttacta gactacatac gtgttcttgc 14160
tcaggagtta caggacggta tccttgatgc tttccaatca ctcactgaca tgtcgtatgt 14220
aaataacttt atgaatgagg ccttttatgc tcagatgtat gagcaaagtc ctacactaca 14280
ggccagcggt gtttgtgtgg tgtgtaattc acccactata ctgcgctgtg gtgattgcat 14340
tcgtcgacca ctactttgtt gcgtctgtgc ctaccagcat gttacgcaga ctacacataa 14400
acgtatcatt gctatcaaca actacatctg tagtgttgag aattgcaatg aggacaatgt 14460
tgaaaaactt ttcatttctg gcactgcgat ttattgtgag aatcacaaac ccacgctgtg 14520
catacccatt gtagctaatg gttctgtttt tggtatctat cgccacactg cccgtggtag 14580
tgatgacata gacctcttta acgagcttgc tacatctaac tatgacacta ttgaacctta 14640
tcagaaggcc aatcgtgcac ctttatcact tatgctcttc gctgctgaaa ccattaaggc 14700
actcgaggag tctatcaaga agtcatatgc taccgcaacc gtcaaggatg tgtatgacca 14760
acgcttcatt aaacttctat gggaacaggg taaaaagccg ccacccataa cgaagaacca 14820
cattttcact ggctaccatt ttaacaagaa tggaaaaacc caagttggtg attacattct 14880
tgctaaaaca gatggcagtg acacttatac ttacagagga acatctacct acaaactcca 14940
aacaggtgat gttctagtct taatggcaca tgttgttaca ccgctctcag caccccctgt 15000
gttaacgcag acaacatatg tcagaaaatc acttttaccc gactctgttg gtgcgtctta 15060
ttatgtgcaa cattttaagt catataatga gatagctatg cagagggtta caacagtatt 15120
aggtccacca ggcacaggta agtcaacctt tgctattggt ttggctaagt actttcccag 15180
tgcacgtatt tgctacactg cgtcttcgca tgcagcaatc gatgcactct gtgaaaaagc 15240
tttcaagaca atacctgtag gccaatgcag tcgtatcgta cccacacgta caactgttga 15300
gtgctttcag gagtttgtcg taaataacac aactgcacag tatatcttct cgactatcaa 15360
tgccttacct gacattaagt gtgacattgt agttgtagat gaggtttcta tgttgaccaa 15420
ttatgagctt tcctctgtga atgctcgttt ggtttacaat cacattgtgt atgttggtga 15480
tccttatcag ttaccttcac ctagaactat gcttacgtct ggccagcttt cgccagctga 15540
ctataacgta gttactgata taatggtaca tgcaggagcg gatgttatgc tcgacatgtg 15600
ctacagatgc ccacgtgaaa tcgttgagac agtgtctaaa cttgtctacg ataacaaact 15660
aaaagcggcg aaaccgaact caagacagtg ttacaagacc attgtgaact ttggtcctgg 15720
agacgttgct catgagggac aatctgccta caacgaagca cagttgcgtt tcgcactcgc 15780
atttagacaa caaaagcggt gggataacgt gactttcata tctccatata atgctatgaa 15840
tgtgaaagca tccttagcag gtttctctac tcagaccgtt gactcttctc aaggttctga 15900
gtatgattat gttatctttt gcgtgaccac tgattcagca cacgcactta acatggctcg 15960
tttgaacgtt gcccttacac gcgcaaagat aggtatcctt gtggtgttta ggcaggcaaa 16020
cgaactttac aatagtttgc agtttgaatc tattgattca cagcttcagt cgagtgctga 16080
gaaaaacctc acaccactgt ttaagcgctg cggctatgag tataatggcg tccatccagc 16140
tcatgctttg acctggcatg attgtggtgc agagtaccgc tgtgaggagc cacttgctaa 16200
attagtagga gttgccgatg gcactcttat atcatacaaa accctagtat ccacacttgg 16260
gtttcttcca tcacttaaaa ttgatgcata tcataatatg ttcctaacac gtgacgcgtg 16320
tcgcacctat gttcagagtt ggatcggcat agatgttgaa gcagcacacg ccataaaacc 16380
taacaccggg actaacctgc cattgcaaat aggttttagt accggaaaga atttttcagt 16440
cactccagag ggaatttggg taaacgagca cggatcttgc actgagcccg tccctgccaa 16500
aatacctcct ggagaacaat ttcgtcacct taaaaaggac atgcgccagg cgcgtccttg 16560
gaaggttgtt cgacgtgaga ttgctactca cattgctgag gtagctcctc acactgatta 16620
tatatgcttt gtcacttggg ctcaccagct tgagctagcg acaatgcgct actttgtcaa 16680
actaggtatg gaagagaaat gcttttgtgg caggcgggct tgtttcacta atggaactga 16740
gttcgcttgc aaagcacacc attctctcac cattccacaa tgtgattatg tgtacaatcc 16800
attcctcatc gacgtggcta cgtggggatt ctcgggacgg ctttccacca accatgacgc 16860
tgtgtgcaca tatcatgcta atgcccatgt tgcatcagct gatgcaatca tgacggtatg 16920
tttagctatc catgaactgt tcagtactgt tgactggaac tttgaatttc cagtaactgc 16980
tgagcaatcg caacttaaca aggcctgtcg cttagtacag gcgaattact taaatatact 17040
actcactaca accaaagcca cggtggttca cgatattggt aacccaaaag gtatccctat 17100
cgtgcgcaaa cctggtgtta aatatcactt ctatgatcaa gcacccattg tcaaacacgt 17160
tcaaaaacta aagtacaagc cagagatgga ggcccgtttc accgatggtt tgactatgtt 17220
ttggaattgt aatgttgaca cataccctgc taacgccctt gtgtgccgct acgacactca 17280
tcggcagaag catttaattg gacctaatgg ttcagcacta tatgttaata agcatgcttt 17340
tctcacccct gagatgcata cttatgctac acataaactc aacttggctc cactcatcta 17400
ctactccacc acagattgta gtagtgaaca gcctattgtt gttacctaca gagattgtgt 17460
cacccggtgc aatactggaa aaactctctg tccaaatcat gctcttgaat accaagagtt 17520
tatcaatgca tacaatctca tggctcgcca tggatttaat gtttacatac cacgcaatgt 17580
caacgtttac aactgttggc ttactttcac taatctccaa aaccttgaaa acttagctta 17640
caactgttat tataagaact gcaatgctca cgttgatggg cagcttgatg tagttattaa 17700
taataacgct gtatatgcta aggtcgacaa taatcttgtc aaacttttcg acaaccgcac 17760
taacttacct gtctcagtgg cctttgaaca ttacactaac aggcataccc gttcactgcc 17820
aactacacag ctgttatctg gtttaggcgt aaccgccacc agaaatttca ctgtgtggtt 17880
cgacaatgat acaattttcc aatacactat taatgtatct acgtatactg acatcgaccc 17940
tagtacccat gttgtcctct gtgatgatag gtacggaaca gattggagtc agtttaacca 18000
acttcctaat gcagtattcc tcaccaaaac taaggtgaag aaaacagaac cgtttgtttg 18060
tacagcactg accctaaatg gcgtcgccat tgacggtgaa gagctataca tctatgtacg 18120
ctataacaat caactgacca catttgctac tacttgtaca cagggtagaa atgttgagca 18180
gtttatacct aaaacaccta tggaaagaga cttccttgag atgtctcaac agtccttcat 18240
ccagcaacat caattgcagg aactgggtgt tgaacacatt atctatggtg atgattccag 18300
tccagtcatt ggcggaactc acacacttat ctcactagtt aaaaacaagt ttgaacatca 18360
gcttgtcaac catgtttaca acccagtcca gaactgtgtt gttacctcac ctaacgcaag 18420
ctccaagaac gtttgcactg ttcttgatgt tcttcttgat gactacattg acatcataag 18480
acaagcacat gccagttaca caagtaaatc caaagtattc actgtgtcaa ttgacaacca 18540
acaaattaga ttcatgcttt ggcatgatga gcaagtcaag acttgctacc caatcttaca 18600
gtcacttacc aatggttacc agatgccatc tgtgtacaaa acattggtta ctgacttaca 18660
accagctgac atccctaatt atcattccta caccccccgg gtgcctggag tagttaagaa 18720
tgttatcaag taccgccaac ttttcaacta catagttaaa aaggataggt tggcagtacc 18780
acacaatatg accgtattac accttggagc tgcatctgca ctaggtacag caccaggttc 18840
ttcagtcata aaacaaatgt ttcctgaagg aactgttctt attgacctcg atataagaga 18900
gttcacttca gatgctaacc aaataatagt tacagactac agaacttaca taccaccaca 18960
ccacgtagac gtcatatttt ctgacctcta ctgttgtgat gacatacact tctttgacaa 19020
tctaataagg atagttaagg agaggctcgc cctcggtggt tctatctttg ttaagataac 19080
tgaacattca ttctcacccg aactctactc acttgcgggt tggttcgatg attatcaact 19140
attttgcaca gcagtcaatg cctcgtcttc agaagcattt ttatgctgtt ttaattattt 19200
ggggcttgct aaggaaaaca ttaatggttt taacttacat gcttcctata ttcaatggcg 19260
caatgaaata gcgttgacac caacctattc tcctttagcg gacaacccgg ctacggcctg 19320
taagctaaaa gcaacgccta ttatctcggc tcgtgagtta gagaagaagc ctattcttcg 19380
ctatctcgtt gcatcagggc gccttcttgt gaggccacca gaatgcagag agctctattg 19440
attatgacct tattttgtct cgttcgagca aagtttgctg atgatctact cgatttgctc 19500
accttcccgg gtgcacatcg cttcttacat aaacccacga ggaattccag cagtctctac 19560
tcgcgggcta ataataattt tgatgttggc gttcttcctg gctaccccac taagaacgtt 19620
aacctcttct caccacttac taactccact ttgcccatta atggccttca tcggagttac 19680
caaccactca tgctgaattg tcttactaaa ataactaacc acactctcag catgtatctc 19740
ctacctagtg agatacaaac ttatagctgc ggcggtgcca tggttaaata ccagacacat 19800
gatgcagttc gtatcatttt agacctcact gccactgacc acatctctgt tgaagtcgtt 19860
ggccaacatg gtgaaaatta tgtgtttgtt tgcagtgagc agtttaacta caccactgca 19920
ttacacaact ctaccttctt ctcacttaat tctgagcttt attgctttac taataacacc 19980
tacttaggta ttcttccacc tgatttaact gactttacgg tctaccgtac tggtcagttc 20040
tatgctaatg gttacctttt aggtacttta cctattacgg ttaactatgt taggttgtat 20100
cggggtcatt ttagtgccca ctttgccctt gcaaacctaa ccgatacact cataacactt 20160
accaatacta ctatatcgca aatcacttat tgtgataagt ctgtagttga ttcaatagca 20220
tgccagcgct cttctcacga agtggaggat gggttttact ccgaccctaa atctgccgtt 20280
agagctaggc aacgtactat tgttacacta cctaagctcc ctgagcttga agtagtgcag 20340
ttaaatattt ctgcacacat ggattttggc gaagccagac ttgacagcgt taccatcaat 20400
ggtaacacat cctattgtgt cactaagcct tacttcaggc ttgaaactaa ctttatgtgt 20460
acaggttgca ctatgaatct gcgcactgat acctgtagtt ttgacctgtc agcagtaaac 20520
aatggcatgt cattctctca attctgtcta agcactgaat ctggtgcttg tgagatgaaa 20580
attattgtta cctacgtatg gaattacttg ctaaggcagc gtttgtatgt tactgctgta 20640
gagggccaga ctcacactgg aaccacttca gtacatgcaa cagacacttc tagtgtaatc 20700
actgatgtct gcactgacta cactatctat ggagtctctg gtactggcat tattaagcca 20760
tcagatctct tattgcacaa tggcatagca ttcacctctc caacaggtga gctttatgca 20820
tttaaaaata taaccactgg caaaaccctt caggtcttac cgtgtgaaac cccttctcaa 20880
ctgattgtga taaacaacac cgttgtcggt gctatcacat ccagtaattc aactgaaaat 20940
aataggttta ctactactat tgtcacacct actttctttt attccacaaa tgccaccact 21000
ttcaactgca ctaagcctgt tttgtcctat ggacctatca gcgtgtgtag tgatggtgca 21060
attgtgggaa catccacatt acagaatact cgaccatcca tagtttcact atacgatggc 21120
gaagttgaaa taccatctgc attttctctt tccgttcaga cggagtactt gcaagttcaa 21180
gcagagcaag ttatagttga ttgtcctcag tatgtatgca atggcaacag ccgttgtcta 21240
caattactgg cacaatacac ctcagcttgc tctaacattg aagcagctct gcattcctct 21300
gcacagttgg atagcagaga gattataaat atgtttcaaa catcaacaca gtccttgcag 21360
ttagctaata ttaccaactt caagggtgac tacaatttta gcagcatact aaccaccaga 21420
attggtggca gatctgctat tgaagacctt ctttttaata aagttgttac tagtggcctt 21480
ggcactgttg atcaggacta caaatcctgc tctagagaca tggccatcgc tgacttagtt 21540
tgttcccagt attacaatgg catcatggtt ctacctggtg ttgttgatgc tgagaaaatg 21600
gcaatgtata ctggctctct tactggagct atggtatttg gaggactgac tgccgcagcg 21660
gcaataccat ttgccacggc agtacaagct cgcctcaatt atgtcgcact gcaaacaaat 21720
gtactacaag aaaaccagaa aattcttgca gaatcattta accaagcagt tggcaatata 21780
tcacttgcac tatcttctgt taatgatgcc atccagcaaa cttctgaggc tcttaacacc 21840
gtagctattg ctattaaaaa gattcaaaca gttgttaacc agcagggcga ggcattatca 21900
cacctgactg cacagctgtc aaacaatttt caagcaattt cgacttctat tcaagacatt 21960
tacaaccgtc ttgaggaagt agaggctaac cagcaagttg accgtctcat cacaggacgg 22020
ttggctgcac ttaatgcata tgttactcag ttactcaatc agatgtctca gattagacaa 22080
tctcgattgt tagctcagca aaagattaat gagtgtgtca aatcacagtc gtccagatac 22140
ggtttctgtg gaaatggcac acacatcttc tcacttacac agactgcacc aaatggcata 22200
tttttcatgc atgcagtgct agtacccaac aaattcacac gtgtcaacgc ttctgccggc 22260
atttgtgtgg ataatacgag aggctactca ttgcagcctc aacttatact ctaccagttt 22320
aataactcct ggagagttac acctagaaat atgtatgaac ccagactgcc ccggcaggct 22380
gatttcatac aattaactga ttgcagcgtt actttttaca acaccaccgc tgctaatctt 22440
cccaatatta tccctgacat tatagatgtc aatcaaacag tcagtgatat tattgacaat 22500
ttacctacag caacacctcc tcagtgggat gttggtatct ataacaacac tattctcaac 22560
ctcaccgttg agattaatga tctacaagag cggtctaaaa acctctcaca gattgcagat 22620
cgtttacaaa attatattga caatcttaac aatactctag ttgaccttga atggctcaac 22680
agagtggaaa cttaccttaa atggccgtgg tatatatggc ttgccattgc cctggctctt 22740
attgcatttg tgacaatcct cataacaatc tttctttgta ctggttgttg tggtggttgc 22800
tttggttgtt gtggcggttg ttttggcctt ttctctaaga agaaaaggta taccgacgac 22860
caaccaacac cgtcctttaa gtttaaggaa tggtagtcga cgactgggcc gttaccatcc 22920
ctggacaata tattattgct atactagttg tcatctgcat tggtgtggca ctacttttta 22980
ttaatacttg cttagcttgt gttaaattat tttacaagtg ctacctaggg gcagcatatc 23040
ttgttaggcc tattatagtg tactactcca agccgaaccc cgtacctgag gatgagtttg 23100
taaaagtaca ccaatttcct agaaacactc actatgtctg acgcagaaga gtggcaaatt 23160
attgttttca ttgcgatcat atgggcgctt ggcgtcatcc tccagggagg ctatgccacg 23220
cgtaatcgtg tgatctatgt tattaaactt attctgcttt ggctgctcca acccttcacc 23280
ctagtggtga ccatttggac cgcagtcgac agatcatcta agaaggacgc agttttcatt 23340
gtgtccataa tttttgccgt actgaccttc atatcctggg ccaagtactg gtatgactca 23400
attcgtttat taatgaaaac cagatctgca tgggcactct cacctgagag tagactcctt 23460
gcagggatta tggatccaat gggtacatgg aggtgcattc ccatcgacca catggctcca 23520
attctcacac cagtcgttaa gcatggcaag ctcaagctac atgggcaaga gctggccaat 23580
ggcatatcag tcagaaatcc gccacaggat atggtgatag tgtcaccaag tgacaccttt 23640
cactacactt ttaagaaacc tgtggaatca aacaacgatc cagaatttgc tgttctgata 23700
taccagggtg accgcgcttc aaacgctgga cttcacacca taaccacttc aaaggccggt 23760
gacgctcgcc tgtataagta tatgtaatgt gcaactgcca tctgcagctg cgagatttat 23820
atagattgtg caataagcgg cacatcagaa gagaggatgt tcctgagctt attgaccctc 23880
tcgttaaaac tcgctgtttt gcttacagtc tcgtggttct tgctaatgct aatccaattg 23940
catttagcat actacctcgg aaaattctta tcaatggtga gcctttactg cttgaatatg 24000
gtagcatata tggtaaagac tttatcattc gaccatcgct ccaagtcatt cttgaagatg 24060
aattaaatta aagttttgac accaatctat catggctgca ccagtagtcc ctactactga 24120
cgcgtcttgg tttcaggtgc tcaaagctca aaacaaaaag gctactcatc ctcagtttcg 24180
tggcaatgga gttccgctta actccgccat caaacccgtt gaaaaccatg gctactggct 24240
gcgttacacc agacaaaagc caggtggtac tcccattcct ccatcctatg ccttttatta 24300
tactggcaca ggtcccagag gaaatcttaa gtatggtgaa ctccctccta atgatacccc 24360
agcaaccact cgtgttactt gggttaaggg ttcgggagct gacacttcta ttaaacctca 24420
tgttgccaaa cgcaacccca acaatcctaa acatcagctg ctacctctcc gattcccaac 24480
cggagatggc ccagctcaag gtttcagagt tgaccccttc aacgctagag gaagacctca 24540
ggagcgtgga agtggcccaa gatctcaatc tgttaactcc agaggcacag gcaatcagcc 24600
caggaaacgc gaccaatctg caccagctgc ggtacgtcgt aagacccagc atcaagctcc 24660
caagcggact ttacccaagg gtaaaaccat ttctcaggta tttggcaacc ggtctcgtac 24720
tggtgccaat gtcggctctg cagacactga gaagacgggt atggctgatc ctcgcatcat 24780
ggctctagcc agacatgtgc ctggtgttca ggaaatgctt tttgctggcc accttgagag 24840
caactttcag gcgggggcaa taacccttac cttctcctac tcaatcacag tcaaggaggg 24900
ttctcctgac tatgagagac ttaaggatgc gctcaatacg gtcgttaacc agacctatga 24960
gccacctacc aaaccaacta aggacaagaa gcctgacaaa caagaccagt ctgctaaacc 25020
caaacagcag aagaaaccta aaaaggtaac tctgccagca gacaaacagg attgggagtg 25080
ggatgatgct tttgagataa agcaggaatc agcagcgtag acatcaatct atgtctgtta 25140
aacccaccca actccactca aatatctctt tggttccaga gagtcgtagt gtatagccag 25200
agagccagtc agagggcgct atcatgcaaa ctagggctgg ctactctagc acagaatcac 25260
atcccgataa tcaacagtgc tagaaggttg attataccat ttaatatgcc gaggccacgc 25320
ggagtacgat cgagggtaca gcataatctc aacttttgtt gagccacaat tttaatccta 25380
attggagaag gccaaaggac tgtactactt ttgtgggtgt agcagtcgcc cagtgggaaa 25440
gcgccaacta ggttacaatt gtggtgggga caaattaggg gaaattaaat tggcttat 25498
<210> 10
<211> 25498
<212> RNA
<213> Porcine Deltacorona Virus ssRNA +
<400> 10
cuuugagcac cugaaaccaa gacgcgucag uaguagggac uacuggugca gccaugauag 60
auuggugucg guagagaacu agcgaagcua gggagauaaa auuauagacu aaugcuauaa 120
uuuuuaucuu uagucuauaa uuuuaucucc cuagcuucgc uaguucucua ccgacaccaa 180
uccaggugcg ucugccacca aguuggcuac ccuuccuagg ggcgcuuucg cgcuugcuca 240
ccauuagauu accuggaaac cagccauuca gguuggaguu uccccaggcu cuuuugugug 300
ggcauuagcg gcuugugguu uuugcacaaa aucuaagcua cuuaccguuc cucugaccau 360
ccaccacuuc uauagacagc acugauuacc guaggguuua agucacaccg gucugcaccg 420
cccgucagcg gacacauuac ccagcauagc acuccuugca ccgcgccuag guaggauaaa 480
acccccuacc gggugacucu uaaggcguuu ccuccacggg auagccacua gucacuaggu 540
guaagugauc ugaucugggc guauuguguu gcgcaagugu gauacccaua ggagcgugga 600
auccuauucu gcggcucagu gccugauaua gcugugaaau ggccaagaac aaguccaagc 660
gcgacgcuau ugcguugccu gaaaauguac caccaccucu gcaacuuuuc auucauguug 720
cagcugcuga agagggucac ccuaagguua cuacuuaccu uggcaacuau aaccucuaug 780
ccaccaaggc uccgccuggc gugcagguuc uuagugcuaa aaccucucuu acugacuuug 840
agaaugucuu uggagcucaa cccaccuugc gaucaauucg uaaucugguu ugugaggcuc 900
gcucggcuga auggacaacu uccaagaaug cuuuugcacu caaagccacu caacuugacu 960
acucugaugc cguuuugagg gcaaugauuc guuucugccc uccaaaggug uccacacucg 1020
cugccuuugc ucuuuuuggc cgauugguua aaauugagga caaggaacuu gcugaguuag 1080
cucgugauac ugcccuugag uuggcguaca cggcuaaaau ugguacaucu cuugcugaca 1140
cgagaucugu cucacuuauu cauaaggacg cuuaucuaac ucucaguaau gagguuguug 1200
gcguaacuuu uacugccgca cuuauggcaa aggcuaccac uguuaaugga gcaaugcaau 1260
acucaaacuu uuaccucuac ccucgugcca cuauuaaggu aaccgauggu aaggcugaag 1320
caauugcaac uaagccucug ucugcugcca cuaaaggcaa gcaaaucaca gaggauguca 1380
accuucuccc ugacuaucag cagcugcuug uugaucaagu gacuggcacu gagguuaagg 1440
uuggagcucu aaccuauguu aagaccacug auucgccacc ccuuuacuuu cccaaaguca 1500
aggguggugu uauugguauu gcacuuaagc agcagggcac uguggcuaag aagcucaaug 1560
uagucuucca ugcucaaccu gaugauguuc ugcuagccuu cauacaacuu cagcaauucu 1620
ugaaccguac uucggauuca aguguugaaa uuacugauug ccagaguuau gaaguaucuc 1680
caacugugac ggucaaaauu ggcccgucua aaccugggga ugucaucgug gcuacugaug 1740
aggaauaccu uaaaugcuuu gaaaccccug agguagguag gcucuauaag guuuuccaaa 1800
cucaaucuug ggcuaucauu gagcgugccu ucuccaguuu gaagauccgc guguccaaag 1860
cuuuaucagc auuuauaagu uuucugcaaa accuugcaga uaacuuuacu gcaauaagug 1920
guguugucac ugcacucauu cgugaacucc aggaucuuac ccuggaugug gcgacacgua 1980
ucacuaacau acaauuuguu uaccgugccg guaagcuuau ugucgacacg acaaguguca 2040
uagcuaaacu uuuccagcca uuuugugauu uuauaucacc uuuccuucgg aaaguugcug 2100
guuuugcaau uuacacuguu gguaaucgca ugcuuauguu uaccagcacu ggcaccuuuc 2160
uucucacaaa ggcaacuacu aagauacuca auaaggcaaa guacaucuuu gauguggagc 2220
cugaguaccc aguagaugua acaacaucca aaguuguagu acaugaagca cuccagcaaa 2280
ccgacacuaa gccuacuaga gcucuggagg cuguugaugu cguuguuggu aauacuguac 2340
ugcaaauggc uacugauggc acugcguucu acccaucgga ugguacgcac gccucucuuc 2400
caggauucaa agcagguucg gaugagcuuu ucauaagcuu caacugcgac cucuuugaug 2460
augagacuaa ugcucaaauc aacgaaacac ucgcugcaua ugagcuuaac caacuagugg 2520
cuccagguga uucuacaccg cgucaaauug cgacguuggu ugucgauaca cuugcagaug 2580
cuauaacaga ccacuuuccg gagaaaacca uugaucuacc ugaagacuau caagucuuuu 2640
cugaccauga ugaccuccca cucgcacaau accacauccc ugaucaccug agccuguaua 2700
uucaggcuau ggaaggugaa gaugauagug gugaugaaau auguauugag gacgaugauu 2760
acgacugucc ucaagccgac gaagacacag aaggaguaau uccccaacag ugggaacuuc 2820
cugauguuga uaaauuuuua cucaagaucc aggaacggaa gaccagcagc gacgaaguac 2880
uuagcgucga cgucuauccu aaaccagagc cggucggcaa uguugggauu gacgacagcg 2940
cgucggaaaa gaagccaaau ggggacucag uaccggaucc ugagguccau ccaacacuag 3000
agagugugga uguugaacga ccaaccgaaa cagcaaacca ggcuguugaa gacaaaccuu 3060
cugauaccac cuuugugguu gaugaggaac aauuacaaga aucaacacca gaacaugaac 3120
uccgcuccua ugaaggggag uuugauucug augaugaaau uauuauuccu auaguaccag 3180
uaacaccugc ggauuuaaaa ccacagacua uuacuauaaa ggaguacuuu aagucugaaa 3240
aacuugagac uauuaacgaa ggauccacag agucaguuac acaaucugac gauucguuug 3300
acgagucauu uguugaugcu gagucugaug auccacaaga uccugcugua uaugaugaua 3360
caacaauuau aacggacagc acugauguag gcgaugagcc ugagacaacu cuagcuacca 3420
ucguuaacac accucugaca cucgauaaua acuugccacc ugaagccauu aaacaaccca 3480
gcccaacuaa gguugaguua guuguuggug aauuggcgag uauuaaauuu gacaauucug 3540
uucuagucaa cccugcuaau gcgcaauuaa caaauggcgg uggagcugcu cgugcaauug 3600
caaaauuagc ugguccaaaa uaucaagagu acuguaauag uguggcuccu aucucaggac 3660
cgcuuaccac ggacucuuuu gaugccaaga aauuuggugu agccugcauc uugcauguag 3720
ugccacccaa agguucugac ccuaauguac aagaacuccu guaucaagcu uacaagagua 3780
uccuuacuga accagcacac uauguuauac cuauacuagg ugcugguauc uuuggaugca 3840
acccagucca cucucuggau gcguucagga aagcaugucc aagugacaua ggucguguca 3900
cccuugucac uaugaacaaa aaccauuugc agguguggga ugcucucaau aggaccauug 3960
uacgcaccac uacugacuau gaucaaguua ccaccaaggc ccuuacaccc cagggagugu 4020
uagaagccaa ucucuuugau ggugaggacu uuguucaaga accaaaaccc ggucaaaucu 4080
accuugaggu uacugaagaa guucagaacc aagccaagga acuugaccuu aaccuucagc 4140
aauacugcgu cuaccugaag acuugccacc auaaaugggu ugugagucgu acgaacgggu 4200
ugaugcaucu aaaacaaaaa gauaacaauu guuuuguuag ugcaggugua aaccuguuuc 4260
aaaacacugc uuaucaacuu agaccugcua uugaugcucu cuauagggag uaucuuaaug 4320
guaauccaaa uagauuuguu gcuuggaucu acgcauccac uaaccgucgu guuggugaga 4380
uggguugucc acagcaaguu auuucuuugc ucguuaguaa cucugacgca gcauuuucag 4440
caacuacagc cuguuguaac accuacuuua accacacagg uguuauuuca guagcucgug 4500
aauaugaccc aauacaacca aaggucuacu gcaugaagug ugaugugugg acucccuuua 4560
caccccagag uggaaaaggu gcaguugcaa uugguauuuc ugcagaugaa ccuaccgguc 4620
cugccauuaa auuugccgca gcucacugcu gguacacuaa uggcaagaaa acaguuaaug 4680
gcuaugacac uaaagcuaau guuguagcua ccuaucauag guuugacgug ccuaagccuc 4740
aacuugucga ggacgugguu gcgcugccua cuaaaaauga cuuugaaguu cucaauguug 4800
aagaacugcc gcaggauagu gugcuccauu uggacccacc uccuguacag gccuuacaac 4860
cuaaggcuaa ccaacacauu gagauucuag aaaacccaga uuaucuggac auuuuggauc 4920
uuuggauucg uaaacccaaa uucauccucg uaaagucgug gaguguuuug gguagagcac 4980
uauguaaggc agguaaaguu gucuuuguca gugcuucgcu uuugacccgu uucuacaauu 5040
accuuguaga gauuggugcu cuugacucaa caauaagguu gucagucgau cuuaccugua 5100
aauuuguuag aacgguucuc ccaucgucua acacuguaca caaaacuugu cuuggucugu 5160
auuauucagc ccagacacuu uuuguuucuu uagcaccauu ccuuauguua ccagcuguag 5220
uuagucugcu uaauucaggc uauacaauug gcacauauuu guaugcaaaa acuggcuggc 5280
cuuguaauua caaugccacg caacacuuug auuauaauuc uuacugugca ggugacuugg 5340
uuugucaagc cuguuuugac ggucaagacu cccuacauuu guauccgcau uuacguguua 5400
aucagcagcc ccuucagacc acugacuaca cuguuuaugc gcuuucacua auacuacuau 5460
uagcuaacau gacucuuguc augggcacgc uaauaguuac uuucuuugug aacuucuaug 5520
gugugcaaau accauuuuau gguacacuuu ugauagauua ucaauccgca cuggugauua 5580
cuuucucagu guacuacuuu uauaagguaa ugaaguuuuu ccgccaucuc acacauggau 5640
guaaaauucc aacgugugug guaugugcca aacuucguac cccaccuacu auaacaguug 5700
agacugucgu ucagggcagg aaauacccau cuguuauuga aacaaauggc ggguuuacaa 5760
uuuguaaaga acacaacuuc uauugcaagg acugcucuuu acaaacaccc ggcacuuuca 5820
ucccgacaga agcuauugag ucgcucucac gagcuaccag gcuuaguguc aaaccaacag 5880
caccagcauu cuuacuugcu agagauguug agugccaaac ugauguuguc guugcucgcg 5940
caaugcauaa ccaaaaugcg caugugugca uuucaaaaua cucugauauc cguaccguug 6000
accaacuacu uaagccuacu ccacuguuuu cauacacucc cgauguuauc aucgcggcag 6060
acuuugacaa cagagguagu cuuaagacag cuaaagaauu agcugugguu uugucaaugg 6120
accuuaaacg uacuauaauu aucauugauc aggccuauuc uagaccuauu gauaauuauc 6180
aggaaguugc uucucguauu gagaaguauu acccaguugc aaagaucaca cccacaggug 6240
acaucuuuac agacauuaag caagcgacca auggccaagc uagugacucu gcuauuaaug 6300
cagcuguucu ggcuguccag cgcggucuug auuuuacaau ugacaacccu aacaacauau 6360
uaccacauua cgccuuugac uuuucaaccc ucaaugcaga agaccagucu accauuuugg 6420
agagugguug ugcuaaaggc aaucucaagg gcacuaaugu ugguguuguu cuuucagcua 6480
gccuuguuac acgucuuagu cagcaggcua uacgugugau ugcuaaugcu gcuucacgua 6540
augguguuac augcgcuguu acuccuucua cacuuguuau gcgugggaau auugcaacac 6600
agcccuugac ucgcaucaaa gcuggugcac cucccaugcg ucaaaaaauu uuauguguua 6660
uccuggcacu ugcuauugug uacuuugcug cuauggcuuu uggcuuuuug gcaagucaaa 6720
uuacgcuuaa uacagugccu acgauuaaau cugauauccg cgccucuacc uucuacguug 6780
uuagagaugg agucuuggau acuguucguu caaaugacaa gugcuuugca aauaaguuuu 6840
uggcauuuga uagcuucauu caagcaccuu acacuaauuc accugacugu ccaguuguug 6900
ugggaguugu ugauguaacg acgcacucua uuccuggaau uccagcaggu gucauucaua 6960
gagacggucu cauacuuaac auuuaugaac agucucuuua ugaaacucau cagcgucagu 7020
cuaugguuag ggaugcguug ucacucaaga cagcaaaucu cuuuaaccua ggcaagcgug 7080
uuguaguagg auacacucaa caugaaguug uuguggguac cuccuauuuu aauucuccug 7140
cacuuuuuaa ugcaaagugc accuucuuac aguaucagga cacuagacaa cucuauugcu 7200
augauacugu uccuacugaa cauaagcucu acucugaugu gcuuccgcac gucgaguaua 7260
aggcuauuga cauuaauggu gaucuuguuc cuuucaagau accagagcag auaauguucu 7320
auccacauau ugugcgcuau acuagcaauu ccuauugccg uauggggcau uguuuuaaua 7380
cuaacccugg uauuugcauu ucauuuacgg acgaauuucc guauagugaa aaugucaaac 7440
cuggugugua cugugcugau accucuuugc aguuguuuuc aaaccucguu uugggcacug 7500
uaucugguau ucacaucuuu acaucaacag cugcauugcu uggaucuacu auugugauca 7560
uacuaugcgu uguugcuguu cuugcaguuc agcgauucuu caaggaguac acaacuuuug 7620
uuauguacac uuguggucuu gcucuuguca acauuguagg cauugcacuu auguacaagu 7680
gccuugucuu cgcgauuuuc uauuaugcaa ucuaccuuua cuuuguccuu acuuuccccu 7740
ccuuuaagag gaauguggca uuguuuuacu ucgcuguagu gaucgugccg cacgugagua 7800
acaugcaauu gcuugcgcuc auugugugua gcauuaucua cuuucucuac accuauguuc 7860
auacuguagc uaagacagcu gggaaauuuu cuuccuucuu agacgcagcu aaagcuacuu 7920
uugucauuga caaugaaaag uacguguugc uuaaagaccu cgcuggugcu gaauuugacc 7980
aguaucuggc cucuuacaac aaguacaaau auuuuucugg uacugcuucu gauaaggauu 8040
augauaaggu cuguauggca uuucuugcca aggcuuuguc aucuuuucgu gaaggaggcg 8100
guucacaguu guacacacca ccuaaauuug caguuguuca gagucuuaag accaagcugc 8160
aagcagguau caaaauccuc cugcacccuu cagguguagu ugagcgaugu auggucucag 8220
uugucuacaa uggaucugca uugaauggca ucuggcuuaa gaauguuguc uacugcccac 8280
gccauguaau uggaaaauuc cguggugacc aguggacuca cauggucuca auugcugauu 8340
gccgcgacuu uauagucaag uguccaauac aggguauuca gcuaaauguc caaucaguua 8400
agaugguagg agcucuccuc caguuaacug uucauaccaa caacacagcc acuccagacu 8460
auaaguuuga aaggcuacaa ccaggaucau cgaugacaau ugcuugugcu uaugauggca 8520
uuguacggca ugucuaucac gugguccucc aacuuaauaa ucuuauuuau gcaagcuucc 8580
uuaacggagc uugugguagu guggguuaca cucuuaaggg uaaaacacuc uacuuacauu 8640
acaugcacca cauugaguuu aacaacaaaa cucauagugg uacagaucuu gaagguaacu 8700
ucuauggccc cuauguggau gaggaaguua uucagcaaca aacagcauuc caguauuaca 8760
cugauaaugu uguugcucaa uuauaugcac acuuacugac uguugaugcu agaccaaaau 8820
ggcuggcaca aucucagaua aguaucgagg auuucaacuc augggcugcu aacaauuccu 8880
uugcuaacuu cccaugugaa caaacuaaua uguccuacau uaugggacuc ucgcaaacag 8940
cucgaguccc uguagaacgu auccucaaua ccauuauaca gcuaaccacc aauagagaug 9000
gugcuuguau uaugggaucu uaugauuucg agugcgauug gacgccagag augguauaca 9060
aucaggcucc aauuucauug cagucaggag uaguuaagaa aacuuguacg ugguucuucc 9120
acuucuuguu uauggcuauu accaugcuac ucgcugccau gcauguuuuc ccuguacacu 9180
uguauccaau aguacugcca ugcuucacug ucguggcauu ccuguugacu uuaaccauua 9240
aacacacugu uguguuuacc acuacauacu ugcuuccguc acuuuugaug augguuguaa 9300
augcuaacac uuuuuggaua ccgaacacau uucugcgcac cugcuacgaa acuauauucg 9360
guuccccaau ugcucagcga cuguaugguu acacuguugc ucuuuauaug cugaucuaug 9420
cuggacuugc aaucaacuau acguugaaaa cacuccggua uagagcaacu ucauucuuau 9480
cuuuuugcau gcagugguuu caauaugguu auguugcaca cauugcguac aaacugcuua 9540
auaaacccug gacagaauca cuacucuuca cagccuucac aaugcuaacc agucaucccu 9600
uguuggcugc ucuuagcugg uggcuagcug gucgcguaac ucugcccauu aucaugccug 9660
acuuagcuau ucguguuuug gcguauaacg ucauuggcua ugucauaugu guucgauuug 9720
gccuuaugug gcuugcaaau cgguucacaa cuguaccuau gggcacauac caguauaugg 9780
ugucuguaga gcaacuuaag uacaugaugg caguuaagau guccccaccg cguaaugcgu 9840
uugaggugcu uauagccaac auuagacuuc uugguuuggg uggaaaccgu aacauugcug 9900
uuucuacugu ccaaaacaaa auucuugaug caaaagcuac ugcuguuguu guugcuaacc 9960
uucuugaaaa ggcuggcguc acaaacaagc acgcuauuug caaaaagauu gugaaacucc 10020
acaaugauac ccuuaaagcc accacuuaug aggagguuga gguagcacuu gugaaacuuc 10080
uuucucacau aauugaguuc uugccaacug aucagguaga ugcuuaucua gcugaugcgg 10140
ccaaugcuca acauguuaau accuauuuag acaacuugcu ugagaacaaa gcuguuguuc 10200
aggcuguugc cgauaucaac auuaaucugg auucuuauag aauuuauaag gaggcagaug 10260
cuauuuauaa acgaucuguu gagaugaacg aaucuccgca ggagcaaaag aaaaagcuua 10320
aagcuguuaa cauugcaaag gcggaauggg agcgugaggc ugcuucucag cguaagcuug 10380
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aauuaaauua aaguuuugac accaaucuau cauggcugca ccaguagucc cuacuacuga 24120
cgcgucuugg uuucaggugc ucaaagcuca aaacaaaaag gcuacucauc cucaguuucg 24180
uggcaaugga guuccgcuua acuccgccau caaacccguu gaaaaccaug gcuacuggcu 24240
gcguuacacc agacaaaagc caggugguac ucccauuccu ccauccuaug ccuuuuauua 24300
uacuggcaca ggucccagag gaaaucuuaa guauggugaa cucccuccua augauacccc 24360
agcaaccacu cguguuacuu ggguuaaggg uucgggagcu gacacuucua uuaaaccuca 24420
uguugccaaa cgcaacccca acaauccuaa acaucagcug cuaccucucc gauucccaac 24480
cggagauggc ccagcucaag guuucagagu ugaccccuuc aacgcuagag gaagaccuca 24540
ggagcgugga aguggcccaa gaucucaauc uguuaacucc agaggcacag gcaaucagcc 24600
caggaaacgc gaccaaucug caccagcugc gguacgucgu aagacccagc aucaagcucc 24660
caagcggacu uuacccaagg guaaaaccau uucucaggua uuuggcaacc ggucucguac 24720
uggugccaau gucggcucug cagacacuga gaagacgggu auggcugauc cucgcaucau 24780
ggcucuagcc agacaugugc cugguguuca ggaaaugcuu uuugcuggcc accuugagag 24840
caacuuucag gcgggggcaa uaacccuuac cuucuccuac ucaaucacag ucaaggaggg 24900
uucuccugac uaugagagac uuaaggaugc gcucaauacg gucguuaacc agaccuauga 24960
gccaccuacc aaaccaacua aggacaagaa gccugacaaa caagaccagu cugcuaaacc 25020
caaacagcag aagaaaccua aaaagguaac ucugccagca gacaaacagg auugggagug 25080
ggaugaugcu uuugagauaa agcaggaauc agcagcguag acaucaaucu augucuguua 25140
aacccaccca acuccacuca aauaucucuu ugguuccaga gagucguagu guauagccag 25200
agagccaguc agagggcgcu aucaugcaaa cuagggcugg cuacucuagc acagaaucac 25260
aucccgauaa ucaacagugc uagaagguug auuauaccau uuaauaugcc gaggccacgc 25320
ggaguacgau cgaggguaca gcauaaucuc aacuuuuguu gagccacaau uuuaauccua 25380
auuggagaag gccaaaggac uguacuacuu uugugggugu agcagucgcc cagugggaaa 25440
gcgccaacua gguuacaauu guggugggga caaauuaggg gaaauuaaau uggcuuau 25498
<210> 11
<211> 3471
<212> DNA
<213> Artificial Sequence
<220>
<223> Porcine Deltacorona Virus BIV-Isolate 5.0327 Spike Antigen cDNA
<400> 11
atgcagagag ctctattgat tatgacctta ttttgtctcg ttcgagcaaa gtttgctgat 60
gatctactcg atttgctcac cttcccgggt gcacatcgct tcttacataa acccacgagg 120
aattccagca gtctctactc gcgggctaat aataattttg atgttggcgt tcttcctggc 180
taccccacta agaacgttaa cctcttctca ccacttacta actccacttt gcccattaat 240
ggccttcatc ggagttacca accactcatg ctgaattgtc ttactaaaat aactaaccac 300
actctcagca tgtatctcct acctagtgag atacaaactt atagctgcgg cggtgccatg 360
gttaaatacc agacacatga tgcagttcgt atcattttag acctcactgc cactgaccac 420
atctctgttg aagtcgttgg ccaacatggt gaaaattatg tgtttgtttg cagtgagcag 480
tttaactaca ccactgcatt acacaactct accttcttct cacttaattc tgagctttat 540
tgctttacta ataacaccta cttaggtatt cttccacctg atttaactga ctttacggtc 600
taccgtactg gtcagttcta tgctaatggt taccttttag gtactttacc tattacggtt 660
aactatgtta ggttgtatcg gggtcatttt agtgcccact ttgcccttgc aaacctaacc 720
gatacactca taacacttac caatactact atatcgcaaa tcacttattg tgataagtct 780
gtagttgatt caatagcatg ccagcgctct tctcacgaag tggaggatgg gttttactcc 840
gaccctaaat ctgccgttag agctaggcaa cgtactattg ttacactacc taagctccct 900
gagcttgaag tagtgcagtt aaatatttct gcacacatgg attttggcga agccagactt 960
gacagcgtta ccatcaatgg taacacatcc tattgtgtca ctaagcctta cttcaggctt 1020
gaaactaact ttatgtgtac aggttgcact atgaatctgc gcactgatac ctgtagtttt 1080
gacctgtcag cagtaaacaa tggcatgtca ttctctcaat tctgtctaag cactgaatct 1140
ggtgcttgtg agatgaaaat tattgttacc tacgtatgga attacttgct aaggcagcgt 1200
ttgtatgtta ctgctgtaga gggccagact cacactggaa ccacttcagt acatgcaaca 1260
gacacttcta gtgtaatcac tgatgtctgc actgactaca ctatctatgg agtctctggt 1320
actggcatta ttaagccatc agatctctta ttgcacaatg gcatagcatt cacctctcca 1380
acaggtgagc tttatgcatt taaaaatata accactggca aaacccttca ggtcttaccg 1440
tgtgaaaccc cttctcaact gattgtgata aacaacaccg ttgtcggtgc tatcacatcc 1500
agtaattcaa ctgaaaataa taggtttact actactattg tcacacctac tttcttttat 1560
tccacaaatg ccaccacttt caactgcact aagcctgttt tgtcctatgg acctatcagc 1620
gtgtgtagtg atggtgcaat tgtgggaaca tccacattac agaatactcg accatccata 1680
gtttcactat acgatggcga agttgaaata ccatctgcat tttctctttc cgttcagacg 1740
gagtacttgc aagttcaagc agagcaagtt atagttgatt gtcctcagta tgtatgcaat 1800
ggcaacagcc gttgtctaca attactggca caatacacct cagcttgctc taacattgaa 1860
gcagctctgc attcctctgc acagttggat agcagagaga ttataaatat gtttcaaaca 1920
tcaacacagt ccttgcagtt agctaatatt accaacttca agggtgacta caattttagc 1980
agcatactaa ccaccagaat tggtggcaga tctgctattg aagaccttct ttttaataaa 2040
gttgttacta gtggccttgg cactgttgat caggactaca aatcctgctc tagagacatg 2100
gccatcgctg acttagtttg ttcccagtat tacaatggca tcatggttct acctggtgtt 2160
gttgatgctg agaaaatggc aatgtatact ggctctctta ctggagctat ggtatttgga 2220
ggactgactg ccgcagcggc aataccattt gccacggcag tacaagctcg cctcaattat 2280
gtcgcactgc aaacaaatgt actacaagaa aaccagaaaa ttcttgcaga atcatttaac 2340
caagcagttg gcaatatatc acttgcacta tcttctgtta atgatgccat ccagcaaact 2400
tctgaggctc ttaacaccgt agctattgct attaaaaaga ttcaaacagt tgttaaccag 2460
cagggcgagg cattatcaca cctgactgca cagctgtcaa acaattttca agcaatttcg 2520
acttctattc aagacattta caaccgtctt gaggaagtag aggctaacca gcaagttgac 2580
cgtctcatca caggacggtt ggctgcactt aatgcatatg ttactcagtt actcaatcag 2640
atgtctcaga ttagacaatc tcgattgtta gctcagcaaa agattaatga gtgtgtcaaa 2700
tcacagtcgt ccagatacgg tttctgtgga aatggcacac acatcttctc acttacacag 2760
actgcaccaa atggcatatt tttcatgcat gcagtgctag tacccaacaa attcacacgt 2820
gtcaacgctt ctgccggcat ttgtgtggat aatacgagag gctactcatt gcagcctcaa 2880
cttatactct accagtttaa taactcctgg agagttacac ctagaaatat gtatgaaccc 2940
agactgcccc ggcaggctga tttcatacaa ttaactgatt gcagcgttac tttttacaac 3000
accaccgctg ctaatcttcc caatattatc cctgacatta tagatgtcaa tcaaacagtc 3060
agtgatatta ttgacaattt acctacagca acacctcctc agtgggatgt tggtatctat 3120
aacaacacta ttctcaacct caccgttgag attaatgatc tacaagagcg gtctaaaaac 3180
ctctcacaga ttgcagatcg tttacaaaat tatattgaca atcttaacaa tactctagtt 3240
gaccttgaat ggctcaacag agtggaaact taccttaaat ggccgtggta tatatggctt 3300
gccattgccc tggctcttat tgcatttgtg acaatcctca taacaatctt tctttgtact 3360
ggttgttgtg gtggttgctt tggttgttgt ggcggttgtt ttggcctttt ctctaagaag 3420
aaaaggtata ccgacgacca accaacaccg tcctttaagt ttaaggaatg g 3471
<210> 12
<211> 1157
<212> PRT
<213> Porcine Deltacorona Virus
<400> 12
Met Gln Arg Ala Leu Leu Ile Met Thr Leu Phe Cys Leu Val Arg Ala
1 5 10 15
Lys Phe Ala Asp Asp Leu Leu Asp Leu Leu Thr Phe Pro Gly Ala His
20 25 30
Arg Phe Leu His Lys Pro Thr Arg Asn Ser Ser Ser Leu Tyr Ser Arg
35 40 45
Ala Asn Asn Asn Phe Asp Val Gly Val Leu Pro Gly Tyr Pro Thr Lys
50 55 60
Asn Val Asn Leu Phe Ser Pro Leu Thr Asn Ser Thr Leu Pro Ile Asn
65 70 75 80
Gly Leu His Arg Ser Tyr Gln Pro Leu Met Leu Asn Cys Leu Thr Lys
85 90 95
Ile Thr Asn His Thr Leu Ser Met Tyr Leu Leu Pro Ser Glu Ile Gln
100 105 110
Thr Tyr Ser Cys Gly Gly Ala Met Val Lys Tyr Gln Thr His Asp Ala
115 120 125
Val Arg Ile Ile Leu Asp Leu Thr Ala Thr Asp His Ile Ser Val Glu
130 135 140
Val Val Gly Gln His Gly Glu Asn Tyr Val Phe Val Cys Ser Glu Gln
145 150 155 160
Phe Asn Tyr Thr Thr Ala Leu His Asn Ser Thr Phe Phe Ser Leu Asn
165 170 175
Ser Glu Leu Tyr Cys Phe Thr Asn Asn Thr Tyr Leu Gly Ile Leu Pro
180 185 190
Pro Asp Leu Thr Asp Phe Thr Val Tyr Arg Thr Gly Gln Phe Tyr Ala
195 200 205
Asn Gly Tyr Leu Leu Gly Thr Leu Pro Ile Thr Val Asn Tyr Val Arg
210 215 220
Leu Tyr Arg Gly His Phe Ser Ala His Phe Ala Leu Ala Asn Leu Thr
225 230 235 240
Asp Thr Leu Ile Thr Leu Thr Asn Thr Thr Ile Ser Gln Ile Thr Tyr
245 250 255
Cys Asp Lys Ser Val Val Asp Ser Ile Ala Cys Gln Arg Ser Ser His
260 265 270
Glu Val Glu Asp Gly Phe Tyr Ser Asp Pro Lys Ser Ala Val Arg Ala
275 280 285
Arg Gln Arg Thr Ile Val Thr Leu Pro Lys Leu Pro Glu Leu Glu Val
290 295 300
Val Gln Leu Asn Ile Ser Ala His Met Asp Phe Gly Glu Ala Arg Leu
305 310 315 320
Asp Ser Val Thr Ile Asn Gly Asn Thr Ser Tyr Cys Val Thr Lys Pro
325 330 335
Tyr Phe Arg Leu Glu Thr Asn Phe Met Cys Thr Gly Cys Thr Met Asn
340 345 350
Leu Arg Thr Asp Thr Cys Ser Phe Asp Leu Ser Ala Val Asn Asn Gly
355 360 365
Met Ser Phe Ser Gln Phe Cys Leu Ser Thr Glu Ser Gly Ala Cys Glu
370 375 380
Met Lys Ile Ile Val Thr Tyr Val Trp Asn Tyr Leu Leu Arg Gln Arg
385 390 395 400
Leu Tyr Val Thr Ala Val Glu Gly Gln Thr His Thr Gly Thr Thr Ser
405 410 415
Val His Ala Thr Asp Thr Ser Ser Val Ile Thr Asp Val Cys Thr Asp
420 425 430
Tyr Thr Ile Tyr Gly Val Ser Gly Thr Gly Ile Ile Lys Pro Ser Asp
435 440 445
Leu Leu Leu His Asn Gly Ile Ala Phe Thr Ser Pro Thr Gly Glu Leu
450 455 460
Tyr Ala Phe Lys Asn Ile Thr Thr Gly Lys Thr Leu Gln Val Leu Pro
465 470 475 480
Cys Glu Thr Pro Ser Gln Leu Ile Val Ile Asn Asn Thr Val Val Gly
485 490 495
Ala Ile Thr Ser Ser Asn Ser Thr Glu Asn Asn Arg Phe Thr Thr Thr
500 505 510
Ile Val Thr Pro Thr Phe Phe Tyr Ser Thr Asn Ala Thr Thr Phe Asn
515 520 525
Cys Thr Lys Pro Val Leu Ser Tyr Gly Pro Ile Ser Val Cys Ser Asp
530 535 540
Gly Ala Ile Val Gly Thr Ser Thr Leu Gln Asn Thr Arg Pro Ser Ile
545 550 555 560
Val Ser Leu Tyr Asp Gly Glu Val Glu Ile Pro Ser Ala Phe Ser Leu
565 570 575
Ser Val Gln Thr Glu Tyr Leu Gln Val Gln Ala Glu Gln Val Ile Val
580 585 590
Asp Cys Pro Gln Tyr Val Cys Asn Gly Asn Ser Arg Cys Leu Gln Leu
595 600 605
Leu Ala Gln Tyr Thr Ser Ala Cys Ser Asn Ile Glu Ala Ala Leu His
610 615 620
Ser Ser Ala Gln Leu Asp Ser Arg Glu Ile Ile Asn Met Phe Gln Thr
625 630 635 640
Ser Thr Gln Ser Leu Gln Leu Ala Asn Ile Thr Asn Phe Lys Gly Asp
645 650 655
Tyr Asn Phe Ser Ser Ile Leu Thr Thr Arg Ile Gly Gly Arg Ser Ala
660 665 670
Ile Glu Asp Leu Leu Phe Asn Lys Val Val Thr Ser Gly Leu Gly Thr
675 680 685
Val Asp Gln Asp Tyr Lys Ser Cys Ser Arg Asp Met Ala Ile Ala Asp
690 695 700
Leu Val Cys Ser Gln Tyr Tyr Asn Gly Ile Met Val Leu Pro Gly Val
705 710 715 720
Val Asp Ala Glu Lys Met Ala Met Tyr Thr Gly Ser Leu Thr Gly Ala
725 730 735
Met Val Phe Gly Gly Leu Thr Ala Ala Ala Ala Ile Pro Phe Ala Thr
740 745 750
Ala Val Gln Ala Arg Leu Asn Tyr Val Ala Leu Gln Thr Asn Val Leu
755 760 765
Gln Glu Asn Gln Lys Ile Leu Ala Glu Ser Phe Asn Gln Ala Val Gly
770 775 780
Asn Ile Ser Leu Ala Leu Ser Ser Val Asn Asp Ala Ile Gln Gln Thr
785 790 795 800
Ser Glu Ala Leu Asn Thr Val Ala Ile Ala Ile Lys Lys Ile Gln Thr
805 810 815
Val Val Asn Gln Gln Gly Glu Ala Leu Ser His Leu Thr Ala Gln Leu
820 825 830
Ser Asn Asn Phe Gln Ala Ile Ser Thr Ser Ile Gln Asp Ile Tyr Asn
835 840 845
Arg Leu Glu Glu Val Glu Ala Asn Gln Gln Val Asp Arg Leu Ile Thr
850 855 860
Gly Arg Leu Ala Ala Leu Asn Ala Tyr Val Thr Gln Leu Leu Asn Gln
865 870 875 880
Met Ser Gln Ile Arg Gln Ser Arg Leu Leu Ala Gln Gln Lys Ile Asn
885 890 895
Glu Cys Val Lys Ser Gln Ser Ser Arg Tyr Gly Phe Cys Gly Asn Gly
900 905 910
Thr His Ile Phe Ser Leu Thr Gln Thr Ala Pro Asn Gly Ile Phe Phe
915 920 925
Met His Ala Val Leu Val Pro Asn Lys Phe Thr Arg Val Asn Ala Ser
930 935 940
Ala Gly Ile Cys Val Asp Asn Thr Arg Gly Tyr Ser Leu Gln Pro Gln
945 950 955 960
Leu Ile Leu Tyr Gln Phe Asn Asn Ser Trp Arg Val Thr Pro Arg Asn
965 970 975
Met Tyr Glu Pro Arg Leu Pro Arg Gln Ala Asp Phe Ile Gln Leu Thr
980 985 990
Asp Cys Ser Val Thr Phe Tyr Asn Thr Thr Ala Ala Asn Leu Pro Asn
995 1000 1005
Ile Ile Pro Asp Ile Ile Asp Val Asn Gln Thr Val Ser Asp Ile
1010 1015 1020
Ile Asp Asn Leu Pro Thr Ala Thr Pro Pro Gln Trp Asp Val Gly
1025 1030 1035
Ile Tyr Asn Asn Thr Ile Leu Asn Leu Thr Val Glu Ile Asn Asp
1040 1045 1050
Leu Gln Glu Arg Ser Lys Asn Leu Ser Gln Ile Ala Asp Arg Leu
1055 1060 1065
Gln Asn Tyr Ile Asp Asn Leu Asn Asn Thr Leu Val Asp Leu Glu
1070 1075 1080
Trp Leu Asn Arg Val Glu Thr Tyr Leu Lys Trp Pro Trp Tyr Ile
1085 1090 1095
Trp Leu Ala Ile Ala Leu Ala Leu Ile Ala Phe Val Thr Ile Leu
1100 1105 1110
Ile Thr Ile Phe Leu Cys Thr Gly Cys Cys Gly Gly Cys Phe Gly
1115 1120 1125
Cys Cys Gly Gly Cys Phe Gly Leu Phe Ser Lys Lys Lys Arg Tyr
1130 1135 1140
Thr Asp Asp Gln Pro Thr Pro Ser Phe Lys Phe Lys Glu Trp
1145 1150 1155
<210> 13
<211> 33
<212> DNA
<213> Artificial Sequence
<220>
<223> P2967167A PDCoV SPIKE FORWARD PRIMER
<400> 13
ggatccatgc agagagctct attgattatg acc 33
<210> 14
<211> 42
<212> DNA
<213> Artificial Sequence
<220>
<223> P2967167B PDCoV SPIKE REVERSE PRIMER
<400> 14
ggtcgagccg cctctgccac caattctggt ggttagtaag ct 42
<210> 15
<211> 31
<212> DNA
<213> Artificial Sequence
<220>
<223> P2967167C FC FORWARD PRIMER
<400> 15
tggtggcaga ggcggctcga ccaagaccaa g 31
<210> 16
<211> 36
<212> DNA
<213> Artificial Sequence
<220>
<223> P2968014E FC FORWARD PRIMER
<400> 16
gcggccgcct acttgcccgg cgtcttcgaa atggat 36
<210> 17
<211> 2718
<212> DNA
<213> Artificial Sequence
<220>
<223> PDCoV S1-IgG2a Fc CDS cDNA Expression Casette
<400> 17
atgcagagag ctctattgat tatgacctta ctttgtctcg ttcgagcaaa gtttgctgat 60
gatctactcg atttgctcac cttcccgggt gcacatcgct tcttacataa acccacgagg 120
aattccagca gtctctactc gcgggctaat aataattttg atgttggcgt tcttcctggc 180
taccccacta agaacgttaa cctcttctca ccacttacta actccacttt gcccattaat 240
ggccttcatc ggagttacca accactcatg ctgaattgtc ttactaaaat aactaaccac 300
actctcagca tgtatctcct acctagtgag atacaaactt atagctgcgg cggtgccatg 360
gttaaatacc agacacatga tgcagttcgt atcattttag acctcactgc cactgaccac 420
atctctgttg aagtcgttgg ccaacatggt gaaaattatg tgtttgtttg cagtgagcag 480
tttaactaca ccactgcatt acacaactct accttcttct cacttaattc tgagctttat 540
tgctttacta ataacaccta cttaggtatt cttccacctg atttaactga ctttacggtc 600
taccgtactg gtcagttcta tgctaatggt taccttttag gtactttacc tattacggtt 660
aactatgtta ggttgtatcg gggtcatttg tctgccaata gtgcctactt tgcccttgca 720
aacctaaccg atacactcat aacacttacc aatactacta tatcgcaaat cacttattgt 780
gataagtcag tagttgattc aatagcatgc cagcgctctt ctcacgaagt ggaggatggg 840
ttttactccg accctaaatc tgccgttaga gctaggcaac gtactattgt tacactacct 900
aagctccctg agcttgaagt agtgcagtta aatatttctg cacacatgga ttttggcgaa 960
gccagacttg acagcgttac catcaatggt aacacatcct attgtgtcac taagccttac 1020
ttcaggcttg aaactaactt tatgtgtaca ggttgcacta tgaatctgcg cactgatacc 1080
tgtagttttg acctgtcagc agtaaacaat ggcatgtcat tctctcaatt ctgtctaagc 1140
actgaatctg gtgcttgtga gatgaaaatt attgttacct acgtatggaa ttacttgcta 1200
aggcagcgtt tgtatgttac tgctgtagag ggccagactc acactggaac cacttcagta 1260
catgcaacag acacttctag tgtaatcact gatgtctgca ctgactacac tatctatgga 1320
gtctcaggta ctggcattat taagccatca gatctcttat tgcacaatgg catagcattc 1380
acctctccaa caggtgagct ttatgcattt aaaaatataa ccactggcaa aacccttcag 1440
gtcttaccgt gtgaaacccc ttctcaactg attgtgataa acaacaccgt tgtcggtgct 1500
atcacatcca gtaattcaac tgaaaataat aggtttacta ctactattgt cacacctact 1560
ttcttttatt ccacaaatgc caccactttc aactgcacta agcctgtttt gtcctatgga 1620
cctatcagcg tgtgtagtga tggtgcaatt gtgggaacat ccacattaca gaatactcga 1680
ccatccatag tttcactata cgatggcgaa gttgaaatac catctgcatt ttctctttcc 1740
gttcagacgg agtacttgca agttcaagca gagcaagtta tagttgattg tcctcagtat 1800
gtatgcaatg gcaacagccg ttgtctacaa ttactggcac aatacacctc agcttgctct 1860
aacattgaag cagctctgca ttcctctgca cagttggata gcagagagat tataaatatg 1920
tttcaaacat caacacagtc cttgcagtta gctaatatta ccaacttcaa gggtgactac 1980
aattttagca gcttactaac caccagaatt ggtggcagag gcggctcgac caagaccaag 2040
cccccctgtc ctatttgccc ggcatgcgag agtccgggtc cgtccgtctt tattttcccc 2100
ccaaagccaa aagatacgct gatgatcagc cgcacccccc aggtcacatg cgtggttgta 2160
gatgtaagcc aagaaaatcc cgaagtgcag ttttcctggt atgtcgatgg cgtggaggtt 2220
cacacagcac agacgcggcc gaaggaagag cagtttaatt ccacgtaccg agtggtgagt 2280
gtgctgccta ttcagcatca ggattggttg aacggaaagg agttcaagtg caaggtgaac 2340
aacaaagacc tgcccgcacc aatcactcgc ataatcagca aggctaaggg acagacacgc 2400
gagccacagg tgtacaccct gccaccgcat gcagaagagc tgagccgatc caaggtatcc 2460
ataacctgcc tggtgatcgg attctacccc cccgatatcg acgtcgagtg gcagaggaat 2520
ggccagccgg agccagaggg caactaccgg accacgcctc cgcagcagga tgtcgatggt 2580
acatattttc tgtacagcaa atttagcgtg gacaaggcca gttggcaggg cggcggtatc 2640
ttccagtgtg ctgtcatgca cgaggcactc cacaatcatt acacccagaa atccatttcg 2700
aagacgccgg gcaagtag 2718
<210> 18
<211> 905
<212> PRT
<213> Artificial Sequence
<220>
<223> PDCoV S1-IgG2a Fc Amino Acid Sequence
<400> 18
Met Gln Arg Ala Leu Leu Ile Met Thr Leu Leu Cys Leu Val Arg Ala
1 5 10 15
Lys Phe Ala Asp Asp Leu Leu Asp Leu Leu Thr Phe Pro Gly Ala His
20 25 30
Arg Phe Leu His Lys Pro Thr Arg Asn Ser Ser Ser Leu Tyr Ser Arg
35 40 45
Ala Asn Asn Asn Phe Asp Val Gly Val Leu Pro Gly Tyr Pro Thr Lys
50 55 60
Asn Val Asn Leu Phe Ser Pro Leu Thr Asn Ser Thr Leu Pro Ile Asn
65 70 75 80
Gly Leu His Arg Ser Tyr Gln Pro Leu Met Leu Asn Cys Leu Thr Lys
85 90 95
Ile Thr Asn His Thr Leu Ser Met Tyr Leu Leu Pro Ser Glu Ile Gln
100 105 110
Thr Tyr Ser Cys Gly Gly Ala Met Val Lys Tyr Gln Thr His Asp Ala
115 120 125
Val Arg Ile Ile Leu Asp Leu Thr Ala Thr Asp His Ile Ser Val Glu
130 135 140
Val Val Gly Gln His Gly Glu Asn Tyr Val Phe Val Cys Ser Glu Gln
145 150 155 160
Phe Asn Tyr Thr Thr Ala Leu His Asn Ser Thr Phe Phe Ser Leu Asn
165 170 175
Ser Glu Leu Tyr Cys Phe Thr Asn Asn Thr Tyr Leu Gly Ile Leu Pro
180 185 190
Pro Asp Leu Thr Asp Phe Thr Val Tyr Arg Thr Gly Gln Phe Tyr Ala
195 200 205
Asn Gly Tyr Leu Leu Gly Thr Leu Pro Ile Thr Val Asn Tyr Val Arg
210 215 220
Leu Tyr Arg Gly His Leu Ser Ala Asn Ser Ala Tyr Phe Ala Leu Ala
225 230 235 240
Asn Leu Thr Asp Thr Leu Ile Thr Leu Thr Asn Thr Thr Ile Ser Gln
245 250 255
Ile Thr Tyr Cys Asp Lys Ser Val Val Asp Ser Ile Ala Cys Gln Arg
260 265 270
Ser Ser His Glu Val Glu Asp Gly Phe Tyr Ser Asp Pro Lys Ser Ala
275 280 285
Val Arg Ala Arg Gln Arg Thr Ile Val Thr Leu Pro Lys Leu Pro Glu
290 295 300
Leu Glu Val Val Gln Leu Asn Ile Ser Ala His Met Asp Phe Gly Glu
305 310 315 320
Ala Arg Leu Asp Ser Val Thr Ile Asn Gly Asn Thr Ser Tyr Cys Val
325 330 335
Thr Lys Pro Tyr Phe Arg Leu Glu Thr Asn Phe Met Cys Thr Gly Cys
340 345 350
Thr Met Asn Leu Arg Thr Asp Thr Cys Ser Phe Asp Leu Ser Ala Val
355 360 365
Asn Asn Gly Met Ser Phe Ser Gln Phe Cys Leu Ser Thr Glu Ser Gly
370 375 380
Ala Cys Glu Met Lys Ile Ile Val Thr Tyr Val Trp Asn Tyr Leu Leu
385 390 395 400
Arg Gln Arg Leu Tyr Val Thr Ala Val Glu Gly Gln Thr His Thr Gly
405 410 415
Thr Thr Ser Val His Ala Thr Asp Thr Ser Ser Val Ile Thr Asp Val
420 425 430
Cys Thr Asp Tyr Thr Ile Tyr Gly Val Ser Gly Thr Gly Ile Ile Lys
435 440 445
Pro Ser Asp Leu Leu Leu His Asn Gly Ile Ala Phe Thr Ser Pro Thr
450 455 460
Gly Glu Leu Tyr Ala Phe Lys Asn Ile Thr Thr Gly Lys Thr Leu Gln
465 470 475 480
Val Leu Pro Cys Glu Thr Pro Ser Gln Leu Ile Val Ile Asn Asn Thr
485 490 495
Val Val Gly Ala Ile Thr Ser Ser Asn Ser Thr Glu Asn Asn Arg Phe
500 505 510
Thr Thr Thr Ile Val Thr Pro Thr Phe Phe Tyr Ser Thr Asn Ala Thr
515 520 525
Thr Phe Asn Cys Thr Lys Pro Val Leu Ser Tyr Gly Pro Ile Ser Val
530 535 540
Cys Ser Asp Gly Ala Ile Val Gly Thr Ser Thr Leu Gln Asn Thr Arg
545 550 555 560
Pro Ser Ile Val Ser Leu Tyr Asp Gly Glu Val Glu Ile Pro Ser Ala
565 570 575
Phe Ser Leu Ser Val Gln Thr Glu Tyr Leu Gln Val Gln Ala Glu Gln
580 585 590
Val Ile Val Asp Cys Pro Gln Tyr Val Cys Asn Gly Asn Ser Arg Cys
595 600 605
Leu Gln Leu Leu Ala Gln Tyr Thr Ser Ala Cys Ser Asn Ile Glu Ala
610 615 620
Ala Leu His Ser Ser Ala Gln Leu Asp Ser Arg Glu Ile Ile Asn Met
625 630 635 640
Phe Gln Thr Ser Thr Gln Ser Leu Gln Leu Ala Asn Ile Thr Asn Phe
645 650 655
Lys Gly Asp Tyr Asn Phe Ser Ser Leu Leu Thr Thr Arg Ile Gly Gly
660 665 670
Arg Gly Gly Ser Thr Lys Thr Lys Pro Pro Cys Pro Ile Cys Pro Ala
675 680 685
Cys Glu Ser Pro Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys
690 695 700
Asp Thr Leu Met Ile Ser Arg Thr Pro Gln Val Thr Cys Val Val Val
705 710 715 720
Asp Val Ser Gln Glu Asn Pro Glu Val Gln Phe Ser Trp Tyr Val Asp
725 730 735
Gly Val Glu Val His Thr Ala Gln Thr Arg Pro Lys Glu Glu Gln Phe
740 745 750
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp
755 760 765
Trp Leu Asn Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Lys Asp Leu
770 775 780
Pro Ala Pro Ile Thr Arg Ile Ile Ser Lys Ala Lys Gly Gln Thr Arg
785 790 795 800
Glu Pro Gln Val Tyr Thr Leu Pro Pro His Ala Glu Glu Leu Ser Arg
805 810 815
Ser Lys Val Ser Ile Thr Cys Leu Val Ile Gly Phe Tyr Pro Pro Asp
820 825 830
Ile Asp Val Glu Trp Gln Arg Asn Gly Gln Pro Glu Pro Glu Gly Asn
835 840 845
Tyr Arg Thr Thr Pro Pro Gln Gln Asp Val Asp Gly Thr Tyr Phe Leu
850 855 860
Tyr Ser Lys Phe Ser Val Asp Lys Ala Ser Trp Gln Gly Gly Gly Ile
865 870 875 880
Phe Gln Cys Ala Val Met His Glu Ala Leu His Asn His Tyr Thr Gln
885 890 895
Lys Ser Ile Ser Lys Thr Pro Gly Lys
900 905
<210> 19
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> P290194A F Primer PDCoVS BD
<400> 19
ggatccgcca ccatggtatc tg 22
<210> 20
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> P290194B R Primer PDCoVS BD
<400> 20
gcttaaatac atatttagta c 21
<210> 21
<211> 100
<212> DNA
<213> Artificial Sequence
<220>
<223> O2967002 A Primer Gp64 signal sequence
<400> 21
ggatccgcca ccatggtatc tgctattgtt ctttatgttt tgcttgctgc cgccgctcac 60
tcagctttcg cggatgatct actcgatttg ctcaccttcc 100
<210> 22
<211> 63
<212> DNA
<213> Artificial Sequence
<220>
<223> O2967002B AcNPV gp64 C-terminal tail coding sequence
<400> 22
gcggccgctt agtactgacg attgcggttt ctaagaaaga ttgttatgag gattgtcaca 60
aat 63
<210> 23
<211> 28
<212> DNA
<213> Artificial Sequence
<220>
<223> P2967002C F Primer PDCoVS BD
<400> 23
gatgatctac tcgatttgct caccttcc 28
<210> 24
<211> 26
<212> DNA
<213> Artificial Sequence
<220>
<223> P2967002D R Primer PDCoVS BD
<400> 24
cgatggccat gtctctagag caggat 26
<210> 25
<211> 25
<212> DNA
<213> Artificial Sequence
<220>
<223> P2967002E F Primer PDCoVS BD
<400> 25
tgtcctatgg acctatcagc gtgtg 25
<210> 26
<211> 31
<212> DNA
<213> Artificial Sequence
<220>
<223> P2968002F R Primer PDCoVS BD
<400> 26
aagaaagatt gttatgagga ttgtcacaaa t 31
<210> 27
<211> 3390
<212> DNA
<213> Artificial Sequence
<220>
<223> PDCoVS BD CDS Expression Cassette
<400> 27
atggtatctg ctattgttct ttatgttttg cttgctgccg ccgctcactc agctttcgcg 60
gatgatctac tcgatttgct caccttcccg ggtgcacatc gcttcttaca taaacccacg 120
aggaattcca gcagtctcta ctcgcgggct aataataatt ttgatgttgg cgttcttcct 180
ggctacccca ctaagaacgt taacctcttc tcaccactta ctaactccac tttgcccatt 240
aatggccttc atcggagtta ccaaccactc atgctgaatt gtcttactaa aataactaac 300
cacactctca gcatgtatct cctacctagt gagatacaaa cttatagctg cggcggtgcc 360
atggttaaat accagacaca tgatgcagtt cgtatcattt tagacctcac tgccactgac 420
cacatctctg ttgaagtcgt tggccaacat ggtgaaaatt atgtgtttgt ttgcagtgag 480
cagtttaact acaccactgc attacacaac tctaccttct tctcacttaa ttctgagctt 540
tattgcttta ctaataacac ctacttaggt attcttccac ctgatttaac tgactttacg 600
gtctaccgta ctggtcagtt ctatgctaat ggttaccttt taggtacttt acctattacg 660
gttaactatg ttaggttgta tcggggtcat ttgtctgcca atagtgccta ctttgccctt 720
gcaaacctaa ccgatacact cataacactt accaatacta ctatatcgca aatcacttat 780
tgtgataagt cagtagttga ttcaatagca tgccagcgct cttctcacga agtggaggat 840
gggttttact ccgaccctaa atctgccgtt agagctaggc aacgtactat tgttacacta 900
cctaagctcc ctgagcttga agtagtgcag ttaaatattt ctgcacacat ggattttggc 960
gaagccagac ttgacagcgt taccatcaat ggtaacacat cctattgtgt cactaagcct 1020
tacttcaggc ttgaaactaa ctttatgtgt acaggttgca ctatgaatct gcgcactgat 1080
acctgtagtt ttgacctgtc agcagtaaac aatggcatgt cattctctca attctgtcta 1140
agcactgaat ctggtgcttg tgagatgaaa attattgtta cctacgtatg gaattacttg 1200
ctaaggcagc gtttgtatgt tactgctgta gagggccaga ctcacactgg aaccacttca 1260
gtacatgcaa cagacacttc tagtgtaatc actgatgtct gcactgacta cactatctat 1320
ggagtctcag gtactggcat tattaagcca tcagatctct tattgcacaa tggcatagca 1380
ttcacctctc caacaggtga gctttatgca tttaaaaata taaccactgg caaaaccctt 1440
caggtcttac cgtgtgaaac cccttctcaa ctgattgtga taaacaacac cgttgtcggt 1500
gctatcacat ccagtaattc aactgaaaat aataggttta ctactactat tgtcacacct 1560
actttctttt attccacaaa tgccaccact ttcaactgca ctaagcctgt tttgtcctat 1620
ggacctatca gcgtgtgtag tgatggtgca attgtgggaa catccacatt acagaatact 1680
cgaccatcca tagtttcact atacgatggc gaagttgaaa taccatctgc attttctctt 1740
tccgttcaga cggagtactt gcaagttcaa gcagagcaag ttatagttga ttgtcctcag 1800
tatgtatgca atggcaacag ccgttgtcta caattactgg cacaatacac ctcagcttgc 1860
tctaacattg aagcagctct gcattcctct gcacagttgg atagcagaga gattataaat 1920
atgtttcaaa catcaacaca gtccttgcag ttagctaata ttaccaactt caagggtgac 1980
tacaatttta gcagcttact aaccaccaga attggtggca gatctgctat tgaagacctt 2040
ctttttaata aagttgttac tagtggcctt ggcactgttg atcaggacta caaatcctgc 2100
tctagagaca tggccatcgc tgacttagtt tgttcccagt attacaatgg catcatggtt 2160
ctacctggtg ttgttgatgc tgagaaaatg gcaatgtata ctggctctct tactggagct 2220
atggtatttg gaggactgac tgccgcagcg gcaataccat ttgccacggc agtacaagct 2280
cgcctcaatt atgtcgcact gcaaacaaat gtactacaag aaaaccagaa aattcttgca 2340
gaatcattta accaagcagt tggcaatata tcacttgcac tatcttctgt taatgatgcc 2400
atccagcaaa cttctgaggc tcttaacacc gtagctattg ctattaaaaa gattcaaaca 2460
gttgttaacc agcagggcga ggcattatca cacctgactg cacagctgtc aaacaatttt 2520
caagcaattt cgacttctat tcaagacatt tacaaccgtc ttgaggaagt agaggctaac 2580
cagcaagttg accgtctcat cacaggacgg ttggctgcac ttaatgcata tgttactcag 2640
ttactcaatc agatgtctca gattagacaa tctcgattgt tagctcagca aaagattaat 2700
gagtgtgtca aatcacagtc gtccagatac ggtttctgtg gaaatggcac acacatcttc 2760
tcacttacac agactgcacc aaatggcata tttttcatgc atgcagtgct agtacccaac 2820
aaattcacac gtgtcaacgc ttctgccggc atttgtgtgg ataatacgag aggctactca 2880
ttgcagcctc aacttatact ctaccagttt aataactcct ggagagttac acctagaaat 2940
atgtatgaac ccagactgcc ccggcaggct gatttcatac aattaactga ttgcagcgtt 3000
actttttaca acaccaccgc tgctaatctt cccaatatta tccctgacat tatagatgtc 3060
aatcaaacag tcagtgatat tattgacaat ttacctacag caacacctcc tcagtgggat 3120
gttggtatct ataacaacac tattctcaac ctcaccgttg agattaatga tctacaagag 3180
cggtctaaaa acctctcaca gattgcagat cgtttacaaa attatattga caatcttaac 3240
aatactctag ttgaccttga atggctcaac agagtggaaa cttaccttaa atggccgtgg 3300
tatatatggc ttgccattgc cctggctctt attgcatttg tgacaatcct cataacaatc 3360
tttcttagaa accgcaatcg tcagtactaa 3390
<210> 28
<211> 1129
<212> PRT
<213> Artificial Sequence
<220>
<223> PDCoVS BD Protein - expressed baculodisplay
<400> 28
Met Val Ser Ala Ile Val Leu Tyr Val Leu Leu Ala Ala Ala Ala His
1 5 10 15
Ser Ala Phe Ala Asp Asp Leu Leu Asp Leu Leu Thr Phe Pro Gly Ala
20 25 30
His Arg Phe Leu His Lys Pro Thr Arg Asn Ser Ser Ser Leu Tyr Ser
35 40 45
Arg Ala Asn Asn Asn Phe Asp Val Gly Val Leu Pro Gly Tyr Pro Thr
50 55 60
Lys Asn Val Asn Leu Phe Ser Pro Leu Thr Asn Ser Thr Leu Pro Ile
65 70 75 80
Asn Gly Leu His Arg Ser Tyr Gln Pro Leu Met Leu Asn Cys Leu Thr
85 90 95
Lys Ile Thr Asn His Thr Leu Ser Met Tyr Leu Leu Pro Ser Glu Ile
100 105 110
Gln Thr Tyr Ser Cys Gly Gly Ala Met Val Lys Tyr Gln Thr His Asp
115 120 125
Ala Val Arg Ile Ile Leu Asp Leu Thr Ala Thr Asp His Ile Ser Val
130 135 140
Glu Val Val Gly Gln His Gly Glu Asn Tyr Val Phe Val Cys Ser Glu
145 150 155 160
Gln Phe Asn Tyr Thr Thr Ala Leu His Asn Ser Thr Phe Phe Ser Leu
165 170 175
Asn Ser Glu Leu Tyr Cys Phe Thr Asn Asn Thr Tyr Leu Gly Ile Leu
180 185 190
Pro Pro Asp Leu Thr Asp Phe Thr Val Tyr Arg Thr Gly Gln Phe Tyr
195 200 205
Ala Asn Gly Tyr Leu Leu Gly Thr Leu Pro Ile Thr Val Asn Tyr Val
210 215 220
Arg Leu Tyr Arg Gly His Leu Ser Ala Asn Ser Ala Tyr Phe Ala Leu
225 230 235 240
Ala Asn Leu Thr Asp Thr Leu Ile Thr Leu Thr Asn Thr Thr Ile Ser
245 250 255
Gln Ile Thr Tyr Cys Asp Lys Ser Val Val Asp Ser Ile Ala Cys Gln
260 265 270
Arg Ser Ser His Glu Val Glu Asp Gly Phe Tyr Ser Asp Pro Lys Ser
275 280 285
Ala Val Arg Ala Arg Gln Arg Thr Ile Val Thr Leu Pro Lys Leu Pro
290 295 300
Glu Leu Glu Val Val Gln Leu Asn Ile Ser Ala His Met Asp Phe Gly
305 310 315 320
Glu Ala Arg Leu Asp Ser Val Thr Ile Asn Gly Asn Thr Ser Tyr Cys
325 330 335
Val Thr Lys Pro Tyr Phe Arg Leu Glu Thr Asn Phe Met Cys Thr Gly
340 345 350
Cys Thr Met Asn Leu Arg Thr Asp Thr Cys Ser Phe Asp Leu Ser Ala
355 360 365
Val Asn Asn Gly Met Ser Phe Ser Gln Phe Cys Leu Ser Thr Glu Ser
370 375 380
Gly Ala Cys Glu Met Lys Ile Ile Val Thr Tyr Val Trp Asn Tyr Leu
385 390 395 400
Leu Arg Gln Arg Leu Tyr Val Thr Ala Val Glu Gly Gln Thr His Thr
405 410 415
Gly Thr Thr Ser Val His Ala Thr Asp Thr Ser Ser Val Ile Thr Asp
420 425 430
Val Cys Thr Asp Tyr Thr Ile Tyr Gly Val Ser Gly Thr Gly Ile Ile
435 440 445
Lys Pro Ser Asp Leu Leu Leu His Asn Gly Ile Ala Phe Thr Ser Pro
450 455 460
Thr Gly Glu Leu Tyr Ala Phe Lys Asn Ile Thr Thr Gly Lys Thr Leu
465 470 475 480
Gln Val Leu Pro Cys Glu Thr Pro Ser Gln Leu Ile Val Ile Asn Asn
485 490 495
Thr Val Val Gly Ala Ile Thr Ser Ser Asn Ser Thr Glu Asn Asn Arg
500 505 510
Phe Thr Thr Thr Ile Val Thr Pro Thr Phe Phe Tyr Ser Thr Asn Ala
515 520 525
Thr Thr Phe Asn Cys Thr Lys Pro Val Leu Ser Tyr Gly Pro Ile Ser
530 535 540
Val Cys Ser Asp Gly Ala Ile Val Gly Thr Ser Thr Leu Gln Asn Thr
545 550 555 560
Arg Pro Ser Ile Val Ser Leu Tyr Asp Gly Glu Val Glu Ile Pro Ser
565 570 575
Ala Phe Ser Leu Ser Val Gln Thr Glu Tyr Leu Gln Val Gln Ala Glu
580 585 590
Gln Val Ile Val Asp Cys Pro Gln Tyr Val Cys Asn Gly Asn Ser Arg
595 600 605
Cys Leu Gln Leu Leu Ala Gln Tyr Thr Ser Ala Cys Ser Asn Ile Glu
610 615 620
Ala Ala Leu His Ser Ser Ala Gln Leu Asp Ser Arg Glu Ile Ile Asn
625 630 635 640
Met Phe Gln Thr Ser Thr Gln Ser Leu Gln Leu Ala Asn Ile Thr Asn
645 650 655
Phe Lys Gly Asp Tyr Asn Phe Ser Ser Leu Leu Thr Thr Arg Ile Gly
660 665 670
Gly Arg Ser Ala Ile Glu Asp Leu Leu Phe Asn Lys Val Val Thr Ser
675 680 685
Gly Leu Gly Thr Val Asp Gln Asp Tyr Lys Ser Cys Ser Arg Asp Met
690 695 700
Ala Ile Ala Asp Leu Val Cys Ser Gln Tyr Tyr Asn Gly Ile Met Val
705 710 715 720
Leu Pro Gly Val Val Asp Ala Glu Lys Met Ala Met Tyr Thr Gly Ser
725 730 735
Leu Thr Gly Ala Met Val Phe Gly Gly Leu Thr Ala Ala Ala Ala Ile
740 745 750
Pro Phe Ala Thr Ala Val Gln Ala Arg Leu Asn Tyr Val Ala Leu Gln
755 760 765
Thr Asn Val Leu Gln Glu Asn Gln Lys Ile Leu Ala Glu Ser Phe Asn
770 775 780
Gln Ala Val Gly Asn Ile Ser Leu Ala Leu Ser Ser Val Asn Asp Ala
785 790 795 800
Ile Gln Gln Thr Ser Glu Ala Leu Asn Thr Val Ala Ile Ala Ile Lys
805 810 815
Lys Ile Gln Thr Val Val Asn Gln Gln Gly Glu Ala Leu Ser His Leu
820 825 830
Thr Ala Gln Leu Ser Asn Asn Phe Gln Ala Ile Ser Thr Ser Ile Gln
835 840 845
Asp Ile Tyr Asn Arg Leu Glu Glu Val Glu Ala Asn Gln Gln Val Asp
850 855 860
Arg Leu Ile Thr Gly Arg Leu Ala Ala Leu Asn Ala Tyr Val Thr Gln
865 870 875 880
Leu Leu Asn Gln Met Ser Gln Ile Arg Gln Ser Arg Leu Leu Ala Gln
885 890 895
Gln Lys Ile Asn Glu Cys Val Lys Ser Gln Ser Ser Arg Tyr Gly Phe
900 905 910
Cys Gly Asn Gly Thr His Ile Phe Ser Leu Thr Gln Thr Ala Pro Asn
915 920 925
Gly Ile Phe Phe Met His Ala Val Leu Val Pro Asn Lys Phe Thr Arg
930 935 940
Val Asn Ala Ser Ala Gly Ile Cys Val Asp Asn Thr Arg Gly Tyr Ser
945 950 955 960
Leu Gln Pro Gln Leu Ile Leu Tyr Gln Phe Asn Asn Ser Trp Arg Val
965 970 975
Thr Pro Arg Asn Met Tyr Glu Pro Arg Leu Pro Arg Gln Ala Asp Phe
980 985 990
Ile Gln Leu Thr Asp Cys Ser Val Thr Phe Tyr Asn Thr Thr Ala Ala
995 1000 1005
Asn Leu Pro Asn Ile Ile Pro Asp Ile Ile Asp Val Asn Gln Thr
1010 1015 1020
Val Ser Asp Ile Ile Asp Asn Leu Pro Thr Ala Thr Pro Pro Gln
1025 1030 1035
Trp Asp Val Gly Ile Tyr Asn Asn Thr Ile Leu Asn Leu Thr Val
1040 1045 1050
Glu Ile Asn Asp Leu Gln Glu Arg Ser Lys Asn Leu Ser Gln Ile
1055 1060 1065
Ala Asp Arg Leu Gln Asn Tyr Ile Asp Asn Leu Asn Asn Thr Leu
1070 1075 1080
Val Asp Leu Glu Trp Leu Asn Arg Val Glu Thr Tyr Leu Lys Trp
1085 1090 1095
Pro Trp Tyr Ile Trp Leu Ala Ile Ala Leu Ala Leu Ile Ala Phe
1100 1105 1110
Val Thr Ile Leu Ile Thr Ile Phe Leu Arg Asn Arg Asn Arg Gln
1115 1120 1125
Tyr
<210> 29
<211> 27960
<212> DNA
<213> Artificial Sequence
<220>
<223> Complete Genome Nucleic Acid (cDNA) Sequence equivalent of the
(+) single-stranded RNA genome of PEDV Isolate 1251-125-10
(125-10)
<400> 29
agactcttgt ctactcaatt caactaaacg aaattttgtc cttccggccg catgtccatg 60
ctgctggaag ctgacgtgga atttcattag gtttgcttaa gtagccatcg caagtgctgt 120
gctgtcctct agttcctggt tggcgttccg tcgccttcta catactagac aaacagcctt 180
cctccggttc cgtctggggg ttgtgtggat aactagttcc gtctagtttg aaaccagtaa 240
ctgtcggcta tggctagcaa ccatgttaca ttggcttttg ccaatgatgc agaaatttca 300
gcttttggct tttgcactgc tagtgaagcc gtctcatact attctgaggc cgccgctagt 360
ggatttatgc aatgccgttt cgtgtccttc gatctcgctg acactgttga gggattgctt 420
cccgaagact atgtcatggt ggtggtcggc actaccaagc ttagtgcgta tgtggacact 480
tttggtagcc gccccaaaaa catttgtggt tggctgttat tttctaactg taattacttc 540
ctcgaagagt tagagcttac ttttggtcgt cgtggtggta acatcgtgcc agttgaccaa 600
tacatgtgtg gcgctgacgg taaacctgtt cttcaggaat ccgaatggga gtatacagat 660
ttctttgctg actccgaaga cggtcaactc aacattgctg gtatcactta tgtgaaggcc 720
tggattgtag agcgatcgga tgtctcttat gcgagtcaga atttaacatc tattaagtct 780
attacttact gttcaaccta tgagcatact tttcctgatg gtactgccat gaaggttgca 840
cgtactccaa agattaagaa gactgttgtc ttgtctgagc cacttgctac tatctacagg 900
gaaattggtt ctccttttgt ggataatggg agcgatgctc gttctatcat taagagacca 960
gtgttcctcc acgcttttgt taagtgtaag tgtggtagtt atcattggac tgttggtgat 1020
tggacttcct atgtctccac ttgctgtggc tttaagtgta agccagtcct tgtggcttca 1080
tgctctgcta cgcctggttc tgttgtggtt acgcgcgctg gtgctggcac tggtgttaag 1140
tattacaaca acatgttcct gcgccatgtg gcagacattg atgggttggc attctggcga 1200
attctcaagg tgcagtccaa agacgacctc gcttgctctg gtaaattcct tgaacaccat 1260
gaggaaggtt tcacagatcc ttgctacttt ttgaatgact cgagcattgc tactaagctc 1320
aagtttgaca tccttagtgg caagttttct gatgaagtca aacaagctat ctttgctggt 1380
catgttgttg ttggcagcgc gctcgttgac attgttgacg atgcactggg acagccttgg 1440
tttatacgta agcttggtga ccttgcaagt gcagcttggg agcagcttaa ggctgtcgtt 1500
agaggcctta acctcctgtc tgatgaggtc gtgctctttg gcaaaagact tagctgtgcc 1560
actcttagta tcgttaacgg tgtttttgag ttcatcgccg aagtgcctga gaagttggct 1620
gcggctgtta cagtttttgt caacttcttg aatgagcttt ttgagtctgc ctgtgactgc 1680
ttaaaggtcg gaggtaaaac ctttaacaag gttggctctt atgttctttt tgacaacgca 1740
ttggttaagc ttgtcaaggc aaaagttcgc ggcccacgac aggcaggtgt ttgtgaagtt 1800
cgttacacaa gccttgttat tgggagtact accaaggtgg tttccaagcg cgttgaaaat 1860
gccaatgtga atctcgtcgt cgttgacgag gatgtgaccc tcaacaccac tggtcgtaca 1920
gttgttgttg acggacttgc attcttcgag agtgacgggt tttacagaca tcttgctgat 1980
gctgacgttg tcattgaaca tcctgtttat aagtctgctt gtgagctcaa gccagttttt 2040
gagtgtgacc caatacctga ttttcctatg cctgtggccg ctagtgttgc agagctttgt 2100
gtgcaaactg atctgttgct taaaaattac aacactcctt ataaaactta cagctgcgtt 2160
gtgagaggtg ataagtgttg tatcacttgc accttacatt tcacagcacc aagttatatg 2220
gaggctgctg ctaattttgt agacctctgt accaagaaca ttggtactgc tggttttcat 2280
gagttttaca ttacggccca tgaacaacag gatctgcaag ggttcgtaac cacttgttgc 2340
acgatgtcag gttttgagtg ttttatgcct ataatcccac agtgtccagc agtgcttgaa 2400
gagattgatg gtggtagcat ctggcggtct tttatcactg gtcttaatac aatgtgggat 2460
ttttgcaagc atcttaaagt cagctttgga ctagatggca ttgttgtcac tgtagcacgc 2520
aaatttaaac gacttggtgc tctcttggca gaaatgtata acacttacct ttcaactgtg 2580
gtggaaaact tggtactggc cggtgttagc ttcaagtatt atgccaccag tgtcccaaaa 2640
attgttttgg gctgttgttt tcacagtgtt aaaagtgttc ttgcaagtgc cttccagatt 2700
cctgtccagg caggcgttga gaagtttaaa gtcttcctta actgtgttca ccctgttgta 2760
ccacgtgtca ttgaaacttc ttttgtggaa ttagaagaga cgacatttaa accaccagca 2820
ctcaatggta gtattgctat tgttgatggc tttgctttct attatgatgg aacactatac 2880
tatcccaccg atggtaatag cgttgttcct atctgcttta agaagaaagg tggtggtgat 2940
gtcaaattct ctgatgaagt ctctgttaaa accattgacc cagtttataa ggtctccctt 3000
gaatttgagt tcgagtctga gactattatg gctgtgctta ataaggctgt tggtaattgt 3060
atcaaggtta caggtggttg ggacgatgtt gttgagtata tcaatgttgc cattgaggtt 3120
cttaaagatc acatcgatgt gcctaagtac tacatctatg atgaggaagg tggcaccgat 3180
cctaatctgc ccgtaatggt ttctcagtgg ccgttgaatg atgacacgat ctcacaggat 3240
ctgcttgatg ttgaagttgt tactgatgcg ccagttgatt tcgagggtga tgaagtagac 3300
tcctctgacc ctgataaggt ggcagacgtg gctaactctg agcctgagga tgacggtctt 3360
aatgtagctc ctgaaacaaa tgtagagtct gaagttgagg aagttgccgc aaccttgtcc 3420
tttattaaag atacaccttc cacagttact aaggatcctt ttgcttttga ctttgcaagc 3480
tatggaggac ttaaggtttt aagacaatct cataacaact gctgggttac ttctaccttg 3540
gtgcagctac aattgcttgg catcgttgat gaccctgcaa tggagctttt tagtgctggt 3600
agagttggtc caatggttcg caaatgctat gagtcacaaa aggctatctt gggatctttg 3660
ggtgatgtgt cggcttgcct agagtctctg actaaggacc tacacacact taagattacc 3720
tgttctgtag tctgtggttg tggtactggt gaacgtatct atgatggttg tgcttttcgt 3780
atgacgccaa ctttggaacc gttcccatat ggtgcttgtg ctcagtgtgc tcaagttttg 3840
atgcacactt ttaaaagtat tgttggcacc ggcatctttt gtcgagatac tactgctctc 3900
tccttggatt ctttggttgt aaaacctctt tgtgcggctg cttttatagg caaggatagt 3960
ggtcattatg tcactaactt ttatgatgct gctatggcta ttgatggtta tggtcgtcat 4020
cagataaagt atgacacact gaacactatt tgtgttaaag acgttaattg gacagcacct 4080
tttgtcccag acgttgagcc tgtattggag cctgttgtca aacctttcta ttcttataag 4140
aatgttgatt tttaccaagg agattttagt gaccttgtta aacttccatg tgattttgtt 4200
gttaatgctg caaatgagaa tttgtctcac ggtggcggca tagcaaaggc cattgatgtt 4260
tataccaagg gcatgttgca gaagtgctcg aatgattaca ttaaagcaca cggtcccatt 4320
aaagttggac gtggtgtcat gttggaggca ttaggtctta aggtctttaa tgttgttggt 4380
ccacgtaagg gtaagcatgc acctgagctt cttgttaagg cttataagtc cgtttttgct 4440
aattcaggtg ttgctcttac acctttgatt agtgttggaa tttttagtgt tcctttggaa 4500
gaatctttat ctgcttttct tgcatgtgtt ggtgatcgcc actgtaagtg cttttgttat 4560
agtgacaaag agcgcgaggc gatcattaat tacatggatg gcttggtaga tgctattttc 4620
aaagatgcac ttgttgatac tactcctgtc caggaagatg ttcaacaagt ttcacaaaaa 4680
ccagttttgc ctaattttga acctttcagg attgaaggtg ctcatgcttt ctatgagtgc 4740
aaccctgaag gtttgatgtc attaggtgct gacaagctgg tgttgtttac aaattccaat 4800
ttggattttt gtagcgttgg taagtgtctt aacaatgtga ctggcggtgc attgcttgaa 4860
gccataaatg tatttaaaaa gagtaacaaa acagtgcctg ctggcaactg tgttactttt 4920
gagtgtgcag atatgatttc tattactatg gtagtattgc catctgacgg tgatgctaat 4980
tatgacaaaa attatgcacg cgccgtcgtc aaggtatcta agcttaaagg caagttattg 5040
cttgctgttg gtgatgccat gttgtattcc aagttgtccc acctcagcgt gttaggtttc 5100
gtatccacac ctgatgatgt ggagcgtttc tacgcaaata agagtgtggt tattaaagtt 5160
actgaggata cacgtagtgt taagactgtt aaagtagaat ccactgttac ttatggacaa 5220
caaattggac cttgtcttgt taatgacacc gttgtcacag acaacaaacc tgttgttgct 5280
gatgttgtag ctaaggttgt accaagtgct aattgggatt cacattatgg ttttgataag 5340
gctggtgagt tccacatgct agaccatact gggtttgcct ttcctagtga agttgttaac 5400
ggtaggcgtg tgcttaaaac cacagataat aactgttggg ttaatgttac atgtttacaa 5460
ttacagtttg ctagatttag gttcaagtca gcaggtctac aggctatgtg ggagtcctat 5520
tgtactggtg atgttgctat gtttgtgcat tggttgtact ggcttactgg tgttgacaaa 5580
ggtcagccta gtgattcaga aaatgcactt aacatgttgt ctaagtacat tgttcctgct 5640
ggttctgtca ctattgaacg tgtcacgcat gacggttgtt gttgtagtaa gcgtgttgtc 5700
actgcaccag ttgtgaatgc tagcgtgttg aagcttggcg tcgaggatgg tctttgtcca 5760
catggtctta actacattga caaagttgtt gtagttaaag gtactacaat tgttgtcaat 5820
gttggaaaac ctgtagtggc accatcgcac ctctttctta agggtgtttc ctacacaaca 5880
ttcctagata atggtaacgg tgttgccggc cattatactg tttttgatca tgacactggt 5940
atggtgcatg atggagatgt ttttgtacca ggtgatctca atgtgtctcc tgttacaaat 6000
gttgtcgtct cagagcagac ggctgttgtg attaaagacc ctgtgaagaa agtagagtta 6060
gacgctacaa agctgttaga cactatgaat tatgcatcgg aaagattctt ttcctttggt 6120
gattttatgt cacgtaattt aattacagtg tttttgtaca tccttagtat tttgggtctc 6180
tgttttaggg cctttcgtaa gagggatgtt aaagttctag ctggtgtacc ccaacgtact 6240
ggtattatat tgcgtaaaag tgtgcgctat aatgcaaagg ctttgggtgt cttcttcaag 6300
ctaaaacttt attggttcaa agttcttggt aagtttagtt tgggtattta tgcattgtat 6360
gcattactat tcatgacaat acgctttaca cctataggtg gccctgtttg tgatgatgtt 6420
gttgctggtt atgctaattc tagttttgac aagaatgagt attgcaacag tgttatttgt 6480
aaggtctgtc tctatgggta ccaggaactt tcggacttct ctcacacaca ggtagtatgg 6540
caacacctta gagacccatt aattggtaat gtgatgcctt tcttttattt ggcatttctg 6600
gcaatttttg ggggtgttta tgtaaaggct attactctct attttatttt ccagtatctt 6660
aacatacttg gtgtgttttt gggcctacaa cagtccattt ggtttttgca gcttgtgcct 6720
tttgatgtct ttggtgacga gatcgtcgtc tttttcatcg ttacacgcgt attgatgttc 6780
cttaagcatg ttttccttgg ctgcgataag gcatcttgtg tggcttgctc taagagtgct 6840
cgccttaagc gcgttcctgt ccagactatt tttcagggta ctagcaaatc cttctacgta 6900
catgccaatg gtggttctaa gttctgtaag aagcacaatt tcttttgttt aaattgtgat 6960
tcttatggtc caggctgcac ttttattaat gacgtcattg caactgaagt tggtaatgtt 7020
gtcaaactta atgtgcaacc gacaggtcct gccactattc ttattgacaa ggttgaattc 7080
agtaatggtt tttactatct ttatagtggt gacacatttt ggaagtacaa ctttgacata 7140
acagataaca aatacacttg caaagagtca cttaaaaatt gtagcataat cacagacttt 7200
attgttttta acaataatgg ttccaatgta aatcaggtta agaatgcatg tgtgtatttt 7260
tcacagatgc tttgtaaacc tgttaagtta gtggactcag cgttgttggc cagtttgtct 7320
gttgattttg gtgcaagctt acatagtgct tttgttagtg tgttgtcgaa tagttttggc 7380
aaagacctgt caagttgtaa tgacatgcag gattgcaaga gcacattggg ttttgatgat 7440
gtaccattgg atacctttaa tgctgctgtt gctgaggctc atcgttacga tgtcctcttg 7500
actgacatgt cgttcaacaa ttttaccacc agttatgcaa aaccagagga aaaacttccc 7560
gtccatgaca ttgccacgtg tatgcgtgta ggtgccaaga ttgttaatca taacgttctt 7620
gtcaaggata gtatacctgt ggtgtggctt gtacgtgatt tcattgccct ttctgaagaa 7680
actaggaagt acattattcg tacgactaaa gttaagggta taaccttcat gttgaccttt 7740
aatgattgtc gtatgcatac taccatacct actgtttgca ttgcaaataa gaagggtgca 7800
ggtcttccta gtttttcaaa ggttaagaaa ttcttctggt ttttgtgtct gttcatagtt 7860
gctgttttct ttgcactaag cttttttgat tttagtactc aggttagcag tgatagtgat 7920
tatgacttca agtatattga gagtggccag ttgaagactt ttgacaatcc acttagttgt 7980
gtgcataatg tctttagtaa cttcgaccag tggcatgatg ccaagtttgg tttcaccccc 8040
gtcaacaatc ctagttgtcc tatagtcgtt ggtgtatcag acgaagcgcg cactgttcca 8100
ggtatcccag caggtgttta tttagctggt aaaacacttg tttttgctat taacaccatt 8160
tttggtacat ctggtttgtg ctttgatgct agtggcgttg ctgataaggg cgcttgcatt 8220
tttaattcgg cttgcaccac attatctggt ttgggtggaa ctgctgtcta ctgttataag 8280
aatggtctag ttgaaggtgc taaactttat agtgagttgg cacctcatag ctactataaa 8340
atggtagatg gtaatgctgt gtctttacct gaaattatct cacgcggctt tggcatccgt 8400
actatccgta caaaggctat gacctactgt cgcgttggcc agtgtgtgca atctgcagaa 8460
ggtgtttgtt ttggcgccga tagattcttt gtctataatg cagaatctgg ttctgacttt 8520
gtttgtggca cagggctctt tacattgttg atgaacgtta ttagtgtttt ttccaagaca 8580
gtaccagtaa ctgtgttgtc tggtcaaata ctttttaatt gcattattgc ttttgctgct 8640
gttgcggtgt gtttcttatt tacaaagttt aagcgcatgt tcggtgatat gtctgttggc 8700
gttttcactg tcggtgcttg tactttgttg aacaatgttt cctacattgt aacacagaac 8760
acacttggca tgttgggcta tgcaactttg tactttttgt gcactaaagg tgttagatat 8820
atgtggattt ggcatttggg atttttgatc tcatatatac ttattgcacc atggtgggtt 8880
ttgatggttt atgccttttc agccattttt gagtttatgc ctaacctttt taagcttaag 8940
gtttcaacac aactttttga gggtgacaag ttcgtaggct cttttgaaaa tgctgcagca 9000
ggtacatttg tgcttgatat gcatgcctat gagagacttg ccaactctat ctcaactgaa 9060
aaactgcgtc agtatgctag tacttacaat aagtacaagt attattcagg cagtgcttca 9120
gaggctgatt acaggcttgc ttgttttgcc catttggcca aggctatgat ggattatgct 9180
tctaatcaca acgacacgtt atacacacca cccactgtga gttacaattc aactctacag 9240
gctggcttgc gtaagatggc acaaccatct ggtgttgttg agaagtgcat agttcgtgtt 9300
tgctatggta atatggctct taatggccta tggcttggtg atactgttat ctgcccacgc 9360
catgttatag cgtctagtac tactagcact atagattatg actatgccct ttctgtttta 9420
cgcctccaca acttctccat ttcatctggt aatgttttcc taggtgttgt gggtgtaacc 9480
atgcgaggtg ctttgttgca gataaaggtt aatcaaaaca atgtccacac gcctaagtac 9540
acctatcgca cagttagacc gggtgaatct tttaatatct tggcgtgcta tgatggttct 9600
gcagctggtg tttacggcgt taacatgcgc tctaattaca ctattagagg ctcgttcatt 9660
aatggcgctt gtggttcacc tggttataac attaacaatg gtaccgttga gttttgctat 9720
ttacaccagc ttgaacttgg ttcaggctgt catgttggta gcgacttaga tggtgttatg 9780
tatggtggtt atgaggacca acctactttg caagttgaag gcgctagtag tctgtttaca 9840
gagaatgtgt tggcatttct ttatgcagca ctcattaatg gttctacctg gtggcttagt 9900
tcttctagga ttgctgtaga caggtttaat gagtgggctg ttcataatgg tatgacaaca 9960
gtagttaata ctgattgctt ttctattctt gctgctaaga ctggtgttga tgtacaacgt 10020
ttgttggcct caatccagtc tctgcataag aattttggtg gaaagcaaat tcttggctat 10080
acctcgttga cagatgagtt tactacaggt gaagttatac gtcaaatgta tggcgttaat 10140
cttcagagtg gttatgtttc acgcgcctgt agaaatgtct tgctggttgg ttcttttctg 10200
actttctttt ggtcagaatt agtttcctac actaagttct tttgggtaaa tcctggttat 10260
gtcacaccta tgtttgcgtg tttgtcattg ctgtcctcac ttttgatgtt cacactcaag 10320
cataagacat tgtttttcca ggtctttcta atacctgctc tgattgttac atcttgcatt 10380
aatttggcat ttgatgttga agtctacaac tatttggcag agcattttga ttaccatgtt 10440
tctctcatgg gttttaatgc acaaggtctt gttaacatct ttgtctgctt tgttgttacc 10500
attttacacg gcacatacac atggcgcttt tttaacacac ctgtgagttc tgtcacttat 10560
gtggtagctt tgctgactgc ggcatataac tatttttacg ctagtgacat tcttagttgt 10620
gctatgacac tatttgctag tgtgactggc aactggttcg ttggtgctgt ttgttataaa 10680
gctgctgttt atatggcctt gagatttcct acttttgtgg ctatttttgg tgatattaag 10740
agtgttatgt tctgttacct tgtgttgggt tattttacct gttgcttcta cggtattctc 10800
tactggttca acaggttttt taaggttagt gtaggtgtct atgactatac tgttagtgct 10860
gctgagttta agtatatggt tgctaacggc ctacgtgcac caactggaac acttgattca 10920
ctacttctgt ctgccaaatt gattggtatt ggtggtgagc ggaatattaa gatttcttcc 10980
gttcagtcta aactgactga tattaagtgt agtaacgttg tgcttttagg ctgtctctct 11040
agcatgaatg tctcagcaaa ttcaacagaa tgggcctatt gtgttgactt gcataacaag 11100
atcaacttgt gtaatgaccc agaaaaagcg caggaaatgc tacttgcttt gttggcattt 11160
ttccttagta agaatagtgc ttttggttta gatgacttat tggaatccta ttttaatgac 11220
aatagtatgt tgcagagtgt tgcatctact tatgtcggtt tgccttctta tgtcatttat 11280
gaaaatgcac gccaacagta tgaagatgct gttaataatg gttctccacc tcagttggtt 11340
aagcaattgc gccatgccat gaatgtagca aagagcgaat ttgaccgtga ggcttctact 11400
cagcgtaagc ttgatagaat ggcggaacag gctgcagcac agatgtacaa agaggcacga 11460
gcagttaata ggaagtccaa agttgtaagt gctatgcatt cactgctttt tggtatgttg 11520
agacgtttgg acatgtcttc tgtagacacc attctcaact tggcaaagga tggggttgta 11580
cctctgtctg tcataccggc agtcagtgct actaagctta acattgttac ttctgatatc 11640
gattcttata atcgtatcca gcgtgaggga tgtgtccact acgctggtac catttggaat 11700
ataattgata tcaaggacaa tgatggcaag gtggtacacg ttaaggaggt aaccgcacag 11760
aatgctgagt ccctgtcatg gcccctggtc cttgggtgtg agcgtattgt caagctccag 11820
aataatgaaa ttattcctgg taagctgaag cagcgctcca ttaaggcaga aggagatggc 11880
atagttggag aaggtaaggc actttacaat aatgagggtg gacgtacttt tatgtatgct 11940
ttcatctcgg acaaaccgga cctgcgtgta gtcaagtggg agttcgatgg tggttgtaac 12000
actattgagc tagaaccacc acgtaagttc ttggtggatt ctcctaatgg tgcacagatc 12060
aagtatctct actttgttcg taaccttaac acgttacgta ggggtgctgt tctcggctac 12120
ataggtgcca ctgtacgctt gcaggctggt aaacaaacag aacaggctat taactcttca 12180
ttgttgacac tttgcgcttt cgctgtggat cctgctaaga cctacatcga tgctgtcaaa 12240
agtggtcaca aaccagtagg taactgtgtt aagatgttgg ccaatggttc tggtaatgga 12300
caagctgtta ctaatggtgt ggaggctagt actaaccagg attcatacgg tggtgcgtcc 12360
gtgtgtctat attgtagagc acatgttgag catccatcta tggatggttt ttgcagactg 12420
aaaggcaagt acgtacaggt tccactaggt acagtggatc ctatacgttt tgtacttgag 12480
aatgacgttt gcaaggtttg tggttgttgg ctggctaatg gctgcacttg tgacagatcc 12540
attatgcaaa gcactgatat ggcttattta aacgagtacg gggctctagt gcagctcgac 12600
tagagccctg taacggtact gatacacaac atgtgtatcg tgcttttgac atctacaaca 12660
aggatgttgc ttgtctaggt aaattcctca aggtgaactg tgttcgcctg aagaatttgg 12720
ataagcatga tgcattctat gttgtcaaaa gatgtaccaa gtctgcgatg gaacacgagc 12780
aatccatcta tagcagactt gaaaagtgtg gagccgtagc cgaacacgat ttcttcactt 12840
ggaaggatgg tcgtgccatc tatggtaacg tttgtagaaa ggatcttacc gagtatacta 12900
tgatggattt gtgttacgct ttacgtaact ttgatgaaaa caattgcgat gttcttaaga 12960
gcattttaat taaggtaggc gcttgtgagg agtcctactt caataataaa gtctggtttg 13020
accctgttga aaatgaagac attcatcgtg tctatgcatt gttaggtacc attgtttcac 13080
gtgctatgct taaatgcgtt aagttctgtg atgcaatggt tgaacaaggt atagttggtg 13140
ttgtcacatt agataatcag gatcttaatg gtgattttta tgattttggt gattttactt 13200
gtagcatcaa gggaatgggt atacccattt gcacatcata ttactcttat atgatgcctg 13260
ttatgggtat gactaattgc cttgctagtg agtgttttgt taagagtgat atatttggtg 13320
aggatttcaa gtcatatgac ctgctggaat atgatttcac ggagcataag acagcactct 13380
tcaacaagta tttcaagtat tggggactgc aataccaccc taactgtgtg gactgcagtg 13440
atgagcagtg catagttcac tgtgccaact tcaatacgtt gttttccact actataccta 13500
ttacggcatt tggacctttg tgtcgcaagt gttggattga tggtgttcca ctggtaacta 13560
cagctggtta tcattttaaa cagttaggta tagtttggaa caatgacctc aacttacact 13620
ctagcaggct ctctattaac gaattactcc agttttgtag tgatcctgca ttgcttatag 13680
catcatcacc agcccttgtt gatcagcgta ctgtttgctt ttcagttgca gcgctaggta 13740
caggtatgac taaccagact gttaaacctg gccatttcaa taaggagttt tatgacttct 13800
tacttgagca aggtttcttt tctgagggct ctgagcttac tttaaagcac ttcttctttg 13860
cacagaaggg tgatgcagct gttaaggatt ttgactacta taggtataat agacctactg 13920
ttctggacat ttgccaagct cgcgtcgtgt atcaaatagt gcaacgctat tttgatattt 13980
acgaaggtgg ttgtatcact gctaaagagg tggttgttac aaaccttaac aagagcgcag 14040
gttatccttt gaacaagttt ggtaaagctg gtctttacta tgagtcttta tcctatgagg 14100
aacaggatga actttatgct tatactaagc gtaacatcct gcccactatg acacagctca 14160
accttaaata tgctataagt ggcaaagaac gtgcacgcac agtgggtggt gtttcgcttt 14220
tgtcaaccat gactactcgg cagtatcatc agaaacacct taagtccata gttaatacta 14280
ggggcgcttc ggttgttatt ggtactacta agttttatgg tggttgggac aatatgctta 14340
agaaccttat tgatggtgtt gaaaatccgt gtcttatggg ttgggactac ccaaagtgcg 14400
acagagcact gcccaatatg atacgtatga tttcagccat gattttaggc tctaagcaca 14460
ccacatgctg cagttccact gaccgctttt tcaggttgtg caatgaattg gctcaagtcc 14520
ttactgaggt tgtttattct aatggaggtt tttatttgaa gccaggtggt actacctctg 14580
gtgatgcaac caccgcatat gcaaactcag tttttaatat cttccaagca gtaagtgcca 14640
atgttaacaa acttcttagt gttgacagca atgtctgtca taatttagaa gttaagcaat 14700
tgcagcgtaa gctttatgag tgctgttata gatcaactac cgtcgatgac cagttcgtcg 14760
ttgagtatta tggttacttg cgtaaacatt tttcaatgat gattctttct gatgatggcg 14820
ttgtttgtta taacaatgac tatgcatcac ttggttatgt cgctgatctt aacgcattca 14880
aggctgtttt gtattaccag aacaatgtct tcatgagcgc ctctaaatgt tggatcgagc 14940
ctgacattaa taaaggtcct catgaatttt gctcgcagca tactatgcag attgtcgata 15000
aagatggtac ttattacctt ccttaccctg atccttcaag aattctctct gcaggtgtgt 15060
ttgttgatga cgttgttaaa actgatgcag ttgtattgct tgaacgttat gtgtcattgg 15120
ctatagatgc ctacccgtta tctaagcatg aaaaccctga atataagaag gtgttttatg 15180
tgcttttgga ttgggttaag catctgtaca aaactcttaa tgctggtgtg ttagagtctt 15240
tttctgtcac acttttggaa gattctactg ctaaattctg ggatgagagc ttttatgcca 15300
acatgtatga gaaatctgca gttttacaat ctgcagggct ttgtgttgtt tgtggctctc 15360
aaactgtttt acgttgtggt gattgtctac ggcgtcctat gctttgtact aagtgtgctt 15420
atgatcatgt cattggaaca actcacaagt tcattttggc catcactcca tatgtgtgtt 15480
gtgcttcaga ttgtggtgtc aatgatgtaa ctaagctcta cttaggtggt cttagttatt 15540
ggtgtcatga ccacaagcca cgtcttgcat tcccgttgtg ctctgctggt aatgtttttg 15600
gcttgtacaa aaattctgct accggctcac ccgatgttga agactttaat cgcattgcta 15660
catccgattg gactgatgtt tctgactaca ggttggcaaa tgatgtcaag gactcattgc 15720
gtctgtttgc agcggaaact atcaaggcca aggaggagag cgttaagtca tcctatgctt 15780
gtgcaacact acatgaggtt gtaggaccta aagagttgtt gctcaaatgg gaagtcggca 15840
gacccaaacc accccttaat agaaattcgg ttttcacttg ttatcatata acgaagaaca 15900
ccaaatttca aatcggtgag tttgtgtttg agaaggcaga atatgataat gatgctgtaa 15960
catataaaac taccgccaca acaaaacttg ttcctggcat ggtttttgtg cttacctcac 16020
ataatgttca gccattgcgc gcaccgacca ttgctaatca agaacgttat tccactatac 16080
ataagttgca tcctgctttt aacatacctg aagcttattc tagcttagtg ccctattacc 16140
aattgattgg taagcagaag attacaacta ttcagggacc tcccggtagt ggtaaatctc 16200
actgtgttat agggctaggt ttgtactatc caggtgcacg tatagtgttt acagcttgtt 16260
ctcatgcagc ggtcgattca ctttgtgtga aagcttccac tgcttatagc aatgacaaat 16320
gttcacgcat cataccacag cgcgctcgtg ttgagtgtta tgatggtttc aagtctaata 16380
atactagtgc tcagtacctt ttctctactg tcaatgcttt gccagagtgc aatgcggaca 16440
ttgttgtggt ggatgaggtc tctatgtgca ctaattatga cttgtctgtc ataaatcagc 16500
gcatcagcta taggcatgta gtctatgttg gtgaccctca acagctgcct gcaccacgtg 16560
ttatgatttc acgtggtact ttggaaccaa aggactacaa cgttgtcact caacgcatgt 16620
gtgcccttaa gcctgatgtt ttcttgcaca agtgttatcg ctgtcctgct gagatagtgc 16680
gtactgtgtc tgagatggtc tatgaaaacc aattcattcc tgtgcaccca gatagcaagc 16740
agtgttttaa aatcttttgc aagggtaatg ttcaggttga taatggttca agcattaatc 16800
gcaggcaatt ggatgttgtg cgtatgtttt tggctaaaaa tcctaggtgg tcaaaggctg 16860
tttttatttc tccttataac agccagaatt atgttgccag ccgcatgcta ggtctacaaa 16920
ttcagacagt tgactcatcc cagggtagtg agtatgacta tgtcatttac acacaaactt 16980
cagatactgc ccatgcctgt aatgttaaca ggtttaatgt tgccatcaca agggccaaga 17040
aaggcatatt atgtataatg tgcgataggt ccctttttga tgtgcttaaa ttctttgagc 17100
ttaaattgtc tgatttgcag gctaatgagg gttgtggtct ttttaaagac tgtagcagag 17160
gtgatgatct gttgccacca tctcacgcta acaccttcat gtctttagcg gacaatttta 17220
agactgatca agatcttgct gttcaaatag gtgttaatgg acccattaaa tatgagcatg 17280
ttatctcgtt tatgggtttc cgttttgata tcaacatacc caaccatcat actctctttt 17340
gcacacgcga ctttgccatg cgcaatgtta gaggttggtt aggctttgac gttgaaggag 17400
cacatgttgt tggctctaac gtcggtacaa atgtcccatt gcaattaggg ttttctaacg 17460
gtgttgattt tgttgtcaga cctgaaggtt gcgttgtaac agagtctggt gactacatta 17520
aacccgtcag agctcgtgct ccaccagggg aacaattcgc acaccttttg cctttactta 17580
aacgcggcca accatgggat gttgtccgca aacgtatagt gcagatgtgt agtgactacc 17640
tggccaacct atcagacata ctaatttttg tgttgtgggc tggtggtttg gagttgacaa 17700
ctatgcgtta ttttgtcaag attggaccaa gtaagagttg tgattgtggt aaggttgcta 17760
cttgttacaa tagtgcgctg catacgtact gttgtttcaa acatgccctt ggttgtgatt 17820
atctgtataa cccatactgt attgatatac agcagtgggg atacaaggga tcacttagcc 17880
ttaaccacca tgagcattgt aatgtacata gaaacgagca tgtggcttct ggtgatgcca 17940
taatgactcg ctgtctggcc atacatgatt gctttgtcaa gaacgttgac tggtccatca 18000
catacccatt tattggtaat gaggctgtta ttaataagag cggccgaatt gtgcaatcac 18060
acactatgcg gtcagttctt aagttataca atccgaaagc catatatgat attggcaatc 18120
ctaagggcat tagatgtgcc gtaacggatg ctaagtggtt ttgctttgac aagaatccta 18180
ctaattctaa tgtcaagaca ttggagtatg actatataac acatggccaa tttgatgggt 18240
tgtgcttgtt ttggaattgc aatgtagaca tgtatccaga attttctgtg gtctgtcgtt 18300
ttgatactcg ctgtaggtca ccactcaact tggagggttg taatggtggt tcactgtatg 18360
ttaataatca tgcattccat acaccggctt ttgacaagcg tgcttttgct aagttgaagc 18420
caatgccatt tttcttttat gatgatactg agtgtgacaa gttacaggac tccataaact 18480
atgttcctct tagggctagt aactgcatta ctaaatgtaa tgttggtggt gctgtctgta 18540
gtaagcattg tgctatgtat catagctatg ttaatgctta caacactttt acgtcggcgg 18600
gctttactat ttgggtgcct acttcgtttg acacctataa tctgtggcag acatttagta 18660
acaatttgca aggtcttgag aacattgctt tcaatgtcgt aaagaaagga tcttttgttg 18720
gtgccgaagg tgaacttcct gtagctgtgg ttaatgacaa agtgctcgtt agagatggta 18780
ctgttgatac tcttgttttt acaaacaaga catcactacc cactaacgta gcttttgagt 18840
tgtatgccaa gcgtaaggta ggactcaccc cacccattac gatcctacgt aacttgggtg 18900
tagtttgtac atctaagtgt gtcatttggg actatgaagc cgaacgtcca cttactactt 18960
ttacaaagga tgtttgtaaa tataccgact ttgagggtga cgtctgtaca ctctttgata 19020
acagcattgt tggttcatta gagcgattct ccatgaccca aaatgctgtg cttatgtcac 19080
ttacagctgt taaaaagctt actggcataa agttaactta tggttatctt aatggtgtcc 19140
cagttaacac acatgaagat aaacctttta cttggtatat ttacactagg aagaacggca 19200
agttcgagga ccatcctgat ggctatttta cccaaggtag aacaaccgct gattttagcc 19260
ctcgtagcga catggaaaag gacttcctaa gtatggatat gggtctgttt attaacaagt 19320
acggacttga agattacggc tttgagcacg ttgtgtatgg tgatgtttca aaaaccaccc 19380
ttggtggttt gcatctacta atttcgcagg tgcgtctggc ctgtatgggt gtgctcaaaa 19440
tagacgagtt tgtgtctagt aatgatagca cgttaaagtc ttgtactgtt acatatgctg 19500
ataaccctag tagtaagatg gtttgtacgt atatggatct cctgcttgac gattttgtca 19560
gcattcttaa atctttggat ttgggcgttg tatctaaagt tcatgaagtt atggtcgatt 19620
gtaaaatgtg gaggtggatg ttgtggtgta aggatcataa actccagaca ttttatccgc 19680
aacttcaggc cagtgaatgg aagtgtggtt attccatgcc ttctatttac aagatacaac 19740
gtatgtgttt agaaccttgc aatctctaca actatggtgc tggtattaag ttacctgatg 19800
gcattatgtt taacgtagtt aaatacacac agctttgtca atatctcaat agcaccacaa 19860
tgtgtgtacc ccatcacatg cgtgtgctac atcttggtgc tggctccgac aagggtgttg 19920
cacctggcac ggctgtctta cgacgttggt tgccactgga tgccattata gttgacaatg 19980
atagtgtgga ttacgttagc gatgctgatt atagtgttac aggagattgc tctaccttat 20040
acctgtcaga taagtttgat ttagttatat ctgatatgta tgatggtaag attaaaagtt 20100
gtgatgggga gaacgtgtct aaagaaggct tctttcccta tattaatggt gtcatcaccg 20160
aaaagttggc acttggtggt actgtagcta ttaaggtgac ggagtttagt tggaataaga 20220
agttgtatga actcattcag aggtttgagt attggacaat gttctgtacc agtgttaaca 20280
cgtcatcgtc agaggcattc ttaattggtg ttcactattt aggtgatttt gcaagtggcg 20340
ctgtgattga cggcaacact atgcatgcca attatatctt ctggcgtaat tccacaatta 20400
tgactatgtc ttacaatagt gtacttgatt taagcaagtt caattgtaag cataaggcta 20460
cagttgtcat taatttaaaa gattcatcca ttagtgatgt tgtgttaggt ttgttgaaga 20520
atggtaagtt gctagtgcgt aataatgacg ccatttgtgg tttttctaat catttggtca 20580
acgtaaacaa atgaagtctt taacctactt ctggttgttc ttaccagtac tttcaacact 20640
tagcctacca caagatgtca ccaggtgctc agctaacact aattttaggc ggttcttttc 20700
aaaatttaat gttcaggcgc ctgcagttgt tgtactgggc ggttatctac ctattggtga 20760
aaaccagggt gtcaattcaa cttggtactg tgctggccaa catccaactg ctagtggcgt 20820
tcatggtatc tttgttagcc atattagagg tggtcatggc tttgagattg gcatttcgca 20880
agagcctttt gaccctagtg gttaccagct ttatttacat aaggctacta acggtaacac 20940
taatgctact gcgcgactgc gcatttgcca gtttcctagc attaataatg atgttacaat 21000
aggtcgtaat tgcctattta acaaagccat cccagctcat atgagtgaac atagtgttgt 21060
cggcataaca tgggataatg atcgtgtcac tgtcttttct gacaagatct attattttta 21120
ttttaaaaat gattggtccc gtgttgcgac aaagtgttac aacagtggag gttgtgctat 21180
gcaatatgtt tacgaaccca cctattacat gcttaatgtt actagtgctg gtgaggatgg 21240
tatttcttat caaccctgta cagctaattg cattggttat gctgccaatg tatttgctac 21300
tgagcccaat ggccacatac cagaaggttt tagttttaat aattggtttc ttttgtccaa 21360
tgattccact ttggtgcatg gtaaggtggt ttccaaccaa ccattgttgg tcaattgtct 21420
tttggccatt cctaagattt atggactagg ccaatttttc tcctttaatc aaacgatcga 21480
tggtgtttgt aatggagctg ctgtgcagcg tgcaccagag gctctgaggt ttaatattaa 21540
tgacacctct gtcattcttg ctgaaggctc aattgtactt catactgctt taggaacaaa 21600
tttttctttt gtttgcagta attcctcaaa tcctcactta gccaccttcg ccatacctct 21660
gggtgctacc caagtacctt attattgttt tcttaaagtg gatacttaca actccactgt 21720
ttataaattt ttggctgttt tacctcctac cgtcagggaa attgtcatca ccaagtatgg 21780
tgatgtttat gtcaatgggt ttggatactt gcatctcggt ttgttggatg ctgtcacaat 21840
taatttcact ggtcatggca ctgacgatga tgtttctggt ttttggacca tagcatcgac 21900
taattttgtt gatgcactca tcgaagttca aggaaccgcc attcagcgta ttctttattg 21960
tgatgatcct gttagccaac tcaagtgttc tcaggttgct tttgaccttg acgatggttt 22020
ttaccctatt tcttctagaa accttctgag tcatgaacag ccaatttctt ttgttactct 22080
gccatcattt aatgatcatt cttttgttaa cattactgta tctgcttcct ttggtggtca 22140
tagtggtgcc aaccttattg catctgacac tactatcaat gggtttagtt ctttctgtgt 22200
tgacactaga caatttacca tttcactgtt ttataacgtt acaaacagtt atggttatgt 22260
gtctaaatca caggacagta attgcccttt caccttgcaa tctgttaatg attacctgtc 22320
ttttagcaaa ttttgtgttt ccaccagcct tttggctagt gcctgtacca tagatctttt 22380
tggttaccct gagtttggta gtggtgttaa gtttacgtcc ctttattttc aattcacaaa 22440
gggtgagttg attactggca cgactaaacc acttgaaggt gtcacggacg tttcttttat 22500
gactctggat gtgtgtacca agtatactat ctatggcttt aaaggtgagg gtatcattac 22560
ccttacaaat tctagctttt tggcaggtgt ttattacaca tctgattctg gacagttgtt 22620
agcctttaag aatgtcacta gtggtgctgt ttattctgtt acgccatgtt ctttttcaga 22680
gcaggctgca tatgttgatg atgatatagt gggtgttatt tctagtttgt ctagctccac 22740
ttttaacagt actagggagt tgcctggttt cttctaccat tctaatgatg gctctaattg 22800
tacagagcct gtgttggtgt atagtaacat aggtgtttgt aaatctggca gtattggcta 22860
cgtcccatct cagtctggcc aagtcaagat tgcacccacg gttactggga atattagtat 22920
tcccaccaac tttagtatga gtattaggac agaatattta cagctttaca acacgcctgt 22980
tagtgttgat tgtgccacat atgtttgtaa tggtaactct cgttgtaaac aattactcac 23040
ccagtacact gcagcatgta agaccataga gtcagcatta caactcagcg ctaggcttga 23100
gtctgttgaa gttaactcta tgcttactat ttctgaagag gctctacagt tagctaccat 23160
tagttcgttt aatggtgatg gatataattt tactaatgtg ctgggtgttt ctgtgtatga 23220
tcctgcaagt ggcagggtgg tacaaaaaag gtcttttatt gaagacctgc tttttaataa 23280
agtggttact aatggccttg gtactgttga tgaagactat aagcgctgtt ctaatggtcg 23340
ctctgtggca gatctagtct gtgcacagta ttactctggt gtcatggtac tacctggtgt 23400
tgttgacgct gagaagcttc acatgtatag tgcgtctctc atcggtggta tggtgctagg 23460
aggttttact tctgcagcgg cattgccttt tagctatgct gttcaagcta gactcaatta 23520
tcttgctcta cagacggatg ttctacagcg gaaccagcaa ttgcttgctg agtcttttaa 23580
ctctgctatt ggtaatataa cttcagcctt tgagagtgtt aaagaggcta ttagtcaaac 23640
ttccaagggt ttgaacactg tggctcatgc gcttactaag gttcaagagg ttgttaactc 23700
gcagggtgca gctttgactc aacttaccgt acagctgcaa cacaacttcc aagccatttc 23760
tagttctatt gatgacattt actctcgact ggacattctt tcagccgatg ttcaggttga 23820
ccgtctcatc accggcagat tatcagcact taatgctttt gttgctcaaa ccctcactaa 23880
gtatactgag gttcaggcta gcaggaagtt agcacagcaa aaggttaatg agtgcgttaa 23940
atcgcaatct cagcgttatg gtttttgtgg tggtgatggc gagcacattt tctctctggt 24000
acaggcagca cctcagggcc tgctgttttt acatacagta cttgtaccga gtgattttgt 24060
agatgttatt gccatcgctg gcttatgcgt taacgatgaa attgccttga ctctacgtga 24120
gcctggctta gtcttgttta cgcatgaact tcaaaatcat actgcgacgg aatattttgt 24180
ttcatcgcga cgtatgtttg aacctagaaa acctaccgtt agtgattttg ttcaaattga 24240
gagttgtgtg gtcacctatg tcaatttgac tagagaccaa ctaccagatg taatcccaga 24300
ttacatcgat gttaacaaaa cacttgatga gattttagct tctctgccca atagaactgg 24360
tccaagtctt cctttagatg tttttaatgc cacttatctt aatctcactg gtgaaattgc 24420
agatttagag cagcgttcag agtctctccg taatactaca gaggagctcc aaagtcttat 24480
atataatatc aacaacacac tagttgacct tgagtggctc aaccgagttg agacatatat 24540
caagtggccg tggtgggttt ggttgattat tttcattgtt ctcatctttg ttgtgtcatt 24600
actagtgttc tgctgcattt ccacgggttg ttgtggatgc tgcggctgct gctgtgcttg 24660
tttctcaggt tgttgtaggg gtcctagact tcaaccttac gaagtttttg aaaaggtcca 24720
cgtgcagtga tgtttcttgg actttttcaa tacacgattg acacagttgt caaagatgtc 24780
tcaaagtctg ctaacttgtc tttggatgct gtccaagagt tggagctcaa tgtagttcca 24840
attagacaag cttcaaatgt gacgggtttt cttttcacca gtgtttttat ctacttcttt 24900
gcactgttta aagcgtcttc tttgaggcgc aattatatta tgttggcagc gcgttttgct 24960
gtcattgttc tttattgccc acttttatat tattgtggtg catttttaga tgcaactatt 25020
atttgttgca cacttattgg caggctttgt ttagtctgct tttactcctg gcgctataaa 25080
aatgcgctct ttattatttt taatactacg acactttctt tcctcaatgg taaagcagct 25140
tattatgacg gcaaatccat tgtgatttta gaaggtggtg accattacat cacttttggc 25200
aactcttttg ttgcttttgt tagtagcatc gacttgtatc tagctatacg tgggcggcaa 25260
gaagctgacc tacagctgtt gcgaactgtt gagcttcttg atggcaagaa gctttatgtc 25320
ttttcgcaac atcaaattgt tggcattact aatgctgcat ttgactcaat tcaactagac 25380
gagtatgcta caattagtga atgataatgg tctagtagtt aatgttatac tttggctttt 25440
cgtactcttt ttcctgctta ttataagcat tactttcgtc caattggtta atctgtgctt 25500
cacttgtcac cggttgtgta atagcgcagt ttacacacct atagggcgtt tgtatagagt 25560
ttataagtct tacatgcaaa tagaccccct ccctagtact gttattgacg tataaacgaa 25620
atatgtctaa cggttctatt cccgttgatg aggtgattca acaccttaga aactggaatt 25680
tcacatggaa tatcatactg acgatactac ttgtagtgct tcagtatggc cattacaagt 25740
actctgcgtt cttgtatggt gtcaagatgg ctattctatg gatactttgg cctcttgtgt 25800
tagcactgtc actttttgat gcatgggcta gctttcaggt caattgggtc ttttttgctt 25860
tcagcatcct tatggcttgc atcactctta tgctgtggat aatgtacttt gtcaatagca 25920
ttcggttgtg gcgcaggaca cattcttggt ggtctttcaa tcctgaaaca gacgcgcttc 25980
tcactacttc tgtgatgggc cgacaggtct gcattccagt gcttggagca ccaactggtg 26040
taacgctaac actccttagt ggtacattgc ttgtagaggg ctataaggtt gctactggcg 26100
tacaggtaag tcaattacct aatttcgtca cagtcgccaa ggccactaca acaattgtct 26160
acggacgtgt tggtcgttca gtcaatgctt catctggcac tggttgggct ttctatgtcc 26220
ggtccaaaca cggcgactac tcagctgtga gtaatccgag ttcggttctc acagatagtg 26280
agaaagtgct tcatttagtc taaacagaaa ctttatggct tctgtcagtt ttcaggatcg 26340
tggccgcaaa cgggtgccat tatccctcta tgcccctctt agggttacta atgacaaacc 26400
cctttctaag gtacttgcaa ataatgctgt acccactaat aaaggaaata aggaccagca 26460
aattggatac tggaatgagc aaattcgctg gcgcatgcgc cgtggtgagc gaattgaaca 26520
accttccaat tggcatttct actacctcgg aacaggacct cacgccgacc tccgctacag 26580
gactcgtact gagggtgttt tctgggttgc taaagaaggc gcaaagactg aacccactaa 26640
cctgggtgtc agaaaggcgt ctgaaaagcc aattattcca aatttctctc aacagcttcc 26700
cagcgtagtt gagattgttg aacctaacac acctcctact tcacgtgcaa attcacgtag 26760
caggagtcgt ggtaatggca acaacaggtc cagatctcca agtaacaaca gaggcaataa 26820
ccagtcccgc ggtaattcac agaatcgtgg aaataaccag ggtcgtggag cttctcagaa 26880
cagaggaggc aataataata acaataacaa gtctcgtaac cagtccaaga acagaaacca 26940
gtcaaatgac cgtggtggtg taacatcacg cgatgatctg gtggctgctg tcaaggatgc 27000
ccttaaatct ttgggtattg gcgaaaaccc tgacaagctt aagcaacagc agaagcccaa 27060
acaggaaagg tctgacagca gcggcaaaaa tacacctaag aagaacaaat ccagagccac 27120
ttcgaaagaa cgtgacctca aagacatccc agagtggagg agaattccca agggcgaaaa 27180
tagcgtagca gcttgcttcg gacccagggg aggcttcaaa aattttggag atgcggaatt 27240
tgtcgaaaaa ggtgttgatg cctcaggcta tgctcagatc gccagtttag caccaaatgt 27300
tgcagcattg ctctttggtg gtaatgtggc tgttcgtgag ctagcggact cttacgagat 27360
tacatataat tataaaatga ctgtgccaaa gtctgatcca aatgtagagc ttcttgtttc 27420
acaggtggat gcatttaaaa ctgggaatgc aaaaccccag agaaagaagg aaaagaagaa 27480
caagcgtgaa accacgcagc agctgaatga agaggccatc tacgatgatg tgggtgtgcc 27540
atctgatgtg actcatgcca atttggaatg ggacacagct gttgatggtg gtgacacggc 27600
cgttgaaatt atcaacgaga tcttcgacac aggaaattaa acaatgtttg actggcttat 27660
cctggctatg tcccagggta gtgccattac actgttatta ctgagtgttt ttctagcgac 27720
ttggctgctg ggctatggct ttgccctcta actagcggtc ttggtcttgc acacaacggt 27780
aagccagtgg taatgtcagt gcaagaagga tattaccata gcactgtcat gaggggaacg 27840
cagtaccttt tcatctaaac ctttgcacga gtaatcaaag atccgcttga cgagcctata 27900
tggaagagcg tgccaggtat ttgactcaag gactgttagt aactgaagac ctgacggtgt 27960
<210> 30
<211> 4140
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDV Isolate 1251-125-10 (125-10) Spike-protein cDNA Sequence
<400> 30
atgaagtctt taacctactt ctggttgttc ttaccagtac tttcaacact tagcctacca 60
caagatgtca ccaggtgctc agctaacact aattttaggc ggttcttttc aaaatttaat 120
gttcaggcgc ctgcagttgt tgtactgggc ggttatctac ctattggtga aaaccagggt 180
gtcaattcaa cttggtactg tgctggccaa catccaactg ctagtggcgt tcatggtatc 240
tttgttagcc atattagagg tggtcatggc tttgagattg gcatttcgca agagcctttt 300
gaccctagtg gttaccagct ttatttacat aaggctacta acggtaacac taatgctact 360
gcgcgactgc gcatttgcca gtttcctagc attaataatg atgttacaat aggtcgtaat 420
tgcctattta acaaagccat cccagctcat atgagtgaac atagtgttgt cggcataaca 480
tgggataatg atcgtgtcac tgtcttttct gacaagatct attattttta ttttaaaaat 540
gattggtccc gtgttgcgac aaagtgttac aacagtggag gttgtgctat gcaatatgtt 600
tacgaaccca cctattacat gcttaatgtt actagtgctg gtgaggatgg tatttcttat 660
caaccctgta cagctaattg cattggttat gctgccaatg tatttgctac tgagcccaat 720
ggccacatac cagaaggttt tagttttaat aattggtttc ttttgtccaa tgattccact 780
ttggtgcatg gtaaggtggt ttccaaccaa ccattgttgg tcaattgtct tttggccatt 840
cctaagattt atggactagg ccaatttttc tcctttaatc aaacgatcga tggtgtttgt 900
aatggagctg ctgtgcagcg tgcaccagag gctctgaggt ttaatattaa tgacacctct 960
gtcattcttg ctgaaggctc aattgtactt catactgctt taggaacaaa tttttctttt 1020
gtttgcagta attcctcaaa tcctcactta gccaccttcg ccatacctct gggtgctacc 1080
caagtacctt attattgttt tcttaaagtg gatacttaca actccactgt ttataaattt 1140
ttggctgttt tacctcctac cgtcagggaa attgtcatca ccaagtatgg tgatgtttat 1200
gtcaatgggt ttggatactt gcatctcggt ttgttggatg ctgtcacaat taatttcact 1260
ggtcatggca ctgacgatga tgtttctggt ttttggacca tagcatcgac taattttgtt 1320
gatgcactca tcgaagttca aggaaccgcc attcagcgta ttctttattg tgatgatcct 1380
gttagccaac tcaagtgttc tcaggttgct tttgaccttg acgatggttt ttaccctatt 1440
tcttctagaa accttctgag tcatgaacag ccaatttctt ttgttactct gccatcattt 1500
aatgatcatt cttttgttaa cattactgta tctgcttcct ttggtggtca tagtggtgcc 1560
aaccttattg catctgacac tactatcaat gggtttagtt ctttctgtgt tgacactaga 1620
caatttacca tttcactgtt ttataacgtt acaaacagtt atggttatgt gtctaaatca 1680
caggacagta attgcccttt caccttgcaa tctgttaatg attacctgtc ttttagcaaa 1740
ttttgtgttt ccaccagcct tttggctagt gcctgtacca tagatctttt tggttaccct 1800
gagtttggta gtggtgttaa gtttacgtcc ctttattttc aattcacaaa gggtgagttg 1860
attactggca cgactaaacc acttgaaggt gtcacggacg tttcttttat gactctggat 1920
gtgtgtacca agtatactat ctatggcttt aaaggtgagg gtatcattac ccttacaaat 1980
tctagctttt tggcaggtgt ttattacaca tctgattctg gacagttgtt agcctttaag 2040
aatgtcacta gtggtgctgt ttattctgtt acgccatgtt ctttttcaga gcaggctgca 2100
tatgttgatg atgatatagt gggtgttatt tctagtttgt ctagctccac ttttaacagt 2160
actagggagt tgcctggttt cttctaccat tctaatgatg gctctaattg tacagagcct 2220
gtgttggtgt atagtaacat aggtgtttgt aaatctggca gtattggcta cgtcccatct 2280
cagtctggcc aagtcaagat tgcacccacg gttactggga atattagtat tcccaccaac 2340
tttagtatga gtattaggac agaatattta cagctttaca acacgcctgt tagtgttgat 2400
tgtgccacat atgtttgtaa tggtaactct cgttgtaaac aattactcac ccagtacact 2460
gcagcatgta agaccataga gtcagcatta caactcagcg ctaggcttga gtctgttgaa 2520
gttaactcta tgcttactat ttctgaagag gctctacagt tagctaccat tagttcgttt 2580
aatggtgatg gatataattt tactaatgtg ctgggtgttt ctgtgtatga tcctgcaagt 2640
ggcagggtgg tacaaaaaag gtcttttatt gaagacctgc tttttaataa agtggttact 2700
aatggccttg gtactgttga tgaagactat aagcgctgtt ctaatggtcg ctctgtggca 2760
gatctagtct gtgcacagta ttactctggt gtcatggtac tacctggtgt tgttgacgct 2820
gagaagcttc acatgtatag tgcgtctctc atcggtggta tggtgctagg aggttttact 2880
tctgcagcgg cattgccttt tagctatgct gttcaagcta gactcaatta tcttgctcta 2940
cagacggatg ttctacagcg gaaccagcaa ttgcttgctg agtcttttaa ctctgctatt 3000
ggtaatataa cttcagcctt tgagagtgtt aaagaggcta ttagtcaaac ttccaagggt 3060
ttgaacactg tggctcatgc gcttactaag gttcaagagg ttgttaactc gcagggtgca 3120
gctttgactc aacttaccgt acagctgcaa cacaacttcc aagccatttc tagttctatt 3180
gatgacattt actctcgact ggacattctt tcagccgatg ttcaggttga ccgtctcatc 3240
accggcagat tatcagcact taatgctttt gttgctcaaa ccctcactaa gtatactgag 3300
gttcaggcta gcaggaagtt agcacagcaa aaggttaatg agtgcgttaa atcgcaatct 3360
cagcgttatg gtttttgtgg tggtgatggc gagcacattt tctctctggt acaggcagca 3420
cctcagggcc tgctgttttt acatacagta cttgtaccga gtgattttgt agatgttatt 3480
gccatcgctg gcttatgcgt taacgatgaa attgccttga ctctacgtga gcctggctta 3540
gtcttgttta cgcatgaact tcaaaatcat actgcgacgg aatattttgt ttcatcgcga 3600
cgtatgtttg aacctagaaa acctaccgtt agtgattttg ttcaaattga gagttgtgtg 3660
gtcacctatg tcaatttgac tagagaccaa ctaccagatg taatcccaga ttacatcgat 3720
gttaacaaaa cacttgatga gattttagct tctctgccca atagaactgg tccaagtctt 3780
cctttagatg tttttaatgc cacttatctt aatctcactg gtgaaattgc agatttagag 3840
cagcgttcag agtctctccg taatactaca gaggagctcc aaagtcttat atataatatc 3900
aacaacacac tagttgacct tgagtggctc aaccgagttg agacatatat caagtggccg 3960
tggtgggttt ggttgattat tttcattgtt ctcatctttg ttgtgtcatt actagtgttc 4020
tgctgcattt ccacgggttg ttgtggatgc tgcggctgct gctgtgcttg tttctcaggt 4080
tgttgtaggg gtcctagact tcaaccttac gaagtttttg aaaaggtcca cgtgcagtga 4140
<210> 31
<211> 1379
<212> PRT
<213> porcine epidemic diarrhea virus
<400> 31
Met Lys Ser Leu Thr Tyr Phe Trp Leu Phe Leu Pro Val Leu Ser Thr
1 5 10 15
Leu Ser Leu Pro Gln Asp Val Thr Arg Cys Ser Ala Asn Thr Asn Phe
20 25 30
Arg Arg Phe Phe Ser Lys Phe Asn Val Gln Ala Pro Ala Val Val Val
35 40 45
Leu Gly Gly Tyr Leu Pro Ile Gly Glu Asn Gln Gly Val Asn Ser Thr
50 55 60
Trp Tyr Cys Ala Gly Gln His Pro Thr Ala Ser Gly Val His Gly Ile
65 70 75 80
Phe Val Ser His Ile Arg Gly Gly His Gly Phe Glu Ile Gly Ile Ser
85 90 95
Gln Glu Pro Phe Asp Pro Ser Gly Tyr Gln Leu Tyr Leu His Lys Ala
100 105 110
Thr Asn Gly Asn Thr Asn Ala Thr Ala Arg Leu Arg Ile Cys Gln Phe
115 120 125
Pro Ser Ile Asn Asn Asp Val Thr Ile Gly Arg Asn Cys Leu Phe Asn
130 135 140
Lys Ala Ile Pro Ala His Met Ser Glu His Ser Val Val Gly Ile Thr
145 150 155 160
Trp Asp Asn Asp Arg Val Thr Val Phe Ser Asp Lys Ile Tyr Tyr Phe
165 170 175
Tyr Phe Lys Asn Asp Trp Ser Arg Val Ala Thr Lys Cys Tyr Asn Ser
180 185 190
Gly Gly Cys Ala Met Gln Tyr Val Tyr Glu Pro Thr Tyr Tyr Met Leu
195 200 205
Asn Val Thr Ser Ala Gly Glu Asp Gly Ile Ser Tyr Gln Pro Cys Thr
210 215 220
Ala Asn Cys Ile Gly Tyr Ala Ala Asn Val Phe Ala Thr Glu Pro Asn
225 230 235 240
Gly His Ile Pro Glu Gly Phe Ser Phe Asn Asn Trp Phe Leu Leu Ser
245 250 255
Asn Asp Ser Thr Leu Val His Gly Lys Val Val Ser Asn Gln Pro Leu
260 265 270
Leu Val Asn Cys Leu Leu Ala Ile Pro Lys Ile Tyr Gly Leu Gly Gln
275 280 285
Phe Phe Ser Phe Asn Gln Thr Ile Asp Gly Val Cys Asn Gly Ala Ala
290 295 300
Val Gln Arg Ala Pro Glu Ala Leu Arg Phe Asn Ile Asn Asp Thr Ser
305 310 315 320
Val Ile Leu Ala Glu Gly Ser Ile Val Leu His Thr Ala Leu Gly Thr
325 330 335
Asn Phe Ser Phe Val Cys Ser Asn Ser Ser Asn Pro His Leu Ala Thr
340 345 350
Phe Ala Ile Pro Leu Gly Ala Thr Gln Val Pro Tyr Tyr Cys Phe Leu
355 360 365
Lys Val Asp Thr Tyr Asn Ser Thr Val Tyr Lys Phe Leu Ala Val Leu
370 375 380
Pro Pro Thr Val Arg Glu Ile Val Ile Thr Lys Tyr Gly Asp Val Tyr
385 390 395 400
Val Asn Gly Phe Gly Tyr Leu His Leu Gly Leu Leu Asp Ala Val Thr
405 410 415
Ile Asn Phe Thr Gly His Gly Thr Asp Asp Asp Val Ser Gly Phe Trp
420 425 430
Thr Ile Ala Ser Thr Asn Phe Val Asp Ala Leu Ile Glu Val Gln Gly
435 440 445
Thr Ala Ile Gln Arg Ile Leu Tyr Cys Asp Asp Pro Val Ser Gln Leu
450 455 460
Lys Cys Ser Gln Val Ala Phe Asp Leu Asp Asp Gly Phe Tyr Pro Ile
465 470 475 480
Ser Ser Arg Asn Leu Leu Ser His Glu Gln Pro Ile Ser Phe Val Thr
485 490 495
Leu Pro Ser Phe Asn Asp His Ser Phe Val Asn Ile Thr Val Ser Ala
500 505 510
Ser Phe Gly Gly His Ser Gly Ala Asn Leu Ile Ala Ser Asp Thr Thr
515 520 525
Ile Asn Gly Phe Ser Ser Phe Cys Val Asp Thr Arg Gln Phe Thr Ile
530 535 540
Ser Leu Phe Tyr Asn Val Thr Asn Ser Tyr Gly Tyr Val Ser Lys Ser
545 550 555 560
Gln Asp Ser Asn Cys Pro Phe Thr Leu Gln Ser Val Asn Asp Tyr Leu
565 570 575
Ser Phe Ser Lys Phe Cys Val Ser Thr Ser Leu Leu Ala Ser Ala Cys
580 585 590
Thr Ile Asp Leu Phe Gly Tyr Pro Glu Phe Gly Ser Gly Val Lys Phe
595 600 605
Thr Ser Leu Tyr Phe Gln Phe Thr Lys Gly Glu Leu Ile Thr Gly Thr
610 615 620
Thr Lys Pro Leu Glu Gly Val Thr Asp Val Ser Phe Met Thr Leu Asp
625 630 635 640
Val Cys Thr Lys Tyr Thr Ile Tyr Gly Phe Lys Gly Glu Gly Ile Ile
645 650 655
Thr Leu Thr Asn Ser Ser Phe Leu Ala Gly Val Tyr Tyr Thr Ser Asp
660 665 670
Ser Gly Gln Leu Leu Ala Phe Lys Asn Val Thr Ser Gly Ala Val Tyr
675 680 685
Ser Val Thr Pro Cys Ser Phe Ser Glu Gln Ala Ala Tyr Val Asp Asp
690 695 700
Asp Ile Val Gly Val Ile Ser Ser Leu Ser Ser Ser Thr Phe Asn Ser
705 710 715 720
Thr Arg Glu Leu Pro Gly Phe Phe Tyr His Ser Asn Asp Gly Ser Asn
725 730 735
Cys Thr Glu Pro Val Leu Val Tyr Ser Asn Ile Gly Val Cys Lys Ser
740 745 750
Gly Ser Ile Gly Tyr Val Pro Ser Gln Ser Gly Gln Val Lys Ile Ala
755 760 765
Pro Thr Val Thr Gly Asn Ile Ser Ile Pro Thr Asn Phe Ser Met Ser
770 775 780
Ile Arg Thr Glu Tyr Leu Gln Leu Tyr Asn Thr Pro Val Ser Val Asp
785 790 795 800
Cys Ala Thr Tyr Val Cys Asn Gly Asn Ser Arg Cys Lys Gln Leu Leu
805 810 815
Thr Gln Tyr Thr Ala Ala Cys Lys Thr Ile Glu Ser Ala Leu Gln Leu
820 825 830
Ser Ala Arg Leu Glu Ser Val Glu Val Asn Ser Met Leu Thr Ile Ser
835 840 845
Glu Glu Ala Leu Gln Leu Ala Thr Ile Ser Ser Phe Asn Gly Asp Gly
850 855 860
Tyr Asn Phe Thr Asn Val Leu Gly Val Ser Val Tyr Asp Pro Ala Ser
865 870 875 880
Gly Arg Val Val Gln Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn
885 890 895
Lys Val Val Thr Asn Gly Leu Gly Thr Val Asp Glu Asp Tyr Lys Arg
900 905 910
Cys Ser Asn Gly Arg Ser Val Ala Asp Leu Val Cys Ala Gln Tyr Tyr
915 920 925
Ser Gly Val Met Val Leu Pro Gly Val Val Asp Ala Glu Lys Leu His
930 935 940
Met Tyr Ser Ala Ser Leu Ile Gly Gly Met Val Leu Gly Gly Phe Thr
945 950 955 960
Ser Ala Ala Ala Leu Pro Phe Ser Tyr Ala Val Gln Ala Arg Leu Asn
965 970 975
Tyr Leu Ala Leu Gln Thr Asp Val Leu Gln Arg Asn Gln Gln Leu Leu
980 985 990
Ala Glu Ser Phe Asn Ser Ala Ile Gly Asn Ile Thr Ser Ala Phe Glu
995 1000 1005
Ser Val Lys Glu Ala Ile Ser Gln Thr Ser Lys Gly Leu Asn Thr
1010 1015 1020
Val Ala His Ala Leu Thr Lys Val Gln Glu Val Val Asn Ser Gln
1025 1030 1035
Gly Ala Ala Leu Thr Gln Leu Thr Val Gln Leu Gln His Asn Phe
1040 1045 1050
Gln Ala Ile Ser Ser Ser Ile Asp Asp Ile Tyr Ser Arg Leu Asp
1055 1060 1065
Ile Leu Ser Ala Asp Val Gln Val Asp Arg Leu Ile Thr Gly Arg
1070 1075 1080
Leu Ser Ala Leu Asn Ala Phe Val Ala Gln Thr Leu Thr Lys Tyr
1085 1090 1095
Thr Glu Val Gln Ala Ser Arg Lys Leu Ala Gln Gln Lys Val Asn
1100 1105 1110
Glu Cys Val Lys Ser Gln Ser Gln Arg Tyr Gly Phe Cys Gly Gly
1115 1120 1125
Asp Gly Glu His Ile Phe Ser Leu Val Gln Ala Ala Pro Gln Gly
1130 1135 1140
Leu Leu Phe Leu His Thr Val Leu Val Pro Ser Asp Phe Val Asp
1145 1150 1155
Val Ile Ala Ile Ala Gly Leu Cys Val Asn Asp Glu Ile Ala Leu
1160 1165 1170
Thr Leu Arg Glu Pro Gly Leu Val Leu Phe Thr His Glu Leu Gln
1175 1180 1185
Asn His Thr Ala Thr Glu Tyr Phe Val Ser Ser Arg Arg Met Phe
1190 1195 1200
Glu Pro Arg Lys Pro Thr Val Ser Asp Phe Val Gln Ile Glu Ser
1205 1210 1215
Cys Val Val Thr Tyr Val Asn Leu Thr Arg Asp Gln Leu Pro Asp
1220 1225 1230
Val Ile Pro Asp Tyr Ile Asp Val Asn Lys Thr Leu Asp Glu Ile
1235 1240 1245
Leu Ala Ser Leu Pro Asn Arg Thr Gly Pro Ser Leu Pro Leu Asp
1250 1255 1260
Val Phe Asn Ala Thr Tyr Leu Asn Leu Thr Gly Glu Ile Ala Asp
1265 1270 1275
Leu Glu Gln Arg Ser Glu Ser Leu Arg Asn Thr Thr Glu Glu Leu
1280 1285 1290
Gln Ser Leu Ile Tyr Asn Ile Asn Asn Thr Leu Val Asp Leu Glu
1295 1300 1305
Trp Leu Asn Arg Val Glu Thr Tyr Ile Lys Trp Pro Trp Trp Val
1310 1315 1320
Trp Leu Ile Ile Phe Ile Val Leu Ile Phe Val Val Ser Leu Leu
1325 1330 1335
Val Phe Cys Cys Ile Ser Thr Gly Cys Cys Gly Cys Cys Gly Cys
1340 1345 1350
Cys Cys Ala Cys Phe Ser Gly Cys Cys Arg Gly Pro Arg Leu Gln
1355 1360 1365
Pro Tyr Glu Val Phe Glu Lys Val His Val Gln
1370 1375
<210> 32
<211> 27960
<212> RNA
<213> porcine epidemic diarrhea virus
<400> 32
agacucuugu cuacucaauu caacuaaacg aaauuuuguc cuuccggccg cauguccaug 60
cugcuggaag cugacgugga auuucauuag guuugcuuaa guagccaucg caagugcugu 120
gcuguccucu aguuccuggu uggcguuccg ucgccuucua cauacuagac aaacagccuu 180
ccuccgguuc cgucuggggg uuguguggau aacuaguucc gucuaguuug aaaccaguaa 240
cugucggcua uggcuagcaa ccauguuaca uuggcuuuug ccaaugaugc agaaauuuca 300
gcuuuuggcu uuugcacugc uagugaagcc gucucauacu auucugaggc cgccgcuagu 360
ggauuuaugc aaugccguuu cguguccuuc gaucucgcug acacuguuga gggauugcuu 420
cccgaagacu augucauggu gguggucggc acuaccaagc uuagugcgua uguggacacu 480
uuugguagcc gccccaaaaa cauuuguggu uggcuguuau uuucuaacug uaauuacuuc 540
cucgaagagu uagagcuuac uuuuggucgu cgugguggua acaucgugcc aguugaccaa 600
uacaugugug gcgcugacgg uaaaccuguu cuucaggaau ccgaauggga guauacagau 660
uucuuugcug acuccgaaga cggucaacuc aacauugcug guaucacuua ugugaaggcc 720
uggauuguag agcgaucgga ugucucuuau gcgagucaga auuuaacauc uauuaagucu 780
auuacuuacu guucaaccua ugagcauacu uuuccugaug guacugccau gaagguugca 840
cguacuccaa agauuaagaa gacuguuguc uugucugagc cacuugcuac uaucuacagg 900
gaaauugguu cuccuuuugu ggauaauggg agcgaugcuc guucuaucau uaagagacca 960
guguuccucc acgcuuuugu uaaguguaag ugugguaguu aucauuggac uguuggugau 1020
uggacuuccu augucuccac uugcuguggc uuuaagugua agccaguccu uguggcuuca 1080
ugcucugcua cgccugguuc uguugugguu acgcgcgcug gugcuggcac ugguguuaag 1140
uauuacaaca acauguuccu gcgccaugug gcagacauug auggguuggc auucuggcga 1200
auucucaagg ugcaguccaa agacgaccuc gcuugcucug guaaauuccu ugaacaccau 1260
gaggaagguu ucacagaucc uugcuacuuu uugaaugacu cgagcauugc uacuaagcuc 1320
aaguuugaca uccuuagugg caaguuuucu gaugaaguca aacaagcuau cuuugcuggu 1380
cauguuguug uuggcagcgc gcucguugac auuguugacg augcacuggg acagccuugg 1440
uuuauacgua agcuugguga ccuugcaagu gcagcuuggg agcagcuuaa ggcugucguu 1500
agaggccuua accuccuguc ugaugagguc gugcucuuug gcaaaagacu uagcugugcc 1560
acucuuagua ucguuaacgg uguuuuugag uucaucgccg aagugccuga gaaguuggcu 1620
gcggcuguua caguuuuugu caacuucuug aaugagcuuu uugagucugc cugugacugc 1680
uuaaaggucg gagguaaaac cuuuaacaag guuggcucuu auguucuuuu ugacaacgca 1740
uugguuaagc uugucaaggc aaaaguucgc ggcccacgac aggcaggugu uugugaaguu 1800
cguuacacaa gccuuguuau ugggaguacu accaaggugg uuuccaagcg cguugaaaau 1860
gccaauguga aucucgucgu cguugacgag gaugugaccc ucaacaccac uggucguaca 1920
guuguuguug acggacuugc auucuucgag agugacgggu uuuacagaca ucuugcugau 1980
gcugacguug ucauugaaca uccuguuuau aagucugcuu gugagcucaa gccaguuuuu 2040
gagugugacc caauaccuga uuuuccuaug ccuguggccg cuaguguugc agagcuuugu 2100
gugcaaacug aucuguugcu uaaaaauuac aacacuccuu auaaaacuua cagcugcguu 2160
gugagaggug auaaguguug uaucacuugc accuuacauu ucacagcacc aaguuauaug 2220
gaggcugcug cuaauuuugu agaccucugu accaagaaca uugguacugc ugguuuucau 2280
gaguuuuaca uuacggccca ugaacaacag gaucugcaag gguucguaac cacuuguugc 2340
acgaugucag guuuugagug uuuuaugccu auaaucccac aguguccagc agugcuugaa 2400
gagauugaug gugguagcau cuggcggucu uuuaucacug gucuuaauac aaugugggau 2460
uuuugcaagc aucuuaaagu cagcuuugga cuagauggca uuguugucac uguagcacgc 2520
aaauuuaaac gacuuggugc ucucuuggca gaaauguaua acacuuaccu uucaacugug 2580
guggaaaacu ugguacuggc cgguguuagc uucaaguauu augccaccag ugucccaaaa 2640
auuguuuugg gcuguuguuu ucacaguguu aaaaguguuc uugcaagugc cuuccagauu 2700
ccuguccagg caggcguuga gaaguuuaaa gucuuccuua acuguguuca cccuguugua 2760
ccacguguca uugaaacuuc uuuuguggaa uuagaagaga cgacauuuaa accaccagca 2820
cucaauggua guauugcuau uguugauggc uuugcuuucu auuaugaugg aacacuauac 2880
uaucccaccg augguaauag cguuguuccu aucugcuuua agaagaaagg ugguggugau 2940
gucaaauucu cugaugaagu cucuguuaaa accauugacc caguuuauaa ggucucccuu 3000
gaauuugagu ucgagucuga gacuauuaug gcugugcuua auaaggcugu ugguaauugu 3060
aucaagguua caggugguug ggacgauguu guugaguaua ucaauguugc cauugagguu 3120
cuuaaagauc acaucgaugu gccuaaguac uacaucuaug augaggaagg uggcaccgau 3180
ccuaaucugc ccguaauggu uucucagugg ccguugaaug augacacgau cucacaggau 3240
cugcuugaug uugaaguugu uacugaugcg ccaguugauu ucgaggguga ugaaguagac 3300
uccucugacc cugauaaggu ggcagacgug gcuaacucug agccugagga ugacggucuu 3360
aauguagcuc cugaaacaaa uguagagucu gaaguugagg aaguugccgc aaccuugucc 3420
uuuauuaaag auacaccuuc cacaguuacu aaggauccuu uugcuuuuga cuuugcaagc 3480
uauggaggac uuaagguuuu aagacaaucu cauaacaacu gcuggguuac uucuaccuug 3540
gugcagcuac aauugcuugg caucguugau gacccugcaa uggagcuuuu uagugcuggu 3600
agaguugguc caaugguucg caaaugcuau gagucacaaa aggcuaucuu gggaucuuug 3660
ggugaugugu cggcuugccu agagucucug acuaaggacc uacacacacu uaagauuacc 3720
uguucuguag ucugugguug ugguacuggu gaacguaucu augaugguug ugcuuuucgu 3780
augacgccaa cuuuggaacc guucccauau ggugcuugug cucagugugc ucaaguuuug 3840
augcacacuu uuaaaaguau uguuggcacc ggcaucuuuu gucgagauac uacugcucuc 3900
uccuuggauu cuuugguugu aaaaccucuu ugugcggcug cuuuuauagg caaggauagu 3960
ggucauuaug ucacuaacuu uuaugaugcu gcuauggcua uugaugguua uggucgucau 4020
cagauaaagu augacacacu gaacacuauu uguguuaaag acguuaauug gacagcaccu 4080
uuugucccag acguugagcc uguauuggag ccuguuguca aaccuuucua uucuuauaag 4140
aauguugauu uuuaccaagg agauuuuagu gaccuuguua aacuuccaug ugauuuuguu 4200
guuaaugcug caaaugagaa uuugucucac gguggcggca uagcaaaggc cauugauguu 4260
uauaccaagg gcauguugca gaagugcucg aaugauuaca uuaaagcaca cggucccauu 4320
aaaguuggac guggugucau guuggaggca uuaggucuua aggucuuuaa uguuguuggu 4380
ccacguaagg guaagcaugc accugagcuu cuuguuaagg cuuauaaguc cguuuuugcu 4440
aauucaggug uugcucuuac accuuugauu aguguuggaa uuuuuagugu uccuuuggaa 4500
gaaucuuuau cugcuuuucu ugcauguguu ggugaucgcc acuguaagug cuuuuguuau 4560
agugacaaag agcgcgaggc gaucauuaau uacauggaug gcuugguaga ugcuauuuuc 4620
aaagaugcac uuguugauac uacuccuguc caggaagaug uucaacaagu uucacaaaaa 4680
ccaguuuugc cuaauuuuga accuuucagg auugaaggug cucaugcuuu cuaugagugc 4740
aacccugaag guuugauguc auuaggugcu gacaagcugg uguuguuuac aaauuccaau 4800
uuggauuuuu guagcguugg uaagugucuu aacaauguga cuggcggugc auugcuugaa 4860
gccauaaaug uauuuaaaaa gaguaacaaa acagugccug cuggcaacug uguuacuuuu 4920
gagugugcag auaugauuuc uauuacuaug guaguauugc caucugacgg ugaugcuaau 4980
uaugacaaaa auuaugcacg cgccgucguc aagguaucua agcuuaaagg caaguuauug 5040
cuugcuguug gugaugccau guuguauucc aaguuguccc accucagcgu guuagguuuc 5100
guauccacac cugaugaugu ggagcguuuc uacgcaaaua agaguguggu uauuaaaguu 5160
acugaggaua cacguagugu uaagacuguu aaaguagaau ccacuguuac uuauggacaa 5220
caaauuggac cuugucuugu uaaugacacc guugucacag acaacaaacc uguuguugcu 5280
gauguuguag cuaagguugu accaagugcu aauugggauu cacauuaugg uuuugauaag 5340
gcuggugagu uccacaugcu agaccauacu ggguuugccu uuccuaguga aguuguuaac 5400
gguaggcgug ugcuuaaaac cacagauaau aacuguuggg uuaauguuac auguuuacaa 5460
uuacaguuug cuagauuuag guucaaguca gcaggucuac aggcuaugug ggaguccuau 5520
uguacuggug auguugcuau guuugugcau ugguuguacu ggcuuacugg uguugacaaa 5580
ggucagccua gugauucaga aaaugcacuu aacauguugu cuaaguacau uguuccugcu 5640
gguucuguca cuauugaacg ugucacgcau gacgguuguu guuguaguaa gcguguuguc 5700
acugcaccag uugugaaugc uagcguguug aagcuuggcg ucgaggaugg ucuuugucca 5760
cauggucuua acuacauuga caaaguuguu guaguuaaag guacuacaau uguugucaau 5820
guuggaaaac cuguaguggc accaucgcac cucuuucuua aggguguuuc cuacacaaca 5880
uuccuagaua augguaacgg uguugccggc cauuauacug uuuuugauca ugacacuggu 5940
auggugcaug auggagaugu uuuuguacca ggugaucuca augugucucc uguuacaaau 6000
guugucgucu cagagcagac ggcuguugug auuaaagacc cugugaagaa aguagaguua 6060
gacgcuacaa agcuguuaga cacuaugaau uaugcaucgg aaagauucuu uuccuuuggu 6120
gauuuuaugu cacguaauuu aauuacagug uuuuuguaca uccuuaguau uuugggucuc 6180
uguuuuaggg ccuuucguaa gagggauguu aaaguucuag cugguguacc ccaacguacu 6240
gguauuauau ugcguaaaag ugugcgcuau aaugcaaagg cuuugggugu cuucuucaag 6300
cuaaaacuuu auugguucaa aguucuuggu aaguuuaguu uggguauuua ugcauuguau 6360
gcauuacuau ucaugacaau acgcuuuaca ccuauaggug gcccuguuug ugaugauguu 6420
guugcugguu augcuaauuc uaguuuugac aagaaugagu auugcaacag uguuauuugu 6480
aaggucuguc ucuaugggua ccaggaacuu ucggacuucu cucacacaca gguaguaugg 6540
caacaccuua gagacccauu aauugguaau gugaugccuu ucuuuuauuu ggcauuucug 6600
gcaauuuuug gggguguuua uguaaaggcu auuacucucu auuuuauuuu ccaguaucuu 6660
aacauacuug guguguuuuu gggccuacaa caguccauuu gguuuuugca gcuugugccu 6720
uuugaugucu uuggugacga gaucgucguc uuuuucaucg uuacacgcgu auugauguuc 6780
cuuaagcaug uuuuccuugg cugcgauaag gcaucuugug uggcuugcuc uaagagugcu 6840
cgccuuaagc gcguuccugu ccagacuauu uuucagggua cuagcaaauc cuucuacgua 6900
caugccaaug gugguucuaa guucuguaag aagcacaauu ucuuuuguuu aaauugugau 6960
ucuuaugguc caggcugcac uuuuauuaau gacgucauug caacugaagu ugguaauguu 7020
gucaaacuua augugcaacc gacagguccu gccacuauuc uuauugacaa gguugaauuc 7080
aguaaugguu uuuacuaucu uuauaguggu gacacauuuu ggaaguacaa cuuugacaua 7140
acagauaaca aauacacuug caaagaguca cuuaaaaauu guagcauaau cacagacuuu 7200
auuguuuuua acaauaaugg uuccaaugua aaucagguua agaaugcaug uguguauuuu 7260
ucacagaugc uuuguaaacc uguuaaguua guggacucag cguuguuggc caguuugucu 7320
guugauuuug gugcaagcuu acauagugcu uuuguuagug uguugucgaa uaguuuuggc 7380
aaagaccugu caaguuguaa ugacaugcag gauugcaaga gcacauuggg uuuugaugau 7440
guaccauugg auaccuuuaa ugcugcuguu gcugaggcuc aucguuacga uguccucuug 7500
acugacaugu cguucaacaa uuuuaccacc aguuaugcaa aaccagagga aaaacuuccc 7560
guccaugaca uugccacgug uaugcgugua ggugccaaga uuguuaauca uaacguucuu 7620
gucaaggaua guauaccugu gguguggcuu guacgugauu ucauugcccu uucugaagaa 7680
acuaggaagu acauuauucg uacgacuaaa guuaagggua uaaccuucau guugaccuuu 7740
aaugauuguc guaugcauac uaccauaccu acuguuugca uugcaaauaa gaagggugca 7800
ggucuuccua guuuuucaaa gguuaagaaa uucuucuggu uuuugugucu guucauaguu 7860
gcuguuuucu uugcacuaag cuuuuuugau uuuaguacuc agguuagcag ugauagugau 7920
uaugacuuca aguauauuga gaguggccag uugaagacuu uugacaaucc acuuaguugu 7980
gugcauaaug ucuuuaguaa cuucgaccag uggcaugaug ccaaguuugg uuucaccccc 8040
gucaacaauc cuaguugucc uauagucguu gguguaucag acgaagcgcg cacuguucca 8100
gguaucccag cagguguuua uuuagcuggu aaaacacuug uuuuugcuau uaacaccauu 8160
uuugguacau cugguuugug cuuugaugcu aguggcguug cugauaaggg cgcuugcauu 8220
uuuaauucgg cuugcaccac auuaucuggu uuggguggaa cugcugucua cuguuauaag 8280
aauggucuag uugaaggugc uaaacuuuau agugaguugg caccucauag cuacuauaaa 8340
augguagaug guaaugcugu gucuuuaccu gaaauuaucu cacgcggcuu uggcauccgu 8400
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ugaugauccu guuagccaac ucaaguguuc ucagguugcu uuugaccuug acgaugguuu 22020
uuacccuauu ucuucuagaa accuucugag ucaugaacag ccaauuucuu uuguuacucu 22080
gccaucauuu aaugaucauu cuuuuguuaa cauuacugua ucugcuuccu uuggugguca 22140
uaguggugcc aaccuuauug caucugacac uacuaucaau ggguuuaguu cuuucugugu 22200
ugacacuaga caauuuacca uuucacuguu uuauaacguu acaaacaguu augguuaugu 22260
gucuaaauca caggacagua auugcccuuu caccuugcaa ucuguuaaug auuaccuguc 22320
uuuuagcaaa uuuuguguuu ccaccagccu uuuggcuagu gccuguacca uagaucuuuu 22380
ugguuacccu gaguuuggua gugguguuaa guuuacgucc cuuuauuuuc aauucacaaa 22440
gggugaguug auuacuggca cgacuaaacc acuugaaggu gucacggacg uuucuuuuau 22500
gacucuggau guguguacca aguauacuau cuauggcuuu aaaggugagg guaucauuac 22560
ccuuacaaau ucuagcuuuu uggcaggugu uuauuacaca ucugauucug gacaguuguu 22620
agccuuuaag aaugucacua guggugcugu uuauucuguu acgccauguu cuuuuucaga 22680
gcaggcugca uauguugaug augauauagu ggguguuauu ucuaguuugu cuagcuccac 22740
uuuuaacagu acuagggagu ugccugguuu cuucuaccau ucuaaugaug gcucuaauug 22800
uacagagccu guguuggugu auaguaacau agguguuugu aaaucuggca guauuggcua 22860
cgucccaucu cagucuggcc aagucaagau ugcacccacg guuacuggga auauuaguau 22920
ucccaccaac uuuaguauga guauuaggac agaauauuua cagcuuuaca acacgccugu 22980
uaguguugau ugugccacau auguuuguaa ugguaacucu cguuguaaac aauuacucac 23040
ccaguacacu gcagcaugua agaccauaga gucagcauua caacucagcg cuaggcuuga 23100
gucuguugaa guuaacucua ugcuuacuau uucugaagag gcucuacagu uagcuaccau 23160
uaguucguuu aauggugaug gauauaauuu uacuaaugug cuggguguuu cuguguauga 23220
uccugcaagu ggcagggugg uacaaaaaag gucuuuuauu gaagaccugc uuuuuaauaa 23280
agugguuacu aauggccuug guacuguuga ugaagacuau aagcgcuguu cuaauggucg 23340
cucuguggca gaucuagucu gugcacagua uuacucuggu gucaugguac uaccuggugu 23400
uguugacgcu gagaagcuuc acauguauag ugcgucucuc aucgguggua uggugcuagg 23460
agguuuuacu ucugcagcgg cauugccuuu uagcuaugcu guucaagcua gacucaauua 23520
ucuugcucua cagacggaug uucuacagcg gaaccagcaa uugcuugcug agucuuuuaa 23580
cucugcuauu gguaauauaa cuucagccuu ugagaguguu aaagaggcua uuagucaaac 23640
uuccaagggu uugaacacug uggcucaugc gcuuacuaag guucaagagg uuguuaacuc 23700
gcagggugca gcuuugacuc aacuuaccgu acagcugcaa cacaacuucc aagccauuuc 23760
uaguucuauu gaugacauuu acucucgacu ggacauucuu ucagccgaug uucagguuga 23820
ccgucucauc accggcagau uaucagcacu uaaugcuuuu guugcucaaa cccucacuaa 23880
guauacugag guucaggcua gcaggaaguu agcacagcaa aagguuaaug agugcguuaa 23940
aucgcaaucu cagcguuaug guuuuugugg uggugauggc gagcacauuu ucucucuggu 24000
acaggcagca ccucagggcc ugcuguuuuu acauacagua cuuguaccga gugauuuugu 24060
agauguuauu gccaucgcug gcuuaugcgu uaacgaugaa auugccuuga cucuacguga 24120
gccuggcuua gucuuguuua cgcaugaacu ucaaaaucau acugcgacgg aauauuuugu 24180
uucaucgcga cguauguuug aaccuagaaa accuaccguu agugauuuug uucaaauuga 24240
gaguugugug gucaccuaug ucaauuugac uagagaccaa cuaccagaug uaaucccaga 24300
uuacaucgau guuaacaaaa cacuugauga gauuuuagcu ucucugccca auagaacugg 24360
uccaagucuu ccuuuagaug uuuuuaaugc cacuuaucuu aaucucacug gugaaauugc 24420
agauuuagag cagcguucag agucucuccg uaauacuaca gaggagcucc aaagucuuau 24480
auauaauauc aacaacacac uaguugaccu ugaguggcuc aaccgaguug agacauauau 24540
caaguggccg ugguggguuu gguugauuau uuucauuguu cucaucuuug uugugucauu 24600
acuaguguuc ugcugcauuu ccacggguug uuguggaugc ugcggcugcu gcugugcuug 24660
uuucucaggu uguuguaggg guccuagacu ucaaccuuac gaaguuuuug aaaaggucca 24720
cgugcaguga uguuucuugg acuuuuucaa uacacgauug acacaguugu caaagauguc 24780
ucaaagucug cuaacuuguc uuuggaugcu guccaagagu uggagcucaa uguaguucca 24840
auuagacaag cuucaaaugu gacggguuuu cuuuucacca guguuuuuau cuacuucuuu 24900
gcacuguuua aagcgucuuc uuugaggcgc aauuauauua uguuggcagc gcguuuugcu 24960
gucauuguuc uuuauugccc acuuuuauau uauuguggug cauuuuuaga ugcaacuauu 25020
auuuguugca cacuuauugg caggcuuugu uuagucugcu uuuacuccug gcgcuauaaa 25080
aaugcgcucu uuauuauuuu uaauacuacg acacuuucuu uccucaaugg uaaagcagcu 25140
uauuaugacg gcaaauccau ugugauuuua gaagguggug accauuacau cacuuuuggc 25200
aacucuuuug uugcuuuugu uaguagcauc gacuuguauc uagcuauacg ugggcggcaa 25260
gaagcugacc uacagcuguu gcgaacuguu gagcuucuug auggcaagaa gcuuuauguc 25320
uuuucgcaac aucaaauugu uggcauuacu aaugcugcau uugacucaau ucaacuagac 25380
gaguaugcua caauuaguga augauaaugg ucuaguaguu aauguuauac uuuggcuuuu 25440
cguacucuuu uuccugcuua uuauaagcau uacuuucguc caauugguua aucugugcuu 25500
cacuugucac cgguugugua auagcgcagu uuacacaccu auagggcguu uguauagagu 25560
uuauaagucu uacaugcaaa uagacccccu cccuaguacu guuauugacg uauaaacgaa 25620
auaugucuaa cgguucuauu cccguugaug aggugauuca acaccuuaga aacuggaauu 25680
ucacauggaa uaucauacug acgauacuac uuguagugcu ucaguauggc cauuacaagu 25740
acucugcguu cuuguauggu gucaagaugg cuauucuaug gauacuuugg ccucuugugu 25800
uagcacuguc acuuuuugau gcaugggcua gcuuucaggu caauuggguc uuuuuugcuu 25860
ucagcauccu uauggcuugc aucacucuua ugcuguggau aauguacuuu gucaauagca 25920
uucgguugug gcgcaggaca cauucuuggu ggucuuucaa uccugaaaca gacgcgcuuc 25980
ucacuacuuc ugugaugggc cgacaggucu gcauuccagu gcuuggagca ccaacuggug 26040
uaacgcuaac acuccuuagu gguacauugc uuguagaggg cuauaagguu gcuacuggcg 26100
uacagguaag ucaauuaccu aauuucguca cagucgccaa ggccacuaca acaauugucu 26160
acggacgugu uggucguuca gucaaugcuu caucuggcac ugguugggcu uucuaugucc 26220
gguccaaaca cggcgacuac ucagcuguga guaauccgag uucgguucuc acagauagug 26280
agaaagugcu ucauuuaguc uaaacagaaa cuuuauggcu ucugucaguu uucaggaucg 26340
uggccgcaaa cgggugccau uaucccucua ugccccucuu aggguuacua augacaaacc 26400
ccuuucuaag guacuugcaa auaaugcugu acccacuaau aaaggaaaua aggaccagca 26460
aauuggauac uggaaugagc aaauucgcug gcgcaugcgc cguggugagc gaauugaaca 26520
accuuccaau uggcauuucu acuaccucgg aacaggaccu cacgccgacc uccgcuacag 26580
gacucguacu gaggguguuu ucuggguugc uaaagaaggc gcaaagacug aacccacuaa 26640
ccuggguguc agaaaggcgu cugaaaagcc aauuauucca aauuucucuc aacagcuucc 26700
cagcguaguu gagauuguug aaccuaacac accuccuacu ucacgugcaa auucacguag 26760
caggagucgu gguaauggca acaacagguc cagaucucca aguaacaaca gaggcaauaa 26820
ccagucccgc gguaauucac agaaucgugg aaauaaccag ggucguggag cuucucagaa 26880
cagaggaggc aauaauaaua acaauaacaa gucucguaac caguccaaga acagaaacca 26940
gucaaaugac cgugguggug uaacaucacg cgaugaucug guggcugcug ucaaggaugc 27000
ccuuaaaucu uuggguauug gcgaaaaccc ugacaagcuu aagcaacagc agaagcccaa 27060
acaggaaagg ucugacagca gcggcaaaaa uacaccuaag aagaacaaau ccagagccac 27120
uucgaaagaa cgugaccuca aagacauccc agaguggagg agaauuccca agggcgaaaa 27180
uagcguagca gcuugcuucg gacccagggg aggcuucaaa aauuuuggag augcggaauu 27240
ugucgaaaaa gguguugaug ccucaggcua ugcucagauc gccaguuuag caccaaaugu 27300
ugcagcauug cucuuuggug guaauguggc uguucgugag cuagcggacu cuuacgagau 27360
uacauauaau uauaaaauga cugugccaaa gucugaucca aauguagagc uucuuguuuc 27420
acagguggau gcauuuaaaa cugggaaugc aaaaccccag agaaagaagg aaaagaagaa 27480
caagcgugaa accacgcagc agcugaauga agaggccauc uacgaugaug ugggugugcc 27540
aucugaugug acucaugcca auuuggaaug ggacacagcu guugauggug gugacacggc 27600
cguugaaauu aucaacgaga ucuucgacac aggaaauuaa acaauguuug acuggcuuau 27660
ccuggcuaug ucccagggua gugccauuac acuguuauua cugaguguuu uucuagcgac 27720
uuggcugcug ggcuauggcu uugcccucua acuagcgguc uuggucuugc acacaacggu 27780
aagccagugg uaaugucagu gcaagaagga uauuaccaua gcacugucau gaggggaacg 27840
caguaccuuu ucaucuaaac cuuugcacga guaaucaaag auccgcuuga cgagccuaua 27900
uggaagagcg ugccagguau uugacucaag gacuguuagu aacugaagac cugacggugu 27960
<210> 33
<211> 27976
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDV 1251-125-10 (125-10) passage30 Genomic cDNA
<400> 33
aattcaacta aacgaaattt tgtccttccg gccgcatgtc catgctgctg gaagctgacg 60
tggaatttca ttaggtttgc ttaagtagcc atcgcaagtg ctgtgctgtc ctctagttcc 120
tggttggcgt tccgtcgcct tctacatact agacaaacag ccttcctccg gttccgtctg 180
ggggttgtgt ggataactag ttccgtctag tttgaaacca gtaactgtcg gctatggcta 240
gcaaccatgt tacattggct tttgccaatg atgcagaaat ttcagctttt ggcttttgca 300
ctgctagtga agccgtctca tactattctg aggccgccgc tagtggattt atgcaatgcc 360
gtttcgtgtc cttcgatctc gctgacactg ttgagggatt gcttcccgaa gactatgtca 420
tggtggtggt cggcactacc aagcttagtg cgtatgtgga cacttttggt agccgcccca 480
aaaacatttg tggttggctg ttattttcta actgtaatta cttcctcgaa gagttagagc 540
ttacttttgg tcgtcgtggt ggtaacatcg tgccagttga ccaatacatg tgtggcgctg 600
acggtaaacc tgttcttcag gaatccgaat gggagtatac agatttcttt gctgactccg 660
aagacggtca actcaacatt gctggtatca cttatgtgaa ggcctggatt gtagagcgat 720
cggatgtctc ttatgcgagt cagaatttaa catctattaa gtctattact tactgttcaa 780
cctatgagca tacttttcct gatggtactg ccatgaaggt tgcacgtact ccaaagatta 840
agaagactgt tgtcttgtct gagccacttg ctactatcta cagggaaatt ggttctcctt 900
ttgtggataa tgggagcgat gctcgttcta tcattaagag accagtgttc ctccacgctt 960
ttgttaagtg taagtgtggt agttatcatt ggactgttgg tgattggact tcctatgtct 1020
ccacttgctg tggctttaag tgtaagccag tccttgtggc ttcatgctct gctacgcctg 1080
gttctgttgt ggttacgcgc gctggtgctg gcactggtgt taagtattac aacaacatgt 1140
tcctgcgcca tgtggcagac attgatgggt tggcattctg gcgaattctc aaggtgcagt 1200
ccaaagacga cctcgcttgc tctggtaaat tccttgaaca ccatgaggaa ggtttcacag 1260
atccttgcta ctttttgaat gactcgagca ttgctactaa gctcaagttt gacatcctta 1320
gtggcaagtt ttctgatgaa gtcaaacaag ctatctttgc tggtcatgtt gttgttggca 1380
gcgcgmtcgt tgacattgtt gacgatgcac tgggacagcc ttggtttata cgtaagcttg 1440
gtgaccttgc aagtgcagct tgggagcagc ttaaggctgt cgttagaggc cttaacctcc 1500
tgtctgatga ggtcgtgctc tttggcaaaa gacttagctg tgccactctt agtatcgtta 1560
acggtgtttt tgagttcatc gccgaagtgc ctgagaagtt ggctgcggct gttacagttt 1620
ttgtcaactt cttgaatgag ctttttgagt ctgcctgtga ctgcttaaag gtcggaggta 1680
aaacctttaa caaggttggc tcttatgttc tttttgacaa cgcattggtt aagcttgtca 1740
aggcaaaagt tcgcggccca cgacaggcag gtgtttgtga agttcgttac acaagccttg 1800
ttattgggag tactaccaag gtggtttcca agcgcgttga aaatgccaat gtgaatctcg 1860
tcgtcgttga cgaggatgtg accctcaaca ccactggtcg tacagttgtt gttgacggac 1920
ttgcattctt cgagagtgac gggttttaca gacatcttgc tgatgctgac gttgtcattg 1980
aacatcctgt ttataagtct gcttgtgagc tcaagccagt ttttgagtgt gacccaatac 2040
ctgattttcc tatgcctgtg gccgctagtg ttgcagagct ttgtgtgcaa actgatctgt 2100
tgcttaaaaa ttacaacact ccttataaaa cttacagctg cgttgtgaga ggtgataagt 2160
gttgtatcac ttgcacctta catttcacag caccaagtta tatggaggct gctgctaatt 2220
ttgtagacct ctgtaccaag aacattggta ctgctggttt tcatgagttt tacattacgg 2280
cccatgaaca acaggatctg caagggttcg taaccacttg ttgcacgatg tcaggttttg 2340
agtgttttat gcctataatc ccacagtgtc cagcagtgct tgaagagatt gatggtggta 2400
gcatctggcg gtcttttatc actggtctta atacaatgtg ggatttttgc aagcatctta 2460
aagtcagctt tggactagat ggcattgttg tcactgtagc acgcaaattt aaacgacttg 2520
gtgctctctt ggcagaaatg tataacactt acctttcaac tgtggtggaa aacttggtac 2580
tggccggtgt tagcttcaag tattatgcca ccagtgtccc aaaaattgtt ttgggctgtt 2640
gttttcacag tgttaaaagt gttcttgcaa gtgccttcca gattcctgtc caggcaggcg 2700
ttgagaagtt taaagtcttc cttaactgtg ttcaccctgt tgtaccacgt gtcattgaaa 2760
cttcttttgt ggaattagaa gagacgacat ttaaaccacc agcactcaat ggtagtattg 2820
ctattgttga tggctttgct ttctattatg atggaacact atactatccc accgatggta 2880
atagcgttgt tcctatctgc tttaagaaga aaggtggtgg tgatgtcaaa ttctctgatg 2940
aagtctctgt taaaaccatt gacccagttt ataaggtctc ccttgaattt gagttcgagt 3000
ctgagactat tatggctgtg cttaataagg ctgttggtaa ttgtatcaag gttacaggtg 3060
gttgggacga tgttgttgag tatatcaatg ttgccattga ggttcttaaa gatcacatcg 3120
atgtgcctaa gtactacatc tatgatgagg aaggtggcac cgatcctaat ctgcccgtaa 3180
tggtttctca gtggccgttg aatgatgaca cgatctcaca ggatctgctt gatgttgaag 3240
ttgttactga tgcgccagtt gatttcgagg gtgatgaagt agactcctct gaccctgwta 3300
aggtggcaga cgtggctaac tctgagcctg aggatgacgg tcttaatgta gctcctgaaa 3360
caaatgtaga gtctgaagtt gaggaagttg ccgcaacctt gtcctttaaa gatacacctt 3420
ccacagttac taaggatcct tttgcttttg actttgcaag ctatggagga cttaaggttt 3480
taagacaatc tcataacaac tgctgggtta cttctacctt ggtgcagcta caattgcttg 3540
gcatcgttga tgaccctgca atggagcttt ttagtgctgg tagagttggt ccaatggttc 3600
gcaaatgcta tgagtcacaa aaggctatct tgggatcttt gggtgatgtg tcggcttgcc 3660
tagagtctct gactaaggac ctacacacac ttaagattac ctgttctgta gtctgtggtt 3720
gtggtactgg tgaacgtatc tatgatggtt gtgcttttcg tatgacgcca actttggaac 3780
cgttcccata tggtgcttgt gctcagtgtg ctcaagtttt gatgcacact tttaaaagta 3840
ttgttggcac cggcatcttt tgtcgagata ctactgctct ctccttggat tctttggttg 3900
taaaacctct ttgtgcggct gcttttatag gcaaggatag tggtcattat gtcactaact 3960
tttatgatgc tgctatggct attgatggtt atggtcgtca tcagataaag tatgacacac 4020
tgaacactat ttgtgttaaa gacgttaatt ggacagcacc ttttgtccca gacgttgagc 4080
ctgtattgga gcctgttgtc aaacctttct attcttataa gaatgttgat ttttaccaag 4140
gagattttag tgaccttgtt aaacttccat gtgattttgt tgttaatgct gcaaatgaga 4200
atttgtctca cggtggcggc atagcaaagg ccattgatgt ttataccaag ggcatgttgc 4260
agaagtgctc gaatgattac attaaagcac acggtcccat taaagttgga cgtggtgtca 4320
tgttggaggc attaggtctt aaggtcttta atgttgttgg tccacgtaag ggtaagcatg 4380
cacctgagct tcttgttaag gcttataagt ccgtttttgc taattcaggt gttgctctta 4440
cacctttgat tagtgttgga atttttagtg ttcctttgga agaatcttta tctgcttttc 4500
ttgcatgtgt tggtgatcgc cactgtaagt gcttttgtta tagtgacaaa gagcgcgagg 4560
cgatcattaa ttacatggat ggcttggtag atgctatttt caaagatgca cttgttgata 4620
ctactcctgt ccaggaagat gttcaacaag tttcacaaaa accagttttg cctaattttg 4680
aacctttcag gattgaaggt gctcatgctt tctatgagtg caaccctgaa ggtttgatgt 4740
cattaggtgc tgacaagctg gtgttgttta caaattccaa tttggatttt tgtagcgttg 4800
gtaagtgtct taacaatgtg actggcggtg cattgcttga agccataaat gtatttaaaa 4860
agagtaacaa aacagtgcct gctggcaact gtgttacttt tgagtgtgca gatatgattt 4920
ctattactat ggtagtattg ccatctgacg gtgatgctaa ttatgacaaa aattatgcac 4980
gcgccgtcgt caaggtatct aagcttaaag gcaagttatt gcttgctgtt ggtgatgcca 5040
tgttgtattc caagttgtcc cacctcagcg tgttaggttt cgtatccaca cctgatgatg 5100
tggagcgttt ctacgcaaat aagagtgtgg ttattaaagt tactgaggat acacgtagtg 5160
ttaagactgt taaagtagaa tccactgtta cttatggaca acaaattgga ccttgtcttg 5220
ttaatgacac cgttgtcaca gacaacaaac ctgttgttgc tgatgttgta gctaaggttg 5280
taccaagtgc taattgggat tcacattatg gttttgataa ggctggtgag ttccacatgc 5340
tagaccatac tgggtttgcc tttcctagtg aagttgttaa cggtaggcgt gtgcttaaaa 5400
ccacagataa taactgttgg gttaatgtta catgtttaca attacagttt gctagattta 5460
ggttcaagtc agcaggtcta caggctatgt gggagtccta ttgtactggt gatgttgcta 5520
tgtttgtgca ttggttgtac tggcttactg gtgttgacaa aggtcagcct agtgattcag 5580
aaaatgcact taacatgttg tctaagtaca ttgttcctgc tggttctgtc actattgaac 5640
gtgtcacgca tgacggttgt tgttgtagta agcgtgttgt cactgcacca gttgtgaatg 5700
ctagcgtgtt gaagcttggc gtcgaggatg gtctttgtcc acatggtctt aactacattg 5760
acaaagttgt tgtagttaaa ggtactacaa ttgttgtcaa tgttggaaaa cctgtagtgg 5820
caccatcgca cctctttctt aagggtgttt cctacacaac attcctagat aatggtaacg 5880
gtgttgccgg ccattatact gtttttgatc atgacactgg tatggtgcat gatggagatg 5940
tttttgtacc aggtgatctc aatgtgtctc ctgttacaaa tgttgtcgtc tcagagcaga 6000
cggctgttgt gattaaagac cctgtgaaga aagtagagtt agacgctaca aagctgttag 6060
acactatgaa ttatgcatcg gaaagattct tttcctttgg tgattttatg tcacgtaatt 6120
taattacagt gtttttgtac atccttagta ttttgggtct ctgttttagg gcctttcgta 6180
agagggatgt taaagttcta gctggtgtac cccaacgtac tggtattata ttgcgtaaaa 6240
gtgtgcgcta taatgcaaag gctttgggtg tcttcttcaa gctaaaactt tattggttca 6300
aagttcttgg taagtttagt ttgggtattt atgcattgta tgcattacta ttcatgacaa 6360
tacgctttac acctataggt ggccctgttt gtgatgatgt tgttgctggt tatgctaatt 6420
ctagttttga caagaatgag tattgcaaca gtgttatttg taaggtctgt ctctatgggt 6480
accaggaact ttcggacttc tctcacacac aggtagtatg gcaacacctt agagacccat 6540
taattggtaa tgtgatgcct ttcttttatt tggcatttct ggcaattttt gggggtgttt 6600
atgtaaaggc tattactctc tattttattt tccagtatct taacatactt ggtgtgtttt 6660
tgggcctaca acagtccatt tggtttttgc agcttgtgcc ttttgatgtc tttggtgacg 6720
agatcgtcgt ctttttcatc gttacacgcg tattgatgtt ccttaagcat gttttccttg 6780
gctgcgataa ggcatcttgt gtggcttgct ctaagagtgc tcgccttaag cgcgttcctg 6840
tccagactat ttttcagggt actagcaaat ccttctacgt acatgccaat ggtggttcta 6900
agttctgtaa gaagcacaat ttcttttgtt taaattgtga ttcttatggt ccaggctgca 6960
cttttattaa tgacgtcatt gcaactgaag ttggtaatgt tgtcaaactt aatgtgcaac 7020
cgacaggtcc tgccactatt cttattgaca aggttgaatt cagtaatggt ttttactatc 7080
tttatagtgg tgacacattt tggaagtaca actttgacat aacagataac aaatacactt 7140
gcaaagagtc acttaaaaat tgtagcataa tcacagactt tattgttttt aacaataatg 7200
gttccaatgt aaatcaggtt aagaatgcat gtgtgtattt ttcacagatg ctttgtaaac 7260
ctgttaagtt agtggactca gcgttgttgg ccagtttgtc tgttgatttt ggtgcaagct 7320
tacatagtgc ttttgttagt gtgttgtcga atagttttgg caaagacctg tcaagttgta 7380
atgacatgca ggattgcaag agcacattgg gttttgatga tgtaccattg gataccttta 7440
atgctgctgt tgctgaggct catcgttacg atgtcctctt gactgacatg tcgttcaaca 7500
attttaccac cagttatgca aaaccagagg aaaaacttcc cgtccatgac attgccacgt 7560
gtatgcgtgt aggtgccaag attgttaatc ataacgttct tgtcaaggat agtatacctg 7620
tggtgtggct tgtacgtgat ttcattgccc tttctgaaga aactaggaag tacattattc 7680
gtacgactaa agttaagggt ataaccttca tgttgacctt taatgattgt cgtatgcata 7740
ctaccatacc tactgtttgc attgcaaata agaagggtgc aggtcttcct agtttttcaa 7800
aggttaagaa attcttctgg tttttgtgtc tgttcatagt tgctgttttc tttgcactaa 7860
gcttttttga ttttagtact caggttagca gtgatagtga ttatgacttc aagtatattg 7920
agagtggcca gttgaagact tttgacaatc cacttagttg tgtgcataat gtctttagta 7980
acttcgacca gtggcatgat gccaagtttg gtttcacccc cgtcaacaat cctagttgtc 8040
ctatagtcgt tggtgtatca gacgaagcgc gcactgttcc aggtatccca gcaggtgttt 8100
atttagctgg taaaacactt gtttttgcta ttaacaccat ttttggtaca tctggtttgt 8160
gctttgatgc tagtggcgtt gctgataagg gcgcttgcat ttttaattcg gcttgcacca 8220
cattatctgg tttgggtgga actgctgtct actgttataa gaatggtcta gttgaaggtg 8280
ctaaacttta tagtgagttg gcacctcata gctactataa aatggtagat ggtaatgctg 8340
tgtctttacc tgaaattatc tcacgcggct ttggcatccg tactatccgt acaaaggcta 8400
tgacctactg tcgcgttggc cagtgtgtgc aatctgcaga aggtgtttgt tttggcgccg 8460
atagattctt tgtctataat gcagaatctg gttctgactt tgtttgtggc acagggctct 8520
ttacattgtt gatgaacgtt attagtgttt tttccaagac agtaccagta actgtgttgt 8580
ctggtcaaat actttttaat tgcattattg cttttgctgc tgttgcggtg tgtttcttat 8640
ttacaaagtt taagcgcatg ttcggtgata tgtctgttgg cgttttcact gtcggtgctt 8700
gtactttgtt gaacaatgtt tcctacattg taacacagaa cacacttggc atgttgggct 8760
atgcaacttt gtactttttg tgcactaaag gtgttagata tatgtggatt tggcatttgg 8820
gatttttgat ctcatatata cttattgcac catggtgggt tttgatggtt tatgcctttt 8880
cagccatttt tgagtttatg cctaaccttt ttaagcttaa ggtttcaaca caactttttg 8940
agggtgacaa gttcgtaggc tcttttgaaa atgctgcagc aggtacattt gtgcttgata 9000
tgcatgccta tgagagactt gccaactcta tctcaactga aaaactgcgt cagtatgcta 9060
gtacttacaa taagtacaag tattattcag gcagtgcttc agaggctgat tacaggcttg 9120
cttgttttgc ccatttggcc aaggctatga tggattatgc ttctaatcac aacgacacgt 9180
tatacacacc acccactgtg agttacaatt caactctaca ggctggcttg cgtaagatgg 9240
cacaaccatc tggtgttgtt gagaagtgca tagttcgtgt ttgctatggt aatatggctc 9300
ttaatggcct atggcttggt gatactgtta tctgcccacg ccatgttata gcgtctagta 9360
ctactagcac tatagattat gactatgccc tttctgtttt acgcctcyac aacttctcca 9420
tttcatctgg taatgttttc ctaggtgttg tgggtgtaac catgcgaggt gctttgttgc 9480
agataaaggt taatcaaaac aatgtccaca cgcctaagta cacctatcgc acagttagac 9540
cgggtgaatc ttttaatatc ttggcgtgct atgatggttc tgcagctggt gtttacggcg 9600
ttaacatgcg ctctaattac actattagag gctcgttcat taatggcgct tgtggttcac 9660
ctggttataa cattaacaat ggtaccgttg agttttgcta tttacaccag cttgaacttg 9720
gttcaggctg tcatgttggt agcgacttag atggtgttat gtatggtggt tatgaggacc 9780
aacctacttt gcaagttgaa ggcgctagta gtctgtttac agagaatgtg ttggcatttc 9840
tttatgcagc actcattaat ggttctacct ggtggcttag ttcttctagg attgctgtag 9900
acaggtttaa tgagtgggct gttcataatg gtatgacaac agtagttaat actgattgct 9960
tttctattct tgctgctaag actggtgttg atgtacaacg tttgttggcc tcaatccagt 10020
ctctgcataa gaattttggt ggaaagcaaa ttcttggcta tacctcgttg acagatgagt 10080
ttactacagg tgaagttata cgtcaaatgt atggcgttwa tcttcagagt ggttatgttt 10140
cacgcgcctg tagaaatgtc ttgctggttg gttcttttct gactttcttt tggtcagaat 10200
tagtttccta cactaagttc ttttgggtaa atcctggtta tgtcacacct atgtttgcgt 10260
gtttgtcatt gctgtcctca cttttgatgt tcacactcaa gcataagaca ttgtttttcc 10320
aggtctttct aatacctgct ctgattgtta catcttgcat taatttggca tttgatgttg 10380
aagtctacaa ctatttggca gagcattttg attaccatgt ttctctcatg ggttttaatg 10440
cacaaggtct tgttaacatc tttgtctgct ttgttgttac cattttacac ggcacataca 10500
catggcgctt ttttaacaca cctgtgagtt ctgtcactta tgtggtagct ttgctgactg 10560
cggcatataa ctatttttac gctagtgaca ttcttagttg tgctatgaca ctatttgcta 10620
gtgtgactgg caactggttc gttggtgctg tttgttataa agctgctgtt tatatggcct 10680
tgagatttcc tacttttgtg gctatttttg gtgatattaa gagtgttatg ttctgttacc 10740
ttgtgttggg ttattttacc tgttgcttct acggtattct ctactggttc aacaggtttt 10800
ttaaggttag tgtaggtgtc tatgactata ctgttagtgc tgctgagttt aagtatatgg 10860
ttgctaacgg cctacgtgca ccaactggaa cacttgattc actacttctg tctgccaaat 10920
tgattggtat tggtggtgag cggaatatta agatttcttc cgttcagtct aaactgactg 10980
atattaagtg tagtaacgtt gtgcttttag gctgtctctc tagcatgaat gtctcagcaa 11040
attcaacaga atgggcctat tgtgttgact tgcataacaa gatcaacttg tgtaatgacc 11100
cagaaaaagc gcaggaaatg ctacttgctt tgttggcatt tttccttagt aagaatagtg 11160
cttttggttt agatgactta ttggaatcct attttaatga caatagtatg ttgcagagtg 11220
ttgcatctac ttatgtcggt ttgccttctt atgtcattta tgaaaatgca cgccaacagt 11280
atgaagatgc tgttaataat ggttctccac ctcagttggt taagcaattg cgccatgcca 11340
tgaatgtagc aaagagcgaa tttgaccgtg aggcttctac tcagcgtaag cttgatagaa 11400
tggcggaaca ggctgcagca cagatgtaca aagaggcacg agcagttaat aggaagtcca 11460
aagttgtaag tgctatgcat tcactgcttt ttggtatgtt gagacgtttg gacatgtctt 11520
ctgtagacac cattctcaac ttggcaaagg atggggttgt acctctgtct gtcataccgg 11580
cagtcagtgc tactaagctt aacattgtta cttctgatat cgattcttat aatcgtatcc 11640
agcgtgaggg atgtgtccac tacgctggta ccatttggaa tataattgat atcaaggaca 11700
atgatggcaa ggtggtacac gttaaggagg taaccgcaca gaatgctgag tccctgtcat 11760
ggcccctggt ccttgggtgt gagcgtattg tcaagctcca gaataatgaa attattcctg 11820
gtaagctgaa gcagcgctcc attaaggcag aaggagatgg catagttgga gaaggtaagg 11880
cactttacaa taatgagggt ggacgtactt ttatgtatgc tttcatctcg gacaaaccgg 11940
acctgcgtgt agtcaagtgg gagttcgatg gtggttgtaa cactattgag ctagaaccac 12000
cacgtaagtt cttggtggat tctcctaatg gtgcacagat caagtatctc tactttgttc 12060
gtaaccttaa cacgttacgt aggggtgctg ttctcggcta cataggtgcc actgtacgct 12120
tgcaggctgg taaacaaaca gaacaggcta ttaactcttc attgttgaca ctttgcgctt 12180
tcgctgtgga tcctgctaag acctacatcg atgctgtcaa aagtggtcac aaaccagtag 12240
gtaactgtgt taagatgttg gccaatggtt ctggtaatgg acaagctgtt actaatggtg 12300
tggaggctag tactaaccag gattcatacg gtggtgcgtc cgtgtgtcta tattgtagag 12360
cacatgttga gcatccatct atggatggtt tttgcagact gaaaggcaag tacgtacagg 12420
ttccactagg tacagtggat cctatacgtt ttgtacttga gaatgacgtt tgcaaggttt 12480
gtggttgttg gctggctaat ggctgcactt gtgacagatc cattatgcaa agcactgata 12540
tggcttattt aaacgagtac ggggctctag tgcagctcga ctagagccct gtaacggtac 12600
tgatacacaa catgtgtatc gtgcttttga catctacaac aaggatgttg cttgtctagg 12660
taaattcctc aaggtgaact gtgttcgcct gaagaatttg gataagcatg atgcattcta 12720
tgttgtcaaa agatgtacca agtctgcgat ggaacacgag caatccatct atagcagact 12780
tgaaaagtgt ggagccgtag ccgaacacga tttcttcact tggaaggatg gtcgtgccat 12840
ctatggtaac gtttgtagaa aggatcttac cgagtatact atgatggatt tgtgttacgc 12900
tttacgtaac tttgatgaaa acaattgcga tgttcttaag agcattttaa ttaaggtagg 12960
cgcttgtgag gagtcctact tcaataataa agtctggttt gaccctgttg aaaatgaaga 13020
cattcatcgt gtctatgcat tgttaggtac cattgtttca cgtgctatgc ttaaatgcgt 13080
taagttctgt gatgcaatgg ttgaacaagg tatagttggt gttgtcacat tagataatca 13140
ggatcttaat ggtgattttt atgattttgg tgattttact tgtagcatca agggaatggg 13200
tatacccatt tgcacatcat attactctta tatgatgcct gttatgggta tgactaattg 13260
ccttgctagt gagtgttttg ttaagagtga tatatttggt gaggatttca agtcatatga 13320
cctgctggaa tatgatttca cggagcataa gacagcactc ttcaacaagt atttcaagta 13380
ttggggactg caataccacc ctaactgtgt ggactgcagt gatgagcagt gcatagttca 13440
ctgtgccaac ttcaatacgt tgttttccac tactatacct attacggcat ttggaccttt 13500
gtgtcgcaag tgttggattg atggtgttcc actggtaact acagctggtt atcattttaa 13560
acagttaggt atagtttgga acaatgacct caacttacac tctagcaggc tctctattaa 13620
cgaattactc cagttttgta gtgatcctgc attgcttata gcatcatcac cagcccttgt 13680
tgatcagcgt actgtttgct tttcagttgc agcgctaggt acaggtatga ctaaccagac 13740
tgttaaacct ggccatttca ataaggagtt ttatgacttc ttacttgagc aaggtttctt 13800
ttctgagggc tctgagctta ctttaaagca cttcttcttt gcacagaagg gtgatgcagc 13860
tgttaaggat tttgactact ataggtataa tagacctact gttctggaca tttgccaagc 13920
tcgcgtcgtg tatcaaatag tgcaacgcta ttttgatatt tacgaaggtg gttgtatcac 13980
tgctaaagag gtggttgtta caaaccttaa caagagcgca ggttatcctt tgaacaagtt 14040
tggtaaagct ggtctttact atgagtcttt atcctatgag gaacaggatg aactttatgc 14100
ttatactaag cgtaacatcc tgcccactat gacacagctc aaccttaaat atgctataag 14160
tggcaaagaa cgtgcacgca cagtgggtgg tgtttcgctt ttgtcaacca tgactactcg 14220
gcagtatcat cagaaacacc ttaagtccat agttaatact aggggcgctt cggttgttat 14280
tggtactact aagttttatg gtggttggga caatatgctt aagaacctta ttgatggtgt 14340
tgaaaatccg tgtcttatgg gttgggacta cccaaagtgc gacagagcac tgcccaatrt 14400
gatacgtatg atttcagcca tgattttagg ctctaagcac accacatgct gcagttccac 14460
tgaccgcttt ttcaggttgt gcaatgaatt ggctcaagtc cttactgagg ttgtttattc 14520
taatggaggt ttttatttga agccaggtgg tactacctct ggtgatgcaa ccaccgcata 14580
tgcaaactca gtttttaata tcttccaagc agtaagtgcc aatgttaaca aacttcttag 14640
tgttgacagc aatgtctgtc ataatttaga agttaagcaa ttgcagcgta agctttatga 14700
gtgctgttat agatcaacta ccgtcgatga ccagttcgtc gttgagtatt atggttactt 14760
gcgtaaacat ttttcaatga tgattctttc tgatgatggc gttgtttgtt ataacaatga 14820
ctatgcatca cttggttatg tcgctgatct taacgcattc aaggctgttt tgtattacca 14880
gaacaatgtc ttcatgagcg cctctaaatg ttggatcgag cctgacatta ataaaggtcc 14940
tcatgaattt tgctcgcagc atactatgca gattgtcgat aaagatggta cttattacct 15000
tccttaccct gatccttcaa gaattctctc tgcaggtgtg tttgttgatg acgttgttaa 15060
aactgatgca gttgtattgc ttgaacgtta tgtgtcattg gctatagatg cctacccgtt 15120
atctaagcat gaaaaccctg aatataagaa ggtgttttat gtgcttttgg attgggttaa 15180
gcatctgtac aaaactctta atgctggtgt gttagagtct ttttctgtca cacttttgga 15240
agattctact gctaaattct gggatgagag cttttatgcc aacatgtatg agaaatctgc 15300
agttttacaa tctgcagggc tttgtgttgt ttgtggctct caaactgttt tacgttgtgg 15360
tgattgtcta cggcgtccta tgctttgtac taagtgtgct tatgatcatg tcattggaac 15420
aactcacaag ttcattttgg ccatcactcc atatgtgtgt tgtgcttcag attgtggtgt 15480
caatgatgta actaagctct acttaggtgg tcttagttat tggtgtcatg accacaagcc 15540
acgtcttgca ttcccgttgt gctctgctgg taatgttttt ggcttgtaca aaaattctgc 15600
taccggctca cccgatgttg aagactttaa tcgcattgct acatccgatt ggactgatgt 15660
ttctgactac aggttggcaa atgatgtcaa ggactcattg cgtctgtttg cagcggaaac 15720
tatcaaggcc aaggaggaga gcgttaagtc atcctatgct tgtgcaacac tacatgaggt 15780
tgtaggacct aaagagttgt tgctcaaatg ggaagtcggc agacccaaac caccccttaa 15840
tagaaattcg gttttcactt gttatcatat aacgaagaac accaaatttc aaatcggtga 15900
gtttgtgttt gagaaggcag aatatgataa tgatgctgta acatataaaa ctaccgccac 15960
aacaaaactt gttcctggca tggtttttgt gcttacctca cataatgttc agccattgcg 16020
cgcaccgacc attgctaatc aagaacgtta ttccactata cataagttgc atcctgcttt 16080
taacatacct gaagcttatt ctagcttagt gccctattac caattgattg gtaagcagaa 16140
gattacaact attcagggac ctcccggtag tggtaaatct cactgtgtta tagggctagg 16200
tttgtactat ccaggtgcac gtatagtgtt tacagcttgt tctcatgcag cggtcgattc 16260
actttgtgtg aaagcttcca ctgcttatag caatgacaaa tgttcacgca tcataccaca 16320
gcgcgctcgt gttgagtgtt atgatggttt caagtctaat aatactagtg ctcagtacct 16380
tttctctact gtcaatgctt tgccagagtg caatgcggac attgttgtgg tggatgaggt 16440
ctctatgtgc actaattatg acttgtctgt cataaatcag cgcatcagct ataggcatgt 16500
agtctatgtt ggtgaccctc aacagctgcc tgcaccacgt gttatgattt cacgtggtac 16560
tttggaacca aaggactaca acgttgtcac tcaacgcatg tgtgccctta agcctgatgt 16620
tttcttgcac aagtgttatc gctgtcctgc tgagatagtg cgtactgtgt ctgagatggt 16680
ctatgaaaac caattcattc ctgtgcaccc agatagcaag cagtgtttta aaatcttttg 16740
caagggtaat gttcaggttg ataatggttc aagcattaat cgcaggcaat tggatgttgt 16800
gcgtatgttt ttggctaaaa atcctaggtg gtcaaaggct gtttttattt ctccttataa 16860
cagccagaat tatgttgcca gccgcatgct aggtctacaa attcagacag ttgactcatc 16920
ccagggtagt gagtatgact atgtcattta cacacaaact tcagatactg cccatgcctg 16980
taatgttaac aggtttaatg ttgccatcac aagggccaag aaaggcatat tatgtataat 17040
gtgcgatagg tccctttttg atgtgcttaa attctttgag cttaaattgt ctgatttgca 17100
ggctaatgag ggttgtggtc tttttaaaga ctgtagcaga ggtgatgatc tgttgccacc 17160
atctcacgct aacaccttca tgtctttagc ggacaatttt aagactgatc aagatcttgc 17220
tgttcaaata ggtgttaatg gacccattaa atatgagcat gttatctcgt ttatgggttt 17280
ccgttttgat atcaacatac ccaaccatca tactctcttt tgcacacgcg actttgccat 17340
gcgcaatgtt agaggttggt taggctttga cgttgaagga gcacatgttg ttggctctaa 17400
cgtcggtaca aatgtcccat tgcaattagg gttttctaac ggtgttgatt ttgttgtcag 17460
acctgaaggt tgcgttgtaa cagagtctgg tgactacatt aaacccgtca gagctcgtgc 17520
tccaccaggg gaacaattcg cacacctttt gcctttactt aaacgcggcc aaccatggga 17580
tgttgtccgc aaacgtatag tgcagatgtg tagtgactac ctggccaacc tatcagacat 17640
actaattttt gtgttgtggg ctggtggttt ggagttgaca actatgcgtt attttgtcaa 17700
gattggacca agtaagagtt gtgattgtgg taaggttgct acttgttaca atagtgcgct 17760
gcatacgtac tgttgtttca aacatgccct tggttgtgat tatctgtata acccatactg 17820
tattgatata cagcagtggg gatacaaggg atcacttagc cttaaccacc atgagcattg 17880
taatgtacat agaaacgagc atgtggcttc tggtgatgcc ataatgactc gctgtctggc 17940
catacatgat tgctttgtca agaacgttga ctggtccatc acatacccat ttattggtaa 18000
tgaggctgtt attaataaga gcggccgaat tgtgcaatca cacactatgc ggtcagttct 18060
taagttatac aatccgaaag ccatatatga tattggcaat cctaagggca ttagatgtgc 18120
cgtaacggat gctaagtggt tttgctttga caagaatcct aytaattcta atgtcaagac 18180
attggagtat gactatataa cacatggcca atttgatggg ttgtgcttgt tttggaattg 18240
caatgtagac atgtatccag aattttctgt ggtctgtcgt tttgatactc gctgtaggtc 18300
accactcaac ttggagggtt gtaatggtgg ttcactgtat gttaataatc atgcattcca 18360
tacaccggct tttgacaagc gtgcttttgc taagttgaag ccaatgccat ttttctttta 18420
tgatgatact gagtgtgaca agttacagga ctccataaac tatgttcctc ttagggctag 18480
taactgcatt actaaatgta atgttggtgg tgctgtctgt agtaagcatt gtgctatgta 18540
tcatagctat gttaatgctt acaacacttt tacgtcggcg ggctttacta tttgggtgcc 18600
tacttcgttt gacacctata atctgtggca gacatttagt aacaatttgc aaggtcttga 18660
gaacattgct ttcaatgtcg taaagaaagg atcttttgtt ggtgccgaag gtgaacttcc 18720
tgtagctgtg gttaatgaca aagtgctcgt tagagatggt actgttgata ctcttgtttt 18780
tacaaacaag acatcactac ccactaacgt agcttttgag ttgtatgcca agcgtaaggt 18840
aggactcacc ccacccatta cgatcctacg taacttgggt gtagtttgta catctaagtg 18900
tgtcatttgg gactatgaag ccgaacgtcc acttactact tttacaaagg atgtttgtaa 18960
atataccgac tttgagggtg acgtctgtac actctttgat aacagcattg ttggttcatt 19020
agagcgattc tccatgaccc aaaatgctgt gcttatgtca cttacagctg ttaaaaagct 19080
taytggcata aagttaactt atggttatct taatggtgtc ccagttaaca cacatgaaga 19140
taaacctttt acttggtata tttacactag gaagaacggc aagttcgagg accatcctga 19200
tggctatttt acccaaggta gaacaaccgc tgattttagc cctcgtagcg acatggaaaa 19260
ggacttccta agtatggata tgggtctgtt tattaacaag tacggacttg aagattacgg 19320
ctttgagcac gttgtgtatg gtgatgtttc aaaaaccacc cttggtggtt tgcatctact 19380
aatttcgcag gtgcgtctgg cctgtatggg tgtgctcaaa atagacgagt ttgtgtctag 19440
taatgatagc acgttaaagt cttgtactgt tacatatgct gataacccta gtagtaagat 19500
ggtttgtacg tatatggatc tcctgcttga cgattttgtc agcattctta aatctttgga 19560
tttgggcgtt gtatctaaag ttcatgaagt tatggtcgat tgtaaaatgt ggaggtggat 19620
gttgtggtgt aaggatcata aactccagac attttatccg caacttcagg ccagtgaatg 19680
gaagtgtggt tattccatgc cttctattta caagatacaa cgtatgtgtt tagaaccttg 19740
caatctctac aactatggtg ctggtattaa gttacctgat ggcattatgt ttaacgtagt 19800
taaatacaca cagctttgtc aatatctcaa tagcaccaca atgtgtgtac cccatcacat 19860
gcgtgtgcta catcttggtg ctggctccga caagggtgtt gcacctggca cggctgtctt 19920
acgacgttgg ttgccactgg atgccattat agttgacaat gatagtgtgg attacgttag 19980
cgatgctgat tatagtgtta caggagattg ctctacctta tacctgtcag ataagtttga 20040
tttagttata tctgatatgt atgatggtaa gattaaaagt tgtgatgggg agaacgtgtc 20100
taaagaaggc ttctttccct atattaatgg tgtcatcacc gaaaagttgg cacttggtgg 20160
tactgtagct attaaggtga cggagtttag ttggaataag aagttgtatg aactcattca 20220
gaggtttgag tattggacaa tgttctgtac cagtgttaac acgtcatcgt cagaggcatt 20280
cttaattggt gttcactatt taggtgattt tgcaagtggc gctgtgattg acggcaacac 20340
tatgcatgcc aattatatct tctggcgtaa ttccacaatt atgactatgt cttacaatag 20400
tgtacttgat ttaagcaagt tcaattgtaa gcataaggct acagttgtca ttaatttaaa 20460
agattcatcc attagtgatg ttgtgttagg tttgttgaag aatggtaagt tgctagtgcg 20520
taataatgac gccatttgtg gtttttctaa tcatttggtc aacgtaaaca aatgaagtct 20580
ttaacctact tctggttgtt cttaccagta ctttcaacac ttagcctacc acaagatgtc 20640
accaggtgct cagctaacac taattttagg cggttctttt caaaatttaa tgttcaggcg 20700
cctgcagttg ttgtactggg cggttatcta cctattggtg aaaaccaggg tgtcaattca 20760
acttggtact gtgctggcca acatccaact gctagtggcg ttcatggtat ctttgttagc 20820
catattagag gtggtcatgg ctttgagatt ggcatttcgc aagagccttt tgaccctagt 20880
ggttaccagc tttatttaca taaggctact aacggtaaca ctaatgctac tgcgcgactg 20940
cgcatttgcc agtttcctag cattaaaaca ttgggcccca ctgctaataa tgatgttaca 21000
ataggtcgta attgcctatt taacaaagcc atcccagctc atatgagtga acatagtgtt 21060
gtcggcataa catgggataa tgatcgtgtc actgtctttt ctgacaagat ctattatttt 21120
tattttaaaa atgattggtc ccgtgttgcg acaaagtgtt acaacagtgg aggttgtgct 21180
atgcaatatg tttacgaacc cacctattac atgcttaatg ttactagtgc tggtgaggat 21240
ggtatttctt atcaaccctg tacagctaat tgcattggtt atgctgccaa tgtatttgct 21300
actgagccca atggccacat accagaaggt tttagtttta ataattggtt tcttttgtcc 21360
aatgattcca ctttggtgca tggtaaggtg gtttccaacc aaccattgtt ggtcaattgt 21420
cttttggcca ttcctaagat ttatggacta ggccaatttt tctcctttaa tcaaacgatc 21480
gatggtgttt gtaatggagc tgctgtgcag cgtgcaccag aggctctgag gtttaatatt 21540
aatgacacct ctgtcattct tgctgaaggc tcaattgtac ttcatactgc tttaggaaca 21600
aatttttctt ttgtttgcag taattcccca aatcctcact tagccacctt cgccatacct 21660
ctgggtgcta cccaagtacc ttattattgt tttcttaaag tggatactta caactccact 21720
gtttataaat ttttggctgt tttacctcct accgtcaggg aaattgtcat caccaagtat 21780
ggtgatgttt atgtcaatgg gtttggatac ttgcatctcg gtttgttgga tgctgtcaca 21840
attaatttca ctggtcatgg cactgacgat gatgtttctg gtttttggac catagcatcg 21900
actaattttg ttgatgcact catcgaagtt caaggaaccg ccattcagcg tattctttat 21960
tgtgatgatc ctgttagcca actcaagtgt tctcaggttg cttttgacct tgacgatggt 22020
ttttacccta tttcttctag aaaccttctg agtcatgaac agccaatttc ttttgttact 22080
ctgccatcat ttaatgatca ttcttttgtt aacattactg tatctgcttc ctttggtggt 22140
catagtggtg ccaaccttat tgcatctgac actactatca atgggtttag ttctttctgt 22200
gttgacacta gacaatttac catttcactg ttttataacg ttacaaacag ttatggttat 22260
gtgtctaaat cacaggacag taattgccct ttcaccttgc aatctgttaa tgattacctg 22320
tcttttagca aattttgtgt ttccaccagc cttttggcta gtgcctgtac catagatctt 22380
tttggttacc ctgagtttgg tagtggtgtt aagtttacgt ccctttactt tcaattcaca 22440
aagggtgagt tgattactgg cacgactaaa ccacttgaag gtgtcacgga cgtttctttt 22500
atgactctgg atgtgtgtac caagtatact atctatggct ttaaaggtga gggtatcatt 22560
acccttacaa attctagctt tttggcaggt gtttattaca catctgtttc tggacagttg 22620
ttagccttta agaatgtcac tagtggtgct gtttattctg ttacgccatg ttctttttca 22680
gagcaggctg catatgttga tgatgatata gtgggtgtta tttctagttt gtctagctcc 22740
acttttaaca gtactaggga gttgcctggt ttcttctacc attctaatga tggctctaat 22800
tgtacagagc ctgtgttggt gtatagtaac ataggtgttt gtaaatctgg cagtattggc 22860
tacgtcccat ctcagtctgg ccaagtcaag attgcaccca cggttactgg gaatattagt 22920
attcccacca actttagtat gagtattagg acagaatatt tacagcttta caacacgcct 22980
gttagtgttg attgtgccac atatgtttgt aatggtaact ctcgttgtaa acaattactc 23040
acccagtaca ctgcagcatg taagaccata gagtcagcat tacractcag cgctaggctt 23100
gagtctgttg aagttaactc tatgcttact atttctgaag aggctctaca gttagctacc 23160
attagttcgt ttaatggtga tggatataat tttactaatg tgctgggtgt ttctgtgtat 23220
gatcctgcaa ggggcagggt ggtacaaaaa aggtctttta ttgaagacct gctttttaat 23280
aaagtggtta ctaatggcct tggtactgtt gatgaagact ataagcgctg ttctaatggt 23340
cgctctgtgg cagatctagt ctgtgcacag tattactctg gtgtcatggt actacctggt 23400
gttgttgacg ctgagaagct tcacatgtat agtgcgtctc tcatcggtgg tatggtgcta 23460
ggaggtttta cttctgcagc ggcattgcct tttagctatg ctgttcaagc tagactcaat 23520
tatcttgctc tacagacgga tgttctacag cggaaccagc aattgcttgc tgagtctttt 23580
aactctgcta ttggtaatat aacttcagcc tttgagagtg ttaaagaggc tattagtcaa 23640
acttccaagg gtttgaacac tgtggctcat gcgcttacta aggttcaaga ggttgttaac 23700
tcgcagggtg cagctttgac tcaacttacc gtacagctgc aacacaactt ccaagccatt 23760
tctagttcta ttgatgacat ttactctcga ctggacattc tttcagccga tgttcaggtt 23820
gaccgtctca tcaccggcag attatcagca cttaatgctt ttgttgctca aaccctcact 23880
aagtatactg aggttcaggc tagcaggaag ttagcacagc aaaaggttaa tgagtgcgtt 23940
aaatcgcaat cccagcgtta tggtttttgt ggtggtgatg gcgagcacat tttctctctg 24000
gtacaggcag cacctcaggg cctgctgttt ttacatacag tacttgtacc gagtgatttt 24060
gtagatgtta ttgccatcgc tggcttatgc gttaacgatg aaattgcctt gactctacgt 24120
gagcctggct tagtcttgtt tacgcatgaa cttcaaaatc atactgcgac ggaatatttt 24180
gtttcatcgc gacgtatgtt tgaacctaga aaacctaccg ttagtgattt tgttcaaatt 24240
gagagttgtg tggtcaccta tgtcaatttg actagagacc aactaccaga tgtaatccca 24300
gattacatcg atgttaacaa aacacttgat gagattttag cttctctgcc caatagaact 24360
ggtccaagtc ttcctttaga tgtttttaat gccacttatc ttaatctcac tggtgaaatt 24420
gcagatttag agcagcgttc agagtctctc cgtaatacta cagaggagct ccaaagtctt 24480
atatataata tcaacaacac actagttgac cttgagtggc tcaaccgagt tgagacatat 24540
atcaagtggc cgtggtgggt ttggttgatt attttcattg ttctcatctt tgttgtgtca 24600
ttactagtgt tctgctgcat ttccacgggt tgttgtggat gctgcggctg ctgctgtgct 24660
tgtttctcag gttgttgtag gggtcctaga cttcaacctt acgaagtttt tgaaaaggtc 24720
cacgtgcagt gatgtttctt ggactttttc aatacacgat tgacacagtt gtcaaagatg 24780
tctcaaagtc tgctaacttg tctttggatg ctgtccaaga gttggagctc aatgtagttc 24840
caattagaca agcttcaaat gtgacgggtt ttcttttcac cagtgttttt atctacttct 24900
ttgcactgtt taaagcgtct tctttgaggc gcaattatat tatgttggca gcgcgttttg 24960
ctgtcattgt tctttattgc ccacttttat attattgtgg tgcattttta gatgcaacta 25020
ttatttgttg cacacttatt caaagtcggt ggcaggcttt gtttagtctg cttttactcc 25080
tggcgctata aaaatgcgct ctttattatt tttaatacta cgacactttc tttcctcaat 25140
ggtaaagcag ctttgacggc aaatccattg tgattttaga aggtggtgac cattacatca 25200
cttttggcaa ctcttttgtt gcttttgtta gtagcatcga cttgtatcta gctatacgtg 25260
ggcggcaaga agctgaccta cagctgttgc gaactgttga gcttcttgat ggcaagaagc 25320
tttatgtctt ttcgcaacat caaattgttg gcattactaa tgctgcattt gactcaattc 25380
aactagacga gtatgctaca attagtgaat gataatggtc tagtagttaa tgttatactt 25440
tggcttttcg tactcttttt cctgcttatt ataagcatta ctttcgtcca attggttaat 25500
ctgtgcttca cttgtcaccg gttgtgtaat agcgcagttt acacacctat agggcgtttg 25560
tatagagttt ataagtctta catgcaaata gaccccctcc ctagtactgt tattgacgta 25620
taaacgaaat atgtctaacg gttctattcc cgttgatgag gtgattcaac accttagaaa 25680
ctggaatttc acatggaata tcatactgac gatactactt gtagtgcttc agtatggcca 25740
ttacaagtac tctgcgttct tgtatggtgt caagatggct attctatgga tactttggcc 25800
tcttgtgtta gcactgtcac tttttgatgc atgggctagc tttcaggtca attgggtctt 25860
ttttgctttc agcatcctta tggcttgcat cactcttatg ctgtggataa tgtactttgt 25920
caatagcatt cggttgtggc gcaggacaca ttcttggtgg tctttcaatc ctgaaacaga 25980
cgcgcttctc actacttctg tgatgggccg acaggtctgc attccagtgc ttggagcacc 26040
aactggtgta acgctaacac tccttagtgg tacattgctt gtagagggct ataaggttgc 26100
tactggcgta caggtaagtc aattacctaa tttcgtcaca gtcgccaagg ccactacaac 26160
aattgtctac ggacgtgttg gtcgttcagt caatgcttca tctggcactg gttgggcttt 26220
ctatgtccgg tccaaacacg gcgactactc agctgtgagt aatccgagtt cggttctcac 26280
agatagtgag aaagtgcttc atttagtcta aacagaaact ttatggcttc tgtcagtttt 26340
caggatcgtg gccgcaaacg ggtgccatta tccctctatg cccctcttag ggttactaat 26400
gacaaacccc tttctaaggt acttgcaaat aatgctgtac ccactaataa aggaaataag 26460
gaccagcaaa ttggatactg gaatgagcaa attcgctggc gcatgcgccg tggtgagcga 26520
attgaacaac cttccaattg gcatttctac tacctcggaa caggacctca cgccgacctc 26580
cgctatagga ctcgtactga gggtgttttc tgggttgcta aagaaggcgc aaagactgaa 26640
cccactaacc tgggtgtcag aaaggcgtct gaaaagccaa ttattccaaa tttctctcaa 26700
cagcttccca gcgtagttga gattgttgaa cctaacacac ctcctacttc acgtgcaaat 26760
tcacgtagca ggagtcgtgg taatggcaac aacaggtcca gatctccaag taacaacaga 26820
ggcaataacc agtcccgcgg taattcacag aatcgtggaa ataaccaggg tcgtggagct 26880
tctcagaaca gaggaggcaa taataataac aataacaagt ctcgtaacca gtccaagaac 26940
agaaaccagt caaatgaccg tggtggtgta acatcacgcg atgatctggt ggctgctgtc 27000
aaggatgccc ttaaatcttt gggtattggc gaaaaccctg acaagcttaa gcaacagcag 27060
aagcccaaac aggaaaggtc tgacagcagc ggcaaaaata cacctaagaa gaacaaatcc 27120
agagccactt cgaaagaacg tgacctcaaa gacatcccag agtggaggag aattcccaag 27180
ggcgaaaata gcgtagcagc ttgcttcgga cccaggggag gcttcaaaaa ttttggagat 27240
gcggaatttg tcgaaaaagg tgttgatgcc tcaggctatg ctcagatcgc cagtttagca 27300
ccaaatgttg cagcattgct ctttggtggt aatgtggctg ttcgtgagct agcggactct 27360
tacgagatta catataatta taaaatgact gtgccaaagt ctgatccaaa tgtagagctt 27420
cttgtttcac aggtggatgc atttaaaact gggaatgcaa aaccccagag aaagaaggaa 27480
aagaagaaca agcgtgaaac cacgcagcag ctgaatgaag aggccatcta cgatgatgtg 27540
ggtgtgccat ctgatgtgac tcatgccaat ttggaatggg acacagctgt tgatggtggt 27600
gacacggccg ttgaaattat caacgagatc ttcgacacag gaaattaaac aatgtttgac 27660
tggcttatcc tggctatgtc ccagggtagt gccattacac tgttattact gagtgttttt 27720
ctagcgactt ggctgctggg ctatggcttt gccctctaac tagcggtctt ggtcttgcac 27780
acaacggtaa gccagtggta atgtcagtgc aagaaggata ttaccatagc actgtcatga 27840
ggggaacgca gtaccttttc atctaaacct ttgcacgagt aatcaaagat ccgcttgacg 27900
agcctatatg gaagagcgtg ccaggtattt gactcaagga ctgttagtaa ctgaagacct 27960
gacggtgttg atatgg 27976
<210> 34
<211> 4161
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDV 1251-125-10 (125-10) passage30 spike protein CDS sequence
<400> 34
atgaagtctt taacctactt ctggttgttc ttaccagtac tttcaacact tagcctacca 60
caagatgtca ccaggtgctc agctaacact aattttaggc ggttcttttc aaaatttaat 120
gttcaggcgc ctgcagttgt tgtactgggc ggttatctac ctattggtga aaaccagggt 180
gtcaattcaa cttggtactg tgctggccaa catccaactg ctagtggcgt tcatggtatc 240
tttgttagcc atattagagg tggtcatggc tttgagattg gcatttcgca agagcctttt 300
gaccctagtg gttaccagct ttatttacat aaggctacta acggtaacac taatgctact 360
gcgcgactgc gcatttgcca gtttcctagc attaaaacat tgggccccac tgctaataat 420
gatgttacaa taggtcgtaa ttgcctattt aacaaagcca tcccagctca tatgagtgaa 480
catagtgttg tcggcataac atgggataat gatcgtgtca ctgtcttttc tgacaagatc 540
tattattttt attttaaaaa tgattggtcc cgtgttgcga caaagtgtta caacagtgga 600
ggttgtgcta tgcaatatgt ttacgaaccc acctattaca tgcttaatgt tactagtgct 660
ggtgaggatg gtatttctta tcaaccctgt acagctaatt gcattggtta tgctgccaat 720
gtatttgcta ctgagcccaa tggccacata ccagaaggtt ttagttttaa taattggttt 780
cttttgtcca atgattccac tttggtgcat ggtaaggtgg tttccaacca accattgttg 840
gtcaattgtc ttttggccat tcctaagatt tatggactag gccaattttt ctcctttaat 900
caaacgatcg atggtgtttg taatggagct gctgtgcagc gtgcaccaga ggctctgagg 960
tttaatatta atgacacctc tgtcattctt gctgaaggct caattgtact tcatactgct 1020
ttaggaacaa atttttcttt tgtttgcagt aattccccaa atcctcactt agccaccttc 1080
gccatacctc tgggtgctac ccaagtacct tattattgtt ttcttaaagt ggatacttac 1140
aactccactg tttataaatt tttggctgtt ttacctccta ccgtcaggga aattgtcatc 1200
accaagtatg gtgatgttta tgtcaatggg tttggatact tgcatctcgg tttgttggat 1260
gctgtcacaa ttaatttcac tggtcatggc actgacgatg atgtttctgg tttttggacc 1320
atagcatcga ctaattttgt tgatgcactc atcgaagttc aaggaaccgc cattcagcgt 1380
attctttatt gtgatgatcc tgttagccaa ctcaagtgtt ctcaggttgc ttttgacctt 1440
gacgatggtt tttaccctat ttcttctaga aaccttctga gtcatgaaca gccaatttct 1500
tttgttactc tgccatcatt taatgatcat tcttttgtta acattactgt atctgcttcc 1560
tttggtggtc atagtggtgc caaccttatt gcatctgaca ctactatcaa tgggtttagt 1620
tctttctgtg ttgacactag acaatttacc atttcactgt tttataacgt tacaaacagt 1680
tatggttatg tgtctaaatc acaggacagt aattgccctt tcaccttgca atctgttaat 1740
gattacctgt cttttagcaa attttgtgtt tccaccagcc ttttggctag tgcctgtacc 1800
atagatcttt ttggttaccc tgagtttggt agtggtgtta agtttacgtc cctttacttt 1860
caattcacaa agggtgagtt gattactggc acgactaaac cacttgaagg tgtcacggac 1920
gtttctttta tgactctgga tgtgtgtacc aagtatacta tctatggctt taaaggtgag 1980
ggtatcatta cccttacaaa ttctagcttt ttggcaggtg tttattacac atctgtttct 2040
ggacagttgt tagcctttaa gaatgtcact agtggtgctg tttattctgt tacgccatgt 2100
tctttttcag agcaggctgc atatgttgat gatgatatag tgggtgttat ttctagtttg 2160
tctagctcca cttttaacag tactagggag ttgcctggtt tcttctacca ttctaatgat 2220
ggctctaatt gtacagagcc tgtgttggtg tatagtaaca taggtgtttg taaatctggc 2280
agtattggct acgtcccatc tcagtctggc caagtcaaga ttgcacccac ggttactggg 2340
aatattagta ttcccaccaa ctttagtatg agtattagga cagaatattt acagctttac 2400
aacacgcctg ttagtgttga ttgtgccaca tatgtttgta atggtaactc tcgttgtaaa 2460
caattactca cccagtacac tgcagcatgt aagaccatag agtcagcatt acractcagc 2520
gctaggcttg agtctgttga agttaactct atgcttacta tttctgaaga ggctctacag 2580
ttagctacca ttagttcgtt taatggtgat ggatataatt ttactaatgt gctgggtgtt 2640
tctgtgtatg atcctgcaag gggcagggtg gtacaaaaaa ggtcttttat tgaagacctg 2700
ctttttaata aagtggttac taatggcctt ggtactgttg atgaagacta taagcgctgt 2760
tctaatggtc gctctgtggc agatctagtc tgtgcacagt attactctgg tgtcatggta 2820
ctacctggtg ttgttgacgc tgagaagctt cacatgtata gtgcgtctct catcggtggt 2880
atggtgctag gaggttttac ttctgcagcg gcattgcctt ttagctatgc tgttcaagct 2940
agactcaatt atcttgctct acagacggat gttctacagc ggaaccagca attgcttgct 3000
gagtctttta actctgctat tggtaatata acttcagcct ttgagagtgt taaagaggct 3060
attagtcaaa cttccaaggg tttgaacact gtggctcatg cgcttactaa ggttcaagag 3120
gttgttaact cgcagggtgc agctttgact caacttaccg tacagctgca acacaacttc 3180
caagccattt ctagttctat tgatgacatt tactctcgac tggacattct ttcagccgat 3240
gttcaggttg accgtctcat caccggcaga ttatcagcac ttaatgcttt tgttgctcaa 3300
accctcacta agtatactga ggttcaggct agcaggaagt tagcacagca aaaggttaat 3360
gagtgcgtta aatcgcaatc ccagcgttat ggtttttgtg gtggtgatgg cgagcacatt 3420
ttctctctgg tacaggcagc acctcagggc ctgctgtttt tacatacagt acttgtaccg 3480
agtgattttg tagatgttat tgccatcgct ggcttatgcg ttaacgatga aattgccttg 3540
actctacgtg agcctggctt agtcttgttt acgcatgaac ttcaaaatca tactgcgacg 3600
gaatattttg tttcatcgcg acgtatgttt gaacctagaa aacctaccgt tagtgatttt 3660
gttcaaattg agagttgtgt ggtcacctat gtcaatttga ctagagacca actaccagat 3720
gtaatcccag attacatcga tgttaacaaa acacttgatg agattttagc ttctctgccc 3780
aatagaactg gtccaagtct tcctttagat gtttttaatg ccacttatct taatctcact 3840
ggtgaaattg cagatttaga gcagcgttca gagtctctcc gtaatactac agaggagctc 3900
caaagtctta tatataatat caacaacaca ctagttgacc ttgagtggct caaccgagtt 3960
gagacatata tcaagtggcc gtggtgggtt tggttgatta ttttcattgt tctcatcttt 4020
gttgtgtcat tactagtgtt ctgctgcatt tccacgggtt gttgtggatg ctgcggctgc 4080
tgctgtgctt gtttctcagg ttgttgtagg ggtcctagac ttcaacctta cgaagttttt 4140
gaaaaggtcc acgtgcagtg a 4161
<210> 35
<211> 1386
<212> PRT
<213> porcine epidemic diarrhea virus
<220>
<221> misc_feature
<222> (838)..(838)
<223> Xaa can be any naturally occurring amino acid
<400> 35
Met Lys Ser Leu Thr Tyr Phe Trp Leu Phe Leu Pro Val Leu Ser Thr
1 5 10 15
Leu Ser Leu Pro Gln Asp Val Thr Arg Cys Ser Ala Asn Thr Asn Phe
20 25 30
Arg Arg Phe Phe Ser Lys Phe Asn Val Gln Ala Pro Ala Val Val Val
35 40 45
Leu Gly Gly Tyr Leu Pro Ile Gly Glu Asn Gln Gly Val Asn Ser Thr
50 55 60
Trp Tyr Cys Ala Gly Gln His Pro Thr Ala Ser Gly Val His Gly Ile
65 70 75 80
Phe Val Ser His Ile Arg Gly Gly His Gly Phe Glu Ile Gly Ile Ser
85 90 95
Gln Glu Pro Phe Asp Pro Ser Gly Tyr Gln Leu Tyr Leu His Lys Ala
100 105 110
Thr Asn Gly Asn Thr Asn Ala Thr Ala Arg Leu Arg Ile Cys Gln Phe
115 120 125
Pro Ser Ile Lys Thr Leu Gly Pro Thr Ala Asn Asn Asp Val Thr Ile
130 135 140
Gly Arg Asn Cys Leu Phe Asn Lys Ala Ile Pro Ala His Met Ser Glu
145 150 155 160
His Ser Val Val Gly Ile Thr Trp Asp Asn Asp Arg Val Thr Val Phe
165 170 175
Ser Asp Lys Ile Tyr Tyr Phe Tyr Phe Lys Asn Asp Trp Ser Arg Val
180 185 190
Ala Thr Lys Cys Tyr Asn Ser Gly Gly Cys Ala Met Gln Tyr Val Tyr
195 200 205
Glu Pro Thr Tyr Tyr Met Leu Asn Val Thr Ser Ala Gly Glu Asp Gly
210 215 220
Ile Ser Tyr Gln Pro Cys Thr Ala Asn Cys Ile Gly Tyr Ala Ala Asn
225 230 235 240
Val Phe Ala Thr Glu Pro Asn Gly His Ile Pro Glu Gly Phe Ser Phe
245 250 255
Asn Asn Trp Phe Leu Leu Ser Asn Asp Ser Thr Leu Val His Gly Lys
260 265 270
Val Val Ser Asn Gln Pro Leu Leu Val Asn Cys Leu Leu Ala Ile Pro
275 280 285
Lys Ile Tyr Gly Leu Gly Gln Phe Phe Ser Phe Asn Gln Thr Ile Asp
290 295 300
Gly Val Cys Asn Gly Ala Ala Val Gln Arg Ala Pro Glu Ala Leu Arg
305 310 315 320
Phe Asn Ile Asn Asp Thr Ser Val Ile Leu Ala Glu Gly Ser Ile Val
325 330 335
Leu His Thr Ala Leu Gly Thr Asn Phe Ser Phe Val Cys Ser Asn Ser
340 345 350
Pro Asn Pro His Leu Ala Thr Phe Ala Ile Pro Leu Gly Ala Thr Gln
355 360 365
Val Pro Tyr Tyr Cys Phe Leu Lys Val Asp Thr Tyr Asn Ser Thr Val
370 375 380
Tyr Lys Phe Leu Ala Val Leu Pro Pro Thr Val Arg Glu Ile Val Ile
385 390 395 400
Thr Lys Tyr Gly Asp Val Tyr Val Asn Gly Phe Gly Tyr Leu His Leu
405 410 415
Gly Leu Leu Asp Ala Val Thr Ile Asn Phe Thr Gly His Gly Thr Asp
420 425 430
Asp Asp Val Ser Gly Phe Trp Thr Ile Ala Ser Thr Asn Phe Val Asp
435 440 445
Ala Leu Ile Glu Val Gln Gly Thr Ala Ile Gln Arg Ile Leu Tyr Cys
450 455 460
Asp Asp Pro Val Ser Gln Leu Lys Cys Ser Gln Val Ala Phe Asp Leu
465 470 475 480
Asp Asp Gly Phe Tyr Pro Ile Ser Ser Arg Asn Leu Leu Ser His Glu
485 490 495
Gln Pro Ile Ser Phe Val Thr Leu Pro Ser Phe Asn Asp His Ser Phe
500 505 510
Val Asn Ile Thr Val Ser Ala Ser Phe Gly Gly His Ser Gly Ala Asn
515 520 525
Leu Ile Ala Ser Asp Thr Thr Ile Asn Gly Phe Ser Ser Phe Cys Val
530 535 540
Asp Thr Arg Gln Phe Thr Ile Ser Leu Phe Tyr Asn Val Thr Asn Ser
545 550 555 560
Tyr Gly Tyr Val Ser Lys Ser Gln Asp Ser Asn Cys Pro Phe Thr Leu
565 570 575
Gln Ser Val Asn Asp Tyr Leu Ser Phe Ser Lys Phe Cys Val Ser Thr
580 585 590
Ser Leu Leu Ala Ser Ala Cys Thr Ile Asp Leu Phe Gly Tyr Pro Glu
595 600 605
Phe Gly Ser Gly Val Lys Phe Thr Ser Leu Tyr Phe Gln Phe Thr Lys
610 615 620
Gly Glu Leu Ile Thr Gly Thr Thr Lys Pro Leu Glu Gly Val Thr Asp
625 630 635 640
Val Ser Phe Met Thr Leu Asp Val Cys Thr Lys Tyr Thr Ile Tyr Gly
645 650 655
Phe Lys Gly Glu Gly Ile Ile Thr Leu Thr Asn Ser Ser Phe Leu Ala
660 665 670
Gly Val Tyr Tyr Thr Ser Val Ser Gly Gln Leu Leu Ala Phe Lys Asn
675 680 685
Val Thr Ser Gly Ala Val Tyr Ser Val Thr Pro Cys Ser Phe Ser Glu
690 695 700
Gln Ala Ala Tyr Val Asp Asp Asp Ile Val Gly Val Ile Ser Ser Leu
705 710 715 720
Ser Ser Ser Thr Phe Asn Ser Thr Arg Glu Leu Pro Gly Phe Phe Tyr
725 730 735
His Ser Asn Asp Gly Ser Asn Cys Thr Glu Pro Val Leu Val Tyr Ser
740 745 750
Asn Ile Gly Val Cys Lys Ser Gly Ser Ile Gly Tyr Val Pro Ser Gln
755 760 765
Ser Gly Gln Val Lys Ile Ala Pro Thr Val Thr Gly Asn Ile Ser Ile
770 775 780
Pro Thr Asn Phe Ser Met Ser Ile Arg Thr Glu Tyr Leu Gln Leu Tyr
785 790 795 800
Asn Thr Pro Val Ser Val Asp Cys Ala Thr Tyr Val Cys Asn Gly Asn
805 810 815
Ser Arg Cys Lys Gln Leu Leu Thr Gln Tyr Thr Ala Ala Cys Lys Thr
820 825 830
Ile Glu Ser Ala Leu Xaa Leu Ser Ala Arg Leu Glu Ser Val Glu Val
835 840 845
Asn Ser Met Leu Thr Ile Ser Glu Glu Ala Leu Gln Leu Ala Thr Ile
850 855 860
Ser Ser Phe Asn Gly Asp Gly Tyr Asn Phe Thr Asn Val Leu Gly Val
865 870 875 880
Ser Val Tyr Asp Pro Ala Arg Gly Arg Val Val Gln Lys Arg Ser Phe
885 890 895
Ile Glu Asp Leu Leu Phe Asn Lys Val Val Thr Asn Gly Leu Gly Thr
900 905 910
Val Asp Glu Asp Tyr Lys Arg Cys Ser Asn Gly Arg Ser Val Ala Asp
915 920 925
Leu Val Cys Ala Gln Tyr Tyr Ser Gly Val Met Val Leu Pro Gly Val
930 935 940
Val Asp Ala Glu Lys Leu His Met Tyr Ser Ala Ser Leu Ile Gly Gly
945 950 955 960
Met Val Leu Gly Gly Phe Thr Ser Ala Ala Ala Leu Pro Phe Ser Tyr
965 970 975
Ala Val Gln Ala Arg Leu Asn Tyr Leu Ala Leu Gln Thr Asp Val Leu
980 985 990
Gln Arg Asn Gln Gln Leu Leu Ala Glu Ser Phe Asn Ser Ala Ile Gly
995 1000 1005
Asn Ile Thr Ser Ala Phe Glu Ser Val Lys Glu Ala Ile Ser Gln
1010 1015 1020
Thr Ser Lys Gly Leu Asn Thr Val Ala His Ala Leu Thr Lys Val
1025 1030 1035
Gln Glu Val Val Asn Ser Gln Gly Ala Ala Leu Thr Gln Leu Thr
1040 1045 1050
Val Gln Leu Gln His Asn Phe Gln Ala Ile Ser Ser Ser Ile Asp
1055 1060 1065
Asp Ile Tyr Ser Arg Leu Asp Ile Leu Ser Ala Asp Val Gln Val
1070 1075 1080
Asp Arg Leu Ile Thr Gly Arg Leu Ser Ala Leu Asn Ala Phe Val
1085 1090 1095
Ala Gln Thr Leu Thr Lys Tyr Thr Glu Val Gln Ala Ser Arg Lys
1100 1105 1110
Leu Ala Gln Gln Lys Val Asn Glu Cys Val Lys Ser Gln Ser Gln
1115 1120 1125
Arg Tyr Gly Phe Cys Gly Gly Asp Gly Glu His Ile Phe Ser Leu
1130 1135 1140
Val Gln Ala Ala Pro Gln Gly Leu Leu Phe Leu His Thr Val Leu
1145 1150 1155
Val Pro Ser Asp Phe Val Asp Val Ile Ala Ile Ala Gly Leu Cys
1160 1165 1170
Val Asn Asp Glu Ile Ala Leu Thr Leu Arg Glu Pro Gly Leu Val
1175 1180 1185
Leu Phe Thr His Glu Leu Gln Asn His Thr Ala Thr Glu Tyr Phe
1190 1195 1200
Val Ser Ser Arg Arg Met Phe Glu Pro Arg Lys Pro Thr Val Ser
1205 1210 1215
Asp Phe Val Gln Ile Glu Ser Cys Val Val Thr Tyr Val Asn Leu
1220 1225 1230
Thr Arg Asp Gln Leu Pro Asp Val Ile Pro Asp Tyr Ile Asp Val
1235 1240 1245
Asn Lys Thr Leu Asp Glu Ile Leu Ala Ser Leu Pro Asn Arg Thr
1250 1255 1260
Gly Pro Ser Leu Pro Leu Asp Val Phe Asn Ala Thr Tyr Leu Asn
1265 1270 1275
Leu Thr Gly Glu Ile Ala Asp Leu Glu Gln Arg Ser Glu Ser Leu
1280 1285 1290
Arg Asn Thr Thr Glu Glu Leu Gln Ser Leu Ile Tyr Asn Ile Asn
1295 1300 1305
Asn Thr Leu Val Asp Leu Glu Trp Leu Asn Arg Val Glu Thr Tyr
1310 1315 1320
Ile Lys Trp Pro Trp Trp Val Trp Leu Ile Ile Phe Ile Val Leu
1325 1330 1335
Ile Phe Val Val Ser Leu Leu Val Phe Cys Cys Ile Ser Thr Gly
1340 1345 1350
Cys Cys Gly Cys Cys Gly Cys Cys Cys Ala Cys Phe Ser Gly Cys
1355 1360 1365
Cys Arg Gly Pro Arg Leu Gln Pro Tyr Glu Val Phe Glu Lys Val
1370 1375 1380
His Val Gln
1385
<210> 36
<211> 27976
<212> RNA
<213> Artificial Sequence
<220>
<223> PEDV 1251-125-10 (125-10) passage30 spike protein genomic RNA
<400> 36
aauucaacua aacgaaauuu uguccuuccg gccgcauguc caugcugcug gaagcugacg 60
uggaauuuca uuagguuugc uuaaguagcc aucgcaagug cugugcuguc cucuaguucc 120
ugguuggcgu uccgucgccu ucuacauacu agacaaacag ccuuccuccg guuccgucug 180
gggguugugu ggauaacuag uuccgucuag uuugaaacca guaacugucg gcuauggcua 240
gcaaccaugu uacauuggcu uuugccaaug augcagaaau uucagcuuuu ggcuuuugca 300
cugcuaguga agccgucuca uacuauucug aggccgccgc uaguggauuu augcaaugcc 360
guuucguguc cuucgaucuc gcugacacug uugagggauu gcuucccgaa gacuauguca 420
uggugguggu cggcacuacc aagcuuagug cguaugugga cacuuuuggu agccgcccca 480
aaaacauuug ugguuggcug uuauuuucua acuguaauua cuuccucgaa gaguuagagc 540
uuacuuuugg ucgucguggu gguaacaucg ugccaguuga ccaauacaug uguggcgcug 600
acgguaaacc uguucuucag gaauccgaau gggaguauac agauuucuuu gcugacuccg 660
aagacgguca acucaacauu gcugguauca cuuaugugaa ggccuggauu guagagcgau 720
cggaugucuc uuaugcgagu cagaauuuaa caucuauuaa gucuauuacu uacuguucaa 780
ccuaugagca uacuuuuccu gaugguacug ccaugaaggu ugcacguacu ccaaagauua 840
agaagacugu ugucuugucu gagccacuug cuacuaucua cagggaaauu gguucuccuu 900
uuguggauaa ugggagcgau gcucguucua ucauuaagag accaguguuc cuccacgcuu 960
uuguuaagug uaaguguggu aguuaucauu ggacuguugg ugauuggacu uccuaugucu 1020
ccacuugcug uggcuuuaag uguaagccag uccuuguggc uucaugcucu gcuacgccug 1080
guucuguugu gguuacgcgc gcuggugcug gcacuggugu uaaguauuac aacaacaugu 1140
uccugcgcca uguggcagac auugaugggu uggcauucug gcgaauucuc aaggugcagu 1200
ccaaagacga ccucgcuugc ucugguaaau uccuugaaca ccaugaggaa gguuucacag 1260
auccuugcua cuuuuugaau gacucgagca uugcuacuaa gcucaaguuu gacauccuua 1320
guggcaaguu uucugaugaa gucaaacaag cuaucuuugc uggucauguu guuguuggca 1380
gcgcgmucgu ugacauuguu gacgaugcac ugggacagcc uugguuuaua cguaagcuug 1440
gugaccuugc aagugcagcu ugggagcagc uuaaggcugu cguuagaggc cuuaaccucc 1500
ugucugauga ggucgugcuc uuuggcaaaa gacuuagcug ugccacucuu aguaucguua 1560
acgguguuuu ugaguucauc gccgaagugc cugagaaguu ggcugcggcu guuacaguuu 1620
uugucaacuu cuugaaugag cuuuuugagu cugccuguga cugcuuaaag gucggaggua 1680
aaaccuuuaa caagguuggc ucuuauguuc uuuuugacaa cgcauugguu aagcuuguca 1740
aggcaaaagu ucgcggccca cgacaggcag guguuuguga aguucguuac acaagccuug 1800
uuauugggag uacuaccaag gugguuucca agcgcguuga aaaugccaau gugaaucucg 1860
ucgucguuga cgaggaugug acccucaaca ccacuggucg uacaguuguu guugacggac 1920
uugcauucuu cgagagugac ggguuuuaca gacaucuugc ugaugcugac guugucauug 1980
aacauccugu uuauaagucu gcuugugagc ucaagccagu uuuugagugu gacccaauac 2040
cugauuuucc uaugccugug gccgcuagug uugcagagcu uugugugcaa acugaucugu 2100
ugcuuaaaaa uuacaacacu ccuuauaaaa cuuacagcug cguugugaga ggugauaagu 2160
guuguaucac uugcaccuua cauuucacag caccaaguua uauggaggcu gcugcuaauu 2220
uuguagaccu cuguaccaag aacauuggua cugcugguuu ucaugaguuu uacauuacgg 2280
cccaugaaca acaggaucug caaggguucg uaaccacuug uugcacgaug ucagguuuug 2340
aguguuuuau gccuauaauc ccacaguguc cagcagugcu ugaagagauu gaugguggua 2400
gcaucuggcg gucuuuuauc acuggucuua auacaaugug ggauuuuugc aagcaucuua 2460
aagucagcuu uggacuagau ggcauuguug ucacuguagc acgcaaauuu aaacgacuug 2520
gugcucucuu ggcagaaaug uauaacacuu accuuucaac ugugguggaa aacuugguac 2580
uggccggugu uagcuucaag uauuaugcca ccaguguccc aaaaauuguu uugggcuguu 2640
guuuucacag uguuaaaagu guucuugcaa gugccuucca gauuccuguc caggcaggcg 2700
uugagaaguu uaaagucuuc cuuaacugug uucacccugu uguaccacgu gucauugaaa 2760
cuucuuuugu ggaauuagaa gagacgacau uuaaaccacc agcacucaau gguaguauug 2820
cuauuguuga uggcuuugcu uucuauuaug auggaacacu auacuauccc accgauggua 2880
auagcguugu uccuaucugc uuuaagaaga aagguggugg ugaugucaaa uucucugaug 2940
aagucucugu uaaaaccauu gacccaguuu auaaggucuc ccuugaauuu gaguucgagu 3000
cugagacuau uauggcugug cuuaauaagg cuguugguaa uuguaucaag guuacaggug 3060
guugggacga uguuguugag uauaucaaug uugccauuga gguucuuaaa gaucacaucg 3120
augugccuaa guacuacauc uaugaugagg aagguggcac cgauccuaau cugcccguaa 3180
ugguuucuca guggccguug aaugaugaca cgaucucaca ggaucugcuu gauguugaag 3240
uuguuacuga ugcgccaguu gauuucgagg gugaugaagu agacuccucu gacccugwua 3300
agguggcaga cguggcuaac ucugagccug aggaugacgg ucuuaaugua gcuccugaaa 3360
caaauguaga gucugaaguu gaggaaguug ccgcaaccuu guccuuuaaa gauacaccuu 3420
ccacaguuac uaaggauccu uuugcuuuug acuuugcaag cuauggagga cuuaagguuu 3480
uaagacaauc ucauaacaac ugcuggguua cuucuaccuu ggugcagcua caauugcuug 3540
gcaucguuga ugacccugca auggagcuuu uuagugcugg uagaguuggu ccaaugguuc 3600
gcaaaugcua ugagucacaa aaggcuaucu ugggaucuuu gggugaugug ucggcuugcc 3660
uagagucucu gacuaaggac cuacacacac uuaagauuac cuguucugua gucugugguu 3720
gugguacugg ugaacguauc uaugaugguu gugcuuuucg uaugacgcca acuuuggaac 3780
cguucccaua uggugcuugu gcucagugug cucaaguuuu gaugcacacu uuuaaaagua 3840
uuguuggcac cggcaucuuu ugucgagaua cuacugcucu cuccuuggau ucuuugguug 3900
uaaaaccucu uugugcggcu gcuuuuauag gcaaggauag uggucauuau gucacuaacu 3960
uuuaugaugc ugcuauggcu auugaugguu auggucguca ucagauaaag uaugacacac 4020
ugaacacuau uuguguuaaa gacguuaauu ggacagcacc uuuuguccca gacguugagc 4080
cuguauugga gccuguuguc aaaccuuucu auucuuauaa gaauguugau uuuuaccaag 4140
gagauuuuag ugaccuuguu aaacuuccau gugauuuugu uguuaaugcu gcaaaugaga 4200
auuugucuca cgguggcggc auagcaaagg ccauugaugu uuauaccaag ggcauguugc 4260
agaagugcuc gaaugauuac auuaaagcac acggucccau uaaaguugga cgugguguca 4320
uguuggaggc auuaggucuu aaggucuuua auguuguugg uccacguaag gguaagcaug 4380
caccugagcu ucuuguuaag gcuuauaagu ccguuuuugc uaauucaggu guugcucuua 4440
caccuuugau uaguguugga auuuuuagug uuccuuugga agaaucuuua ucugcuuuuc 4500
uugcaugugu uggugaucgc cacuguaagu gcuuuuguua uagugacaaa gagcgcgagg 4560
cgaucauuaa uuacauggau ggcuugguag augcuauuuu caaagaugca cuuguugaua 4620
cuacuccugu ccaggaagau guucaacaag uuucacaaaa accaguuuug ccuaauuuug 4680
aaccuuucag gauugaaggu gcucaugcuu ucuaugagug caacccugaa gguuugaugu 4740
cauuaggugc ugacaagcug guguuguuua caaauuccaa uuuggauuuu uguagcguug 4800
guaagugucu uaacaaugug acuggcggug cauugcuuga agccauaaau guauuuaaaa 4860
agaguaacaa aacagugccu gcuggcaacu guguuacuuu ugagugugca gauaugauuu 4920
cuauuacuau gguaguauug ccaucugacg gugaugcuaa uuaugacaaa aauuaugcac 4980
gcgccgucgu caagguaucu aagcuuaaag gcaaguuauu gcuugcuguu ggugaugcca 5040
uguuguauuc caaguugucc caccucagcg uguuagguuu cguauccaca ccugaugaug 5100
uggagcguuu cuacgcaaau aagagugugg uuauuaaagu uacugaggau acacguagug 5160
uuaagacugu uaaaguagaa uccacuguua cuuauggaca acaaauugga ccuugucuug 5220
uuaaugacac cguugucaca gacaacaaac cuguuguugc ugauguugua gcuaagguug 5280
uaccaagugc uaauugggau ucacauuaug guuuugauaa ggcuggugag uuccacaugc 5340
uagaccauac uggguuugcc uuuccuagug aaguuguuaa cgguaggcgu gugcuuaaaa 5400
ccacagauaa uaacuguugg guuaauguua cauguuuaca auuacaguuu gcuagauuua 5460
gguucaaguc agcaggucua caggcuaugu gggaguccua uuguacuggu gauguugcua 5520
uguuugugca uugguuguac uggcuuacug guguugacaa aggucagccu agugauucag 5580
aaaaugcacu uaacauguug ucuaaguaca uuguuccugc ugguucuguc acuauugaac 5640
gugucacgca ugacgguugu uguuguagua agcguguugu cacugcacca guugugaaug 5700
cuagcguguu gaagcuuggc gucgaggaug gucuuugucc acauggucuu aacuacauug 5760
acaaaguugu uguaguuaaa gguacuacaa uuguugucaa uguuggaaaa ccuguagugg 5820
caccaucgca ccucuuucuu aaggguguuu ccuacacaac auuccuagau aaugguaacg 5880
guguugccgg ccauuauacu guuuuugauc augacacugg uauggugcau gauggagaug 5940
uuuuuguacc aggugaucuc aaugugucuc cuguuacaaa uguugucguc ucagagcaga 6000
cggcuguugu gauuaaagac ccugugaaga aaguagaguu agacgcuaca aagcuguuag 6060
acacuaugaa uuaugcaucg gaaagauucu uuuccuuugg ugauuuuaug ucacguaauu 6120
uaauuacagu guuuuuguac auccuuagua uuuugggucu cuguuuuagg gccuuucgua 6180
agagggaugu uaaaguucua gcugguguac cccaacguac ugguauuaua uugcguaaaa 6240
gugugcgcua uaaugcaaag gcuuugggug ucuucuucaa gcuaaaacuu uauugguuca 6300
aaguucuugg uaaguuuagu uuggguauuu augcauugua ugcauuacua uucaugacaa 6360
uacgcuuuac accuauaggu ggcccuguuu gugaugaugu uguugcuggu uaugcuaauu 6420
cuaguuuuga caagaaugag uauugcaaca guguuauuug uaaggucugu cucuaugggu 6480
accaggaacu uucggacuuc ucucacacac agguaguaug gcaacaccuu agagacccau 6540
uaauugguaa ugugaugccu uucuuuuauu uggcauuucu ggcaauuuuu ggggguguuu 6600
auguaaaggc uauuacucuc uauuuuauuu uccaguaucu uaacauacuu gguguguuuu 6660
ugggccuaca acaguccauu ugguuuuugc agcuugugcc uuuugauguc uuuggugacg 6720
agaucgucgu cuuuuucauc guuacacgcg uauugauguu ccuuaagcau guuuuccuug 6780
gcugcgauaa ggcaucuugu guggcuugcu cuaagagugc ucgccuuaag cgcguuccug 6840
uccagacuau uuuucagggu acuagcaaau ccuucuacgu acaugccaau ggugguucua 6900
aguucuguaa gaagcacaau uucuuuuguu uaaauuguga uucuuauggu ccaggcugca 6960
cuuuuauuaa ugacgucauu gcaacugaag uugguaaugu ugucaaacuu aaugugcaac 7020
cgacaggucc ugccacuauu cuuauugaca agguugaauu caguaauggu uuuuacuauc 7080
uuuauagugg ugacacauuu uggaaguaca acuuugacau aacagauaac aaauacacuu 7140
gcaaagaguc acuuaaaaau uguagcauaa ucacagacuu uauuguuuuu aacaauaaug 7200
guuccaaugu aaaucagguu aagaaugcau guguguauuu uucacagaug cuuuguaaac 7260
cuguuaaguu aguggacuca gcguuguugg ccaguuuguc uguugauuuu ggugcaagcu 7320
uacauagugc uuuuguuagu guguugucga auaguuuugg caaagaccug ucaaguugua 7380
augacaugca ggauugcaag agcacauugg guuuugauga uguaccauug gauaccuuua 7440
augcugcugu ugcugaggcu caucguuacg auguccucuu gacugacaug ucguucaaca 7500
auuuuaccac caguuaugca aaaccagagg aaaaacuucc cguccaugac auugccacgu 7560
guaugcgugu aggugccaag auuguuaauc auaacguucu ugucaaggau aguauaccug 7620
ugguguggcu uguacgugau uucauugccc uuucugaaga aacuaggaag uacauuauuc 7680
guacgacuaa aguuaagggu auaaccuuca uguugaccuu uaaugauugu cguaugcaua 7740
cuaccauacc uacuguuugc auugcaaaua agaagggugc aggucuuccu aguuuuucaa 7800
agguuaagaa auucuucugg uuuuuguguc uguucauagu ugcuguuuuc uuugcacuaa 7860
gcuuuuuuga uuuuaguacu cagguuagca gugauaguga uuaugacuuc aaguauauug 7920
agaguggcca guugaagacu uuugacaauc cacuuaguug ugugcauaau gucuuuagua 7980
acuucgacca guggcaugau gccaaguuug guuucacccc cgucaacaau ccuaguuguc 8040
cuauagucgu ugguguauca gacgaagcgc gcacuguucc agguauccca gcagguguuu 8100
auuuagcugg uaaaacacuu guuuuugcua uuaacaccau uuuugguaca ucugguuugu 8160
gcuuugaugc uaguggcguu gcugauaagg gcgcuugcau uuuuaauucg gcuugcacca 8220
cauuaucugg uuugggugga acugcugucu acuguuauaa gaauggucua guugaaggug 8280
cuaaacuuua uagugaguug gcaccucaua gcuacuauaa aaugguagau gguaaugcug 8340
ugucuuuacc ugaaauuauc ucacgcggcu uuggcauccg uacuauccgu acaaaggcua 8400
ugaccuacug ucgcguuggc cagugugugc aaucugcaga agguguuugu uuuggcgccg 8460
auagauucuu ugucuauaau gcagaaucug guucugacuu uguuuguggc acagggcucu 8520
uuacauuguu gaugaacguu auuaguguuu uuuccaagac aguaccagua acuguguugu 8580
cuggucaaau acuuuuuaau ugcauuauug cuuuugcugc uguugcggug uguuucuuau 8640
uuacaaaguu uaagcgcaug uucggugaua ugucuguugg cguuuucacu gucggugcuu 8700
guacuuuguu gaacaauguu uccuacauug uaacacagaa cacacuuggc auguugggcu 8760
augcaacuuu guacuuuuug ugcacuaaag guguuagaua uauguggauu uggcauuugg 8820
gauuuuugau cucauauaua cuuauugcac cauggugggu uuugaugguu uaugccuuuu 8880
cagccauuuu ugaguuuaug ccuaaccuuu uuaagcuuaa gguuucaaca caacuuuuug 8940
agggugacaa guucguaggc ucuuuugaaa augcugcagc agguacauuu gugcuugaua 9000
ugcaugccua ugagagacuu gccaacucua ucucaacuga aaaacugcgu caguaugcua 9060
guacuuacaa uaaguacaag uauuauucag gcagugcuuc agaggcugau uacaggcuug 9120
cuuguuuugc ccauuuggcc aaggcuauga uggauuaugc uucuaaucac aacgacacgu 9180
uauacacacc acccacugug aguuacaauu caacucuaca ggcuggcuug cguaagaugg 9240
cacaaccauc ugguguuguu gagaagugca uaguucgugu uugcuauggu aauauggcuc 9300
uuaauggccu auggcuuggu gauacuguua ucugcccacg ccauguuaua gcgucuagua 9360
cuacuagcac uauagauuau gacuaugccc uuucuguuuu acgccucyac aacuucucca 9420
uuucaucugg uaauguuuuc cuagguguug uggguguaac caugcgaggu gcuuuguugc 9480
agauaaaggu uaaucaaaac aauguccaca cgccuaagua caccuaucgc acaguuagac 9540
cgggugaauc uuuuaauauc uuggcgugcu augaugguuc ugcagcuggu guuuacggcg 9600
uuaacaugcg cucuaauuac acuauuagag gcucguucau uaauggcgcu ugugguucac 9660
cugguuauaa cauuaacaau gguaccguug aguuuugcua uuuacaccag cuugaacuug 9720
guucaggcug ucauguuggu agcgacuuag augguguuau guaugguggu uaugaggacc 9780
aaccuacuuu gcaaguugaa ggcgcuagua gucuguuuac agagaaugug uuggcauuuc 9840
uuuaugcagc acucauuaau gguucuaccu gguggcuuag uucuucuagg auugcuguag 9900
acagguuuaa ugagugggcu guucauaaug guaugacaac aguaguuaau acugauugcu 9960
uuucuauucu ugcugcuaag acugguguug auguacaacg uuuguuggcc ucaauccagu 10020
cucugcauaa gaauuuuggu ggaaagcaaa uucuuggcua uaccucguug acagaugagu 10080
uuacuacagg ugaaguuaua cgucaaaugu auggcguuwa ucuucagagu gguuauguuu 10140
cacgcgccug uagaaauguc uugcugguug guucuuuucu gacuuucuuu uggucagaau 10200
uaguuuccua cacuaaguuc uuuuggguaa auccugguua ugucacaccu auguuugcgu 10260
guuugucauu gcuguccuca cuuuugaugu ucacacucaa gcauaagaca uuguuuuucc 10320
aggucuuucu aauaccugcu cugauuguua caucuugcau uaauuuggca uuugauguug 10380
aagucuacaa cuauuuggca gagcauuuug auuaccaugu uucucucaug gguuuuaaug 10440
cacaaggucu uguuaacauc uuugucugcu uuguuguuac cauuuuacac ggcacauaca 10500
cauggcgcuu uuuuaacaca ccugugaguu cugucacuua ugugguagcu uugcugacug 10560
cggcauauaa cuauuuuuac gcuagugaca uucuuaguug ugcuaugaca cuauuugcua 10620
gugugacugg caacugguuc guuggugcug uuuguuauaa agcugcuguu uauauggccu 10680
ugagauuucc uacuuuugug gcuauuuuug gugauauuaa gaguguuaug uucuguuacc 10740
uuguguuggg uuauuuuacc uguugcuucu acgguauucu cuacugguuc aacagguuuu 10800
uuaagguuag uguagguguc uaugacuaua cuguuagugc ugcugaguuu aaguauaugg 10860
uugcuaacgg ccuacgugca ccaacuggaa cacuugauuc acuacuucug ucugccaaau 10920
ugauugguau ugguggugag cggaauauua agauuucuuc cguucagucu aaacugacug 10980
auauuaagug uaguaacguu gugcuuuuag gcugucucuc uagcaugaau gucucagcaa 11040
auucaacaga augggccuau uguguugacu ugcauaacaa gaucaacuug uguaaugacc 11100
cagaaaaagc gcaggaaaug cuacuugcuu uguuggcauu uuuccuuagu aagaauagug 11160
cuuuugguuu agaugacuua uuggaauccu auuuuaauga caauaguaug uugcagagug 11220
uugcaucuac uuaugucggu uugccuucuu augucauuua ugaaaaugca cgccaacagu 11280
augaagaugc uguuaauaau gguucuccac cucaguuggu uaagcaauug cgccaugcca 11340
ugaauguagc aaagagcgaa uuugaccgug aggcuucuac ucagcguaag cuugauagaa 11400
uggcggaaca ggcugcagca cagauguaca aagaggcacg agcaguuaau aggaagucca 11460
aaguuguaag ugcuaugcau ucacugcuuu uugguauguu gagacguuug gacaugucuu 11520
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gguaaagcag cuuugacggc aaauccauug ugauuuuaga agguggugac cauuacauca 25200
cuuuuggcaa cucuuuuguu gcuuuuguua guagcaucga cuuguaucua gcuauacgug 25260
ggcggcaaga agcugaccua cagcuguugc gaacuguuga gcuucuugau ggcaagaagc 25320
uuuaugucuu uucgcaacau caaauuguug gcauuacuaa ugcugcauuu gacucaauuc 25380
aacuagacga guaugcuaca auuagugaau gauaaugguc uaguaguuaa uguuauacuu 25440
uggcuuuucg uacucuuuuu ccugcuuauu auaagcauua cuuucgucca auugguuaau 25500
cugugcuuca cuugucaccg guuguguaau agcgcaguuu acacaccuau agggcguuug 25560
uauagaguuu auaagucuua caugcaaaua gacccccucc cuaguacugu uauugacgua 25620
uaaacgaaau augucuaacg guucuauucc cguugaugag gugauucaac accuuagaaa 25680
cuggaauuuc acauggaaua ucauacugac gauacuacuu guagugcuuc aguauggcca 25740
uuacaaguac ucugcguucu uguauggugu caagauggcu auucuaugga uacuuuggcc 25800
ucuuguguua gcacugucac uuuuugaugc augggcuagc uuucagguca auugggucuu 25860
uuuugcuuuc agcauccuua uggcuugcau cacucuuaug cuguggauaa uguacuuugu 25920
caauagcauu cgguuguggc gcaggacaca uucuuggugg ucuuucaauc cugaaacaga 25980
cgcgcuucuc acuacuucug ugaugggccg acaggucugc auuccagugc uuggagcacc 26040
aacuggugua acgcuaacac uccuuagugg uacauugcuu guagagggcu auaagguugc 26100
uacuggcgua cagguaaguc aauuaccuaa uuucgucaca gucgccaagg ccacuacaac 26160
aauugucuac ggacguguug gucguucagu caaugcuuca ucuggcacug guugggcuuu 26220
cuauguccgg uccaaacacg gcgacuacuc agcugugagu aauccgaguu cgguucucac 26280
agauagugag aaagugcuuc auuuagucua aacagaaacu uuauggcuuc ugucaguuuu 26340
caggaucgug gccgcaaacg ggugccauua ucccucuaug ccccucuuag gguuacuaau 26400
gacaaacccc uuucuaaggu acuugcaaau aaugcuguac ccacuaauaa aggaaauaag 26460
gaccagcaaa uuggauacug gaaugagcaa auucgcuggc gcaugcgccg uggugagcga 26520
auugaacaac cuuccaauug gcauuucuac uaccucggaa caggaccuca cgccgaccuc 26580
cgcuauagga cucguacuga ggguguuuuc uggguugcua aagaaggcgc aaagacugaa 26640
cccacuaacc ugggugucag aaaggcgucu gaaaagccaa uuauuccaaa uuucucucaa 26700
cagcuuccca gcguaguuga gauuguugaa ccuaacacac cuccuacuuc acgugcaaau 26760
ucacguagca ggagucgugg uaauggcaac aacaggucca gaucuccaag uaacaacaga 26820
ggcaauaacc agucccgcgg uaauucacag aaucguggaa auaaccaggg ucguggagcu 26880
ucucagaaca gaggaggcaa uaauaauaac aauaacaagu cucguaacca guccaagaac 26940
agaaaccagu caaaugaccg ugguggugua acaucacgcg augaucuggu ggcugcuguc 27000
aaggaugccc uuaaaucuuu ggguauuggc gaaaacccug acaagcuuaa gcaacagcag 27060
aagcccaaac aggaaagguc ugacagcagc ggcaaaaaua caccuaagaa gaacaaaucc 27120
agagccacuu cgaaagaacg ugaccucaaa gacaucccag aguggaggag aauucccaag 27180
ggcgaaaaua gcguagcagc uugcuucgga cccaggggag gcuucaaaaa uuuuggagau 27240
gcggaauuug ucgaaaaagg uguugaugcc ucaggcuaug cucagaucgc caguuuagca 27300
ccaaauguug cagcauugcu cuuugguggu aauguggcug uucgugagcu agcggacucu 27360
uacgagauua cauauaauua uaaaaugacu gugccaaagu cugauccaaa uguagagcuu 27420
cuuguuucac agguggaugc auuuaaaacu gggaaugcaa aaccccagag aaagaaggaa 27480
aagaagaaca agcgugaaac cacgcagcag cugaaugaag aggccaucua cgaugaugug 27540
ggugugccau cugaugugac ucaugccaau uuggaauggg acacagcugu ugaugguggu 27600
gacacggccg uugaaauuau caacgagauc uucgacacag gaaauuaaac aauguuugac 27660
uggcuuaucc uggcuauguc ccaggguagu gccauuacac uguuauuacu gaguguuuuu 27720
cuagcgacuu ggcugcuggg cuauggcuuu gcccucuaac uagcggucuu ggucuugcac 27780
acaacgguaa gccaguggua augucagugc aagaaggaua uuaccauagc acugucauga 27840
ggggaacgca guaccuuuuc aucuaaaccu uugcacgagu aaucaaagau ccgcuugacg 27900
agccuauaug gaagagcgug ccagguauuu gacucaagga cuguuaguaa cugaagaccu 27960
gacgguguug auaugg 27976
<210> 37
<211> 24
<212> DNA
<213> Artificial Sequence
<220>
<223> P1360110A PEDV 2a BD- FORWARD PRIMER N-terminal fragment
<400> 37
gatgtcacca ggtgctcagc taac 24
<210> 38
<211> 32
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDV-S2-R PEDV 2a BD- REVERSE PRIMER N-terminal fragment
<400> 38
ctatacatgt gaagcttctc agcgtcaaca ac 32
<210> 39
<211> 26
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDV-S1-F PEDV 2a BD- FORWARD PRIMER C-terminal fragment
<400> 39
catctgattc tggacagttg ttagcc 26
<210> 40
<211> 33
<212> DNA
<213> Artificial Sequence
<220>
<223> P1360110B.PEDV 2a BD- Reverse PRIMER C-terminal fragment
<400> 40
gaacactagt aatgacacaa caaagatgag aac 33
<210> 41
<211> 107
<212> DNA
<213> Artificial Sequence
<220>
<223> O1360110C gp64 signal sequence (AcNPV) cDNA
<400> 41
gatccgccac catggtatct gctattgttc tttatgtttt gcttgctgcc gccgctcact 60
cagctttcgc ggatgtcacc aggtgctcag ctaacactaa ttttagg 107
<210> 42
<211> 68
<212> DNA
<213> Artificial Sequence
<220>
<223> O1360110D gp64 C-terminal tail coding sequence (AcNPV) cDNA
<400> 42
gcttaaatac atatttagta ctgacgattg cggtttctga acactagtaa tgacacaaca 60
aagatgag 68
<210> 43
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> P290194B Gp64 Forward Primer
<400> 43
gcttaaatac atatttagta c 21
<210> 44
<211> 26
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDV-S1-F Gp64 Reverse Primer
<400> 44
catctgattc tggacagttg ttagcc 26
<210> 45
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> P290194A (OE Primer)
<400> 45
ggatccgcca ccatggtatc tg 22
<210> 46
<211> 4062
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDVS 2a BD Coding Sequence Expression Casette
<400> 46
atggtatctg ctattgttct ttatgttttg cttgctgccg ccgctcactc agctttcgcg 60
gatgtcacca ggtgctcagc taacactaat tttaggcggt tcttttcaaa atttaatgtt 120
caggcgcctg cagttgttgt actgggcggt tatctaccta ttggtgaaaa ccagggtgtc 180
aattcaactt ggtactgtgc tggccaacat ccaactgcta gtggcgttca tggtatcttt 240
gttagccata ttagaggtgg tcatggcttt gagattggca tttcgcaaga gccttttgac 300
cctagtggtt accagcttta tttacataag gctactaacg gtaacactaa tgctactgcg 360
cgactgcgca tttgccagtt tcctagcatt aaaacattgg gccccactgc taataatgat 420
gttacaacag gtcgtaattg cctatttaac aaagccatcc cagctcatat gagtgaacat 480
agtgttgtcg gcataacatg ggataatgat cgtgtcactg tcttttctga caagatctat 540
tatttttatt ttaaaaatga ttggtcccgt gttgcgacaa agtgttacaa cagtggaggt 600
tgtgctatgc aatatgttta cgaacccacc tattacatgc ttaatgttac tagtgctggt 660
gaggatggta tttcttatca accctgtaca gctaattgca ttggttatgc tgccaatgta 720
tttgctactg agcccaatgg ccacatacca gaaggtttta gttttaataa ttggtttctt 780
ttgtccaatg attccacttt ggtgcatggt aaggtggttt ccaaccaacc attgttggtc 840
aattgtcttt tggccattcc taagatttat ggactaggcc aatttttctc ctttaatcaa 900
acgatcgatg gtgtttgtaa tggagctgct gtgcagcgtg caccagaggc tctgaggttt 960
aatattaatg acacctctgt cattcttgct gaaggctcaa ttgtacttca tactgcttta 1020
ggaacaaatt tttcttttgt ttgcagtaat tcctcaaatc ctcacttagc caccttcgcc 1080
atacctttgg gtgctaccca agtaccctat tattgttttc ttaaagtgga tacttacaac 1140
tccactgttt ataaattctt ggctgtttta cctccaaccg tcagggaaat tgtcatcacc 1200
aagtatggtg atgtttatgt caatgggttt ggatacttgc atctcggttt gttggatgct 1260
gtcacaatta atttcactgg tcatggcact gacgatgatg tttctggttt ttggaccata 1320
gcatcgacta attttgttga tgcactcatc gaagttcaag gaaccgccat tcagcgtatt 1380
ctttattgtg atgatcctgt tagccaactc aagtgttctc aggttgcttt tgaccttgac 1440
gatggttttt accctatttc ttctagaaac cttctgagtc atgaacagcc aatttctttt 1500
gttactctgc catcatttaa tgatcattct tttgttaaca ttactgtatc tgcttccttt 1560
ggtggtcata gtggtgccaa ccttattgca tctgacacta ctatcaatgg gtttagttct 1620
ttctgtgttg acactagaca atttaccatt tcactgtttt ataacgttac aaacagttat 1680
ggttatgtgt ctaaatcaca ggacagtaat tgccctttca ccttgcaatc tgttaatgat 1740
tacctgtctt ttagcaaatt ttgtgtttcc accagccttt tggctagtgc ctgtaccata 1800
gatctttttg gttaccctga gtttggtagt ggtgttaagt ttacgtccct ttactttcaa 1860
ttcacaaagg gtgagttgat tactggcacg cctaaaccac ttgaaggtgt cacggacgtt 1920
tcttttatga ctctggatgt gtgtaccaag tatactatct atggctttaa aggtgagggt 1980
atcattaccc ttacaaattc tagctttttg gcaggtgttt attacacatc tgattctgga 2040
cagttgttag cctttaagaa tgtcactagt ggtgctgttt attctgttac gccatgttct 2100
ttttcagagc aggctgcata tgttgatgat gatatagtgg gtgttatttc tagtttgtct 2160
agctccactt ttaacagtac tagggagttg cctggtttct tctaccattc taatgatggc 2220
tctaattgta cagagcctgt gttggtgtat agtaacatag gtgtttgtaa atctggcagt 2280
attggctacg tcccatctca gtctggccaa gtcaagattg cacccacggt tactgggaat 2340
attagtattc ccaccaactt tagtatgagt attaggacag aatatttaca gctttacaac 2400
acgcctgtta gtgttgattg tgccacatat gtttgtaatg gtaactctcg ttgtaaacaa 2460
ttactcaccc agtacactgc agcatgtaag accatagagt cagcattaca actcagcgct 2520
aggcttgagt ctgttgaagt taactctatg cttactattt ctgaagaggc tctacagtta 2580
gctaccatta gttcgtttaa tggtgatgga tataatttta ctaatgtgct gggtgtttct 2640
gtgtatgatc ctgcaagtgg cagggtggta caaaaaaggt cttttattga agacctgctt 2700
tttaataaag tggttactaa tggccttggt actgttgatg aagactataa gcgctgttct 2760
aatggtcgct ctgtggcaga tctagtctgt gcacagtatt actctggtgt catggtacta 2820
cctggtgttg ttgacgctga gaagcttcac atgtatagtg cgtctctcat cggtggtatg 2880
gtgctaggag gttttacttc tgcagcggca ttgcctttta gctatgctgt tcaagctaga 2940
ctcaattatc ttgctctaca gacggatgtt ctacagcgga accagcaatt gcttgctgag 3000
tcttttaact ctgctattgg taatataact tcagcctttg agagtgttaa agaggctatt 3060
agtcaaactt ccaagggttt gaacactgtg gctcatgcgc ttactaaggt tcaagaggtt 3120
gttaactcgc agggtgcagc tttgactcaa cttaccgtac agctgcaaca caacttccaa 3180
gccatttcta gttctattga tgacatttac tctcgactgg acattctttc agccgatgtt 3240
caggttgacc gtctcatcac cggcagatta tcagcactta atgcttttgt tgctcaaacc 3300
ctcactaagt atactgaggt tcaggctagc aggaagttag cacagcaaaa ggttaatgag 3360
tgcgttaaat cgcaatctca gcgttatggt ttttgtggtg gtgatggcga gcacattttc 3420
tctctggtac aggcagcacc tcagggcctg ctgtttttac atacagtact tgtaccgagt 3480
gattttgtag atgttattgc catcgctggc ttatgcgtta acgatgaaat tgccttgact 3540
ctacgtgagc ctggcttagt cttgtttacg catgaacttc aaaatcatac tgcgacggaa 3600
tattttgttt catcgcgacg tatgtttgaa cctagaaaac ctaccgttag tgattttgtt 3660
caaattgaga gttgtgtggt cacctatgtc aatttgacta gagaccaact accagatgta 3720
atcccagatt acatcgatgt taacaaaaca cttgatgaga ttttagcttc tctgcccaat 3780
agaactggtc caagtcttcc tttagatgtt tttaatgcca cttatcttaa tctcactggt 3840
gaaattgcag atttagagca gcgttcagag tctctccgta atactacaga ggagctccaa 3900
agtcttatat ataatatcaa caacacacta gttgaccttg agtggctcaa ccgagttgag 3960
acatatatca agtggccgtg gtgggtttgg ttgattattt tcattgttct catctttgtt 4020
gtgtcattac tagtgttcag aaaccgcaat cgtcagtact aa 4062
<210> 47
<211> 1353
<212> PRT
<213> Artificial Sequence
<220>
<223> PEDVS 2a BD Amino Acid from Expression Casette
<400> 47
Met Val Ser Ala Ile Val Leu Tyr Val Leu Leu Ala Ala Ala Ala His
1 5 10 15
Ser Ala Phe Ala Asp Val Thr Arg Cys Ser Ala Asn Thr Asn Phe Arg
20 25 30
Arg Phe Phe Ser Lys Phe Asn Val Gln Ala Pro Ala Val Val Val Leu
35 40 45
Gly Gly Tyr Leu Pro Ile Gly Glu Asn Gln Gly Val Asn Ser Thr Trp
50 55 60
Tyr Cys Ala Gly Gln His Pro Thr Ala Ser Gly Val His Gly Ile Phe
65 70 75 80
Val Ser His Ile Arg Gly Gly His Gly Phe Glu Ile Gly Ile Ser Gln
85 90 95
Glu Pro Phe Asp Pro Ser Gly Tyr Gln Leu Tyr Leu His Lys Ala Thr
100 105 110
Asn Gly Asn Thr Asn Ala Thr Ala Arg Leu Arg Ile Cys Gln Phe Pro
115 120 125
Ser Ile Lys Thr Leu Gly Pro Thr Ala Asn Asn Asp Val Thr Thr Gly
130 135 140
Arg Asn Cys Leu Phe Asn Lys Ala Ile Pro Ala His Met Ser Glu His
145 150 155 160
Ser Val Val Gly Ile Thr Trp Asp Asn Asp Arg Val Thr Val Phe Ser
165 170 175
Asp Lys Ile Tyr Tyr Phe Tyr Phe Lys Asn Asp Trp Ser Arg Val Ala
180 185 190
Thr Lys Cys Tyr Asn Ser Gly Gly Cys Ala Met Gln Tyr Val Tyr Glu
195 200 205
Pro Thr Tyr Tyr Met Leu Asn Val Thr Ser Ala Gly Glu Asp Gly Ile
210 215 220
Ser Tyr Gln Pro Cys Thr Ala Asn Cys Ile Gly Tyr Ala Ala Asn Val
225 230 235 240
Phe Ala Thr Glu Pro Asn Gly His Ile Pro Glu Gly Phe Ser Phe Asn
245 250 255
Asn Trp Phe Leu Leu Ser Asn Asp Ser Thr Leu Val His Gly Lys Val
260 265 270
Val Ser Asn Gln Pro Leu Leu Val Asn Cys Leu Leu Ala Ile Pro Lys
275 280 285
Ile Tyr Gly Leu Gly Gln Phe Phe Ser Phe Asn Gln Thr Ile Asp Gly
290 295 300
Val Cys Asn Gly Ala Ala Val Gln Arg Ala Pro Glu Ala Leu Arg Phe
305 310 315 320
Asn Ile Asn Asp Thr Ser Val Ile Leu Ala Glu Gly Ser Ile Val Leu
325 330 335
His Thr Ala Leu Gly Thr Asn Phe Ser Phe Val Cys Ser Asn Ser Ser
340 345 350
Asn Pro His Leu Ala Thr Phe Ala Ile Pro Leu Gly Ala Thr Gln Val
355 360 365
Pro Tyr Tyr Cys Phe Leu Lys Val Asp Thr Tyr Asn Ser Thr Val Tyr
370 375 380
Lys Phe Leu Ala Val Leu Pro Pro Thr Val Arg Glu Ile Val Ile Thr
385 390 395 400
Lys Tyr Gly Asp Val Tyr Val Asn Gly Phe Gly Tyr Leu His Leu Gly
405 410 415
Leu Leu Asp Ala Val Thr Ile Asn Phe Thr Gly His Gly Thr Asp Asp
420 425 430
Asp Val Ser Gly Phe Trp Thr Ile Ala Ser Thr Asn Phe Val Asp Ala
435 440 445
Leu Ile Glu Val Gln Gly Thr Ala Ile Gln Arg Ile Leu Tyr Cys Asp
450 455 460
Asp Pro Val Ser Gln Leu Lys Cys Ser Gln Val Ala Phe Asp Leu Asp
465 470 475 480
Asp Gly Phe Tyr Pro Ile Ser Ser Arg Asn Leu Leu Ser His Glu Gln
485 490 495
Pro Ile Ser Phe Val Thr Leu Pro Ser Phe Asn Asp His Ser Phe Val
500 505 510
Asn Ile Thr Val Ser Ala Ser Phe Gly Gly His Ser Gly Ala Asn Leu
515 520 525
Ile Ala Ser Asp Thr Thr Ile Asn Gly Phe Ser Ser Phe Cys Val Asp
530 535 540
Thr Arg Gln Phe Thr Ile Ser Leu Phe Tyr Asn Val Thr Asn Ser Tyr
545 550 555 560
Gly Tyr Val Ser Lys Ser Gln Asp Ser Asn Cys Pro Phe Thr Leu Gln
565 570 575
Ser Val Asn Asp Tyr Leu Ser Phe Ser Lys Phe Cys Val Ser Thr Ser
580 585 590
Leu Leu Ala Ser Ala Cys Thr Ile Asp Leu Phe Gly Tyr Pro Glu Phe
595 600 605
Gly Ser Gly Val Lys Phe Thr Ser Leu Tyr Phe Gln Phe Thr Lys Gly
610 615 620
Glu Leu Ile Thr Gly Thr Pro Lys Pro Leu Glu Gly Val Thr Asp Val
625 630 635 640
Ser Phe Met Thr Leu Asp Val Cys Thr Lys Tyr Thr Ile Tyr Gly Phe
645 650 655
Lys Gly Glu Gly Ile Ile Thr Leu Thr Asn Ser Ser Phe Leu Ala Gly
660 665 670
Val Tyr Tyr Thr Ser Asp Ser Gly Gln Leu Leu Ala Phe Lys Asn Val
675 680 685
Thr Ser Gly Ala Val Tyr Ser Val Thr Pro Cys Ser Phe Ser Glu Gln
690 695 700
Ala Ala Tyr Val Asp Asp Asp Ile Val Gly Val Ile Ser Ser Leu Ser
705 710 715 720
Ser Ser Thr Phe Asn Ser Thr Arg Glu Leu Pro Gly Phe Phe Tyr His
725 730 735
Ser Asn Asp Gly Ser Asn Cys Thr Glu Pro Val Leu Val Tyr Ser Asn
740 745 750
Ile Gly Val Cys Lys Ser Gly Ser Ile Gly Tyr Val Pro Ser Gln Ser
755 760 765
Gly Gln Val Lys Ile Ala Pro Thr Val Thr Gly Asn Ile Ser Ile Pro
770 775 780
Thr Asn Phe Ser Met Ser Ile Arg Thr Glu Tyr Leu Gln Leu Tyr Asn
785 790 795 800
Thr Pro Val Ser Val Asp Cys Ala Thr Tyr Val Cys Asn Gly Asn Ser
805 810 815
Arg Cys Lys Gln Leu Leu Thr Gln Tyr Thr Ala Ala Cys Lys Thr Ile
820 825 830
Glu Ser Ala Leu Gln Leu Ser Ala Arg Leu Glu Ser Val Glu Val Asn
835 840 845
Ser Met Leu Thr Ile Ser Glu Glu Ala Leu Gln Leu Ala Thr Ile Ser
850 855 860
Ser Phe Asn Gly Asp Gly Tyr Asn Phe Thr Asn Val Leu Gly Val Ser
865 870 875 880
Val Tyr Asp Pro Ala Ser Gly Arg Val Val Gln Lys Arg Ser Phe Ile
885 890 895
Glu Asp Leu Leu Phe Asn Lys Val Val Thr Asn Gly Leu Gly Thr Val
900 905 910
Asp Glu Asp Tyr Lys Arg Cys Ser Asn Gly Arg Ser Val Ala Asp Leu
915 920 925
Val Cys Ala Gln Tyr Tyr Ser Gly Val Met Val Leu Pro Gly Val Val
930 935 940
Asp Ala Glu Lys Leu His Met Tyr Ser Ala Ser Leu Ile Gly Gly Met
945 950 955 960
Val Leu Gly Gly Phe Thr Ser Ala Ala Ala Leu Pro Phe Ser Tyr Ala
965 970 975
Val Gln Ala Arg Leu Asn Tyr Leu Ala Leu Gln Thr Asp Val Leu Gln
980 985 990
Arg Asn Gln Gln Leu Leu Ala Glu Ser Phe Asn Ser Ala Ile Gly Asn
995 1000 1005
Ile Thr Ser Ala Phe Glu Ser Val Lys Glu Ala Ile Ser Gln Thr
1010 1015 1020
Ser Lys Gly Leu Asn Thr Val Ala His Ala Leu Thr Lys Val Gln
1025 1030 1035
Glu Val Val Asn Ser Gln Gly Ala Ala Leu Thr Gln Leu Thr Val
1040 1045 1050
Gln Leu Gln His Asn Phe Gln Ala Ile Ser Ser Ser Ile Asp Asp
1055 1060 1065
Ile Tyr Ser Arg Leu Asp Ile Leu Ser Ala Asp Val Gln Val Asp
1070 1075 1080
Arg Leu Ile Thr Gly Arg Leu Ser Ala Leu Asn Ala Phe Val Ala
1085 1090 1095
Gln Thr Leu Thr Lys Tyr Thr Glu Val Gln Ala Ser Arg Lys Leu
1100 1105 1110
Ala Gln Gln Lys Val Asn Glu Cys Val Lys Ser Gln Ser Gln Arg
1115 1120 1125
Tyr Gly Phe Cys Gly Gly Asp Gly Glu His Ile Phe Ser Leu Val
1130 1135 1140
Gln Ala Ala Pro Gln Gly Leu Leu Phe Leu His Thr Val Leu Val
1145 1150 1155
Pro Ser Asp Phe Val Asp Val Ile Ala Ile Ala Gly Leu Cys Val
1160 1165 1170
Asn Asp Glu Ile Ala Leu Thr Leu Arg Glu Pro Gly Leu Val Leu
1175 1180 1185
Phe Thr His Glu Leu Gln Asn His Thr Ala Thr Glu Tyr Phe Val
1190 1195 1200
Ser Ser Arg Arg Met Phe Glu Pro Arg Lys Pro Thr Val Ser Asp
1205 1210 1215
Phe Val Gln Ile Glu Ser Cys Val Val Thr Tyr Val Asn Leu Thr
1220 1225 1230
Arg Asp Gln Leu Pro Asp Val Ile Pro Asp Tyr Ile Asp Val Asn
1235 1240 1245
Lys Thr Leu Asp Glu Ile Leu Ala Ser Leu Pro Asn Arg Thr Gly
1250 1255 1260
Pro Ser Leu Pro Leu Asp Val Phe Asn Ala Thr Tyr Leu Asn Leu
1265 1270 1275
Thr Gly Glu Ile Ala Asp Leu Glu Gln Arg Ser Glu Ser Leu Arg
1280 1285 1290
Asn Thr Thr Glu Glu Leu Gln Ser Leu Ile Tyr Asn Ile Asn Asn
1295 1300 1305
Thr Leu Val Asp Leu Glu Trp Leu Asn Arg Val Glu Thr Tyr Ile
1310 1315 1320
Lys Trp Pro Trp Trp Val Trp Leu Ile Ile Phe Ile Val Leu Ile
1325 1330 1335
Phe Val Val Ser Leu Leu Val Phe Arg Asn Arg Asn Arg Gln Tyr
1340 1345 1350
<210> 48
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> P290194A PEDV 2b BD- FORWARD PRIMER N-terminal fragment
<400> 48
ggatccgcca ccatggtatc tg 22
<210> 49
<211> 25
<212> DNA
<213> Artificial Sequence
<220>
<223> P3183131B PEDV 2b BD- Reverse PRIMER N-terminal fragment
<400> 49
gtcgatgcta tggtccaaaa accag 25
<210> 50
<211> 22
<212> DNA
<213> Artificial Sequence
<220>
<223> P3183131A PEDV 2b BD- FORWARD PRIMER C-terminal fragment
<400> 50
ggatccgcca ccatggtatc tg 22
<210> 51
<211> 21
<212> DNA
<213> Artificial Sequence
<220>
<223> P290194B PEDV 2b BD- REVERSE PRIMER C-terminal fragment
<400> 51
gcttaaatac atatttagta c 21
<210> 52
<211> 4053
<212> DNA
<213> Artificial Sequence
<220>
<223> PEDVS 2b BD CDS cDNA
<400> 52
atggtatctg ctattgttct ttatgttttg cttgctgccg ccgctcactc agctttcgcg 60
gatgtcacta ggtgccagtc cactattaac ttcaggcggt tcttttcaaa atttaatgtg 120
caggcacctg ctgtcgttgt gttgggtggt tatctaccta gtatgaactc ctctagctgg 180
tactgtggca caggtcttga aactgctagt ggcgtgcatg gtattttcct cagttacatc 240
gattctggtc agggctttga gattggcatt tcacaggagc cgtttgatcc tagtggttac 300
cagctttatt tacataaggc cactaatggt aaccataatg ctattgcacg actgcgcatt 360
tgccagtttc caaataataa tacattgggc cctactgtta atgatgttac aacaggtcgt 420
aactgcctat tcaacaaagc cattccagct tatatgcagg atggaaaaaa catcgttgtc 480
ggcataacat gggacaatga tcgtgtcact gtttttgctg acaagatcta tcatttttat 540
ctcaaaaatg attggtcccg tgttgcgaca agatgttaca ataaaagaag ttgtgctatg 600
caatatgttt atacacctac ctactacatg cttaatgtta ctagtgcagg tgaggatggc 660
atttattatg aaccatgtac agctaattgc agtggttacg ctgccaatgt gtttgccact 720
gattctaatg gccacatacc agaaggtttt agttttaata attggtttct tttgtccaat 780
gattccactt tgttgcatgg taaggtggtt tccaaccaac ctttgttggt caattgtctt 840
ttggccattc ctaagattta tggactaggc caatttttct cattcaatca aacgatggat 900
ggcgtttgta atggagctgc tgcgcagcgt gcaccagagg ctctgaggtt taatattaat 960
gacacctctg tcattcttgc tgaaggctca attgtacttc acactgcttt aggaacaaat 1020
ctttcttttg tttgcagtaa ttcttcagat cctcatttag ctaccttcac catacctctg 1080
ggtgctaccc aagtacccta ttattgtttt cttaaagtgg atacttacaa ctccactgtt 1140
tataaatttt tggctgtttt acctcctacc gtcagggaaa ttgtcatcac caagtatggt 1200
gatgtttatg tcaatgggtt tggatacttg catctcggtt tgttggatgc tgtcacaatt 1260
aatttcactg gtcatggcac tgacgatgat gtttctggtt tttggaccat agcatcgact 1320
aattttgttg atgcactcat cgaagttcaa ggaaccgcca ttcagcgtat tctttattgt 1380
gatgatcctg ttagccaact caagtgttct caggttgctt ttgaccttga cgatggtttt 1440
taccctattt cttctagaaa ccttctgagt catgaacagc caatttcttt tgttactctg 1500
ccatcattta atgatcattc ttttgttaac attactgtat ctgcttcctt tggtggtcat 1560
agtggtgcca accttattgc atctgacact actatcaatg ggtttagttc tttctgtgtt 1620
gacactagac aatttaccat ttcactgttt tataacgtta caaacagtta tggttatgtg 1680
tctaaatcac aggacagtaa ttgccctttc accttgcaat ctgttaatga ttacctgtct 1740
tttagcaaat tttgtgtttc caccagcctt ttggctagtg cctgtaccat agatcttttt 1800
ggttaccctg agtttggtag tggtgttaag tttacgtccc tttactttca attcacaaag 1860
ggtgagttga ttactggcac gcctaaacca cttgaaggtg tcacggacgt ttcttttatg 1920
actctggatg tgtgtaccaa gtatactatc tatggcttta aaggtgaggg tatcattacc 1980
cttacaaatt ctagcttttt ggcaggtgtt tattacacat ctgattctgg acagttgtta 2040
gcctttaaga atgtcactag tggtgctgtt tattctgtta cgccatgttc tttttcagag 2100
caggctgcat atgttgatga tgatatagtg ggtgttattt ctagtttgtc tagctccact 2160
tttaacagta ctagggagtt gcctggtttc ttctaccatt ctaatgatgg ctctaattgt 2220
acagagcctg tgttggtgta tagtaacata ggtgtttgta aatctggcag tattggctac 2280
gtcccatctc agtctggcca agtcaagatt gcacccacgg ttactgggaa tattagtatt 2340
cccaccaact ttagtatgag tattaggaca gaatatttac agctttacaa cacgcctgtt 2400
agtgttgatt gtgccacata tgtttgtaat ggtaactctc gttgtaaaca attactcacc 2460
cagtacactg cagcatgtaa gaccatagag tcagcattac aactcagcgc taggcttgag 2520
tctgttgaag ttaactctat gcttactatt tctgaagagg ctctacagtt agctaccatt 2580
agttcgttta atggtgatgg atataatttt actaatgtgc tgggtgtttc tgtgtatgat 2640
cctgcaagtg gcagggtggt acaaaaaagg tcttttattg aagacctgct ttttaataaa 2700
gtggttacta atggccttgg tactgttgat gaagactata agcgctgttc taatggtcgc 2760
tctgtggcag atctagtctg tgcacagtat tactctggtg tcatggtact acctggtgtt 2820
gttgacgctg agaagcttca catgtatagt gcgtctctca tcggtggtat ggtgctagga 2880
ggttttactt ctgcagcggc attgcctttt agctatgctg ttcaagctag actcaattat 2940
cttgctctac agacggatgt tctacagcgg aaccagcaat tgcttgctga gtcttttaac 3000
tctgctattg gtaatataac ttcagccttt gagagtgtta aagaggctat tagtcaaact 3060
tccaagggtt tgaacactgt ggctcatgcg cttactaagg ttcaagaggt tgttaactcg 3120
cagggtgcag ctttgactca acttaccgta cagctgcaac acaacttcca agccatttct 3180
agttctattg atgacattta ctctcgactg gacattcttt cagccgatgt tcaggttgac 3240
cgtctcatca ccggcagatt atcagcactt aatgcttttg ttgctcaaac cctcactaag 3300
tatactgagg ttcaggctag caggaagtta gcacagcaaa aggttaatga gtgcgttaaa 3360
tcgcaatctc agcgttatgg tttttgtggt ggtgatggcg agcacatttt ctctctggta 3420
caggcagcac ctcagggcct gctgttttta catacagtac ttgtaccgag tgattttgta 3480
gatgttattg ccatcgctgg cttatgcgtt aacgatgaaa ttgccttgac tctacgtgag 3540
cctggcttag tcttgtttac gcatgaactt caaaatcata ctgcgacgga atattttgtt 3600
tcatcgcgac gtatgtttga acctagaaaa cctaccgtta gtgattttgt tcaaattgag 3660
agttgtgtgg tcacctatgt caatttgact agagaccaac taccagatgt aatcccagat 3720
tacatcgatg ttaacaaaac acttgatgag attttagctt ctctgcccaa tagaactggt 3780
ccaagtcttc ctttagatgt ttttaatgcc acttatctta atctcactgg tgaaattgca 3840
gatttagagc agcgttcaga gtctctccgt aatactacag aggagctcca aagtcttata 3900
tataatatca acaacacact agttgacctt gagtggctca accgagttga gacatatatc 3960
aagtggccgt ggtgggtttg gttgattatt ttcattgttc tcatctttgt tgtgtcatta 4020
ctagtgttca gaaaccgcaa tcgtcagtac taa 4053
<210> 53
<211> 1350
<212> PRT
<213> Artificial Sequence
<220>
<223> PEDVS 2b BD Expression Casetter Amino Acid Sequence
<400> 53
Met Val Ser Ala Ile Val Leu Tyr Val Leu Leu Ala Ala Ala Ala His
1 5 10 15
Ser Ala Phe Ala Asp Val Thr Arg Cys Gln Ser Thr Ile Asn Phe Arg
20 25 30
Arg Phe Phe Ser Lys Phe Asn Val Gln Ala Pro Ala Val Val Val Leu
35 40 45
Gly Gly Tyr Leu Pro Ser Met Asn Ser Ser Ser Trp Tyr Cys Gly Thr
50 55 60
Gly Leu Glu Thr Ala Ser Gly Val His Gly Ile Phe Leu Ser Tyr Ile
65 70 75 80
Asp Ser Gly Gln Gly Phe Glu Ile Gly Ile Ser Gln Glu Pro Phe Asp
85 90 95
Pro Ser Gly Tyr Gln Leu Tyr Leu His Lys Ala Thr Asn Gly Asn His
100 105 110
Asn Ala Ile Ala Arg Leu Arg Ile Cys Gln Phe Pro Asn Asn Asn Thr
115 120 125
Leu Gly Pro Thr Val Asn Asp Val Thr Thr Gly Arg Asn Cys Leu Phe
130 135 140
Asn Lys Ala Ile Pro Ala Tyr Met Gln Asp Gly Lys Asn Ile Val Val
145 150 155 160
Gly Ile Thr Trp Asp Asn Asp Arg Val Thr Val Phe Ala Asp Lys Ile
165 170 175
Tyr His Phe Tyr Leu Lys Asn Asp Trp Ser Arg Val Ala Thr Arg Cys
180 185 190
Tyr Asn Lys Arg Ser Cys Ala Met Gln Tyr Val Tyr Thr Pro Thr Tyr
195 200 205
Tyr Met Leu Asn Val Thr Ser Ala Gly Glu Asp Gly Ile Tyr Tyr Glu
210 215 220
Pro Cys Thr Ala Asn Cys Ser Gly Tyr Ala Ala Asn Val Phe Ala Thr
225 230 235 240
Asp Ser Asn Gly His Ile Pro Glu Gly Phe Ser Phe Asn Asn Trp Phe
245 250 255
Leu Leu Ser Asn Asp Ser Thr Leu Leu His Gly Lys Val Val Ser Asn
260 265 270
Gln Pro Leu Leu Val Asn Cys Leu Leu Ala Ile Pro Lys Ile Tyr Gly
275 280 285
Leu Gly Gln Phe Phe Ser Phe Asn Gln Thr Met Asp Gly Val Cys Asn
290 295 300
Gly Ala Ala Ala Gln Arg Ala Pro Glu Ala Leu Arg Phe Asn Ile Asn
305 310 315 320
Asp Thr Ser Val Ile Leu Ala Glu Gly Ser Ile Val Leu His Thr Ala
325 330 335
Leu Gly Thr Asn Leu Ser Phe Val Cys Ser Asn Ser Ser Asp Pro His
340 345 350
Leu Ala Thr Phe Thr Ile Pro Leu Gly Ala Thr Gln Val Pro Tyr Tyr
355 360 365
Cys Phe Leu Lys Val Asp Thr Tyr Asn Ser Thr Val Tyr Lys Phe Leu
370 375 380
Ala Val Leu Pro Pro Thr Val Arg Glu Ile Val Ile Thr Lys Tyr Gly
385 390 395 400
Asp Val Tyr Val Asn Gly Phe Gly Tyr Leu His Leu Gly Leu Leu Asp
405 410 415
Ala Val Thr Ile Asn Phe Thr Gly His Gly Thr Asp Asp Asp Val Ser
420 425 430
Gly Phe Trp Thr Ile Ala Ser Thr Asn Phe Val Asp Ala Leu Ile Glu
435 440 445
Val Gln Gly Thr Ala Ile Gln Arg Ile Leu Tyr Cys Asp Asp Pro Val
450 455 460
Ser Gln Leu Lys Cys Ser Gln Val Ala Phe Asp Leu Asp Asp Gly Phe
465 470 475 480
Tyr Pro Ile Ser Ser Arg Asn Leu Leu Ser His Glu Gln Pro Ile Ser
485 490 495
Phe Val Thr Leu Pro Ser Phe Asn Asp His Ser Phe Val Asn Ile Thr
500 505 510
Val Ser Ala Ser Phe Gly Gly His Ser Gly Ala Asn Leu Ile Ala Ser
515 520 525
Asp Thr Thr Ile Asn Gly Phe Ser Ser Phe Cys Val Asp Thr Arg Gln
530 535 540
Phe Thr Ile Ser Leu Phe Tyr Asn Val Thr Asn Ser Tyr Gly Tyr Val
545 550 555 560
Ser Lys Ser Gln Asp Ser Asn Cys Pro Phe Thr Leu Gln Ser Val Asn
565 570 575
Asp Tyr Leu Ser Phe Ser Lys Phe Cys Val Ser Thr Ser Leu Leu Ala
580 585 590
Ser Ala Cys Thr Ile Asp Leu Phe Gly Tyr Pro Glu Phe Gly Ser Gly
595 600 605
Val Lys Phe Thr Ser Leu Tyr Phe Gln Phe Thr Lys Gly Glu Leu Ile
610 615 620
Thr Gly Thr Pro Lys Pro Leu Glu Gly Val Thr Asp Val Ser Phe Met
625 630 635 640
Thr Leu Asp Val Cys Thr Lys Tyr Thr Ile Tyr Gly Phe Lys Gly Glu
645 650 655
Gly Ile Ile Thr Leu Thr Asn Ser Ser Phe Leu Ala Gly Val Tyr Tyr
660 665 670
Thr Ser Asp Ser Gly Gln Leu Leu Ala Phe Lys Asn Val Thr Ser Gly
675 680 685
Ala Val Tyr Ser Val Thr Pro Cys Ser Phe Ser Glu Gln Ala Ala Tyr
690 695 700
Val Asp Asp Asp Ile Val Gly Val Ile Ser Ser Leu Ser Ser Ser Thr
705 710 715 720
Phe Asn Ser Thr Arg Glu Leu Pro Gly Phe Phe Tyr His Ser Asn Asp
725 730 735
Gly Ser Asn Cys Thr Glu Pro Val Leu Val Tyr Ser Asn Ile Gly Val
740 745 750
Cys Lys Ser Gly Ser Ile Gly Tyr Val Pro Ser Gln Ser Gly Gln Val
755 760 765
Lys Ile Ala Pro Thr Val Thr Gly Asn Ile Ser Ile Pro Thr Asn Phe
770 775 780
Ser Met Ser Ile Arg Thr Glu Tyr Leu Gln Leu Tyr Asn Thr Pro Val
785 790 795 800
Ser Val Asp Cys Ala Thr Tyr Val Cys Asn Gly Asn Ser Arg Cys Lys
805 810 815
Gln Leu Leu Thr Gln Tyr Thr Ala Ala Cys Lys Thr Ile Glu Ser Ala
820 825 830
Leu Gln Leu Ser Ala Arg Leu Glu Ser Val Glu Val Asn Ser Met Leu
835 840 845
Thr Ile Ser Glu Glu Ala Leu Gln Leu Ala Thr Ile Ser Ser Phe Asn
850 855 860
Gly Asp Gly Tyr Asn Phe Thr Asn Val Leu Gly Val Ser Val Tyr Asp
865 870 875 880
Pro Ala Ser Gly Arg Val Val Gln Lys Arg Ser Phe Ile Glu Asp Leu
885 890 895
Leu Phe Asn Lys Val Val Thr Asn Gly Leu Gly Thr Val Asp Glu Asp
900 905 910
Tyr Lys Arg Cys Ser Asn Gly Arg Ser Val Ala Asp Leu Val Cys Ala
915 920 925
Gln Tyr Tyr Ser Gly Val Met Val Leu Pro Gly Val Val Asp Ala Glu
930 935 940
Lys Leu His Met Tyr Ser Ala Ser Leu Ile Gly Gly Met Val Leu Gly
945 950 955 960
Gly Phe Thr Ser Ala Ala Ala Leu Pro Phe Ser Tyr Ala Val Gln Ala
965 970 975
Arg Leu Asn Tyr Leu Ala Leu Gln Thr Asp Val Leu Gln Arg Asn Gln
980 985 990
Gln Leu Leu Ala Glu Ser Phe Asn Ser Ala Ile Gly Asn Ile Thr Ser
995 1000 1005
Ala Phe Glu Ser Val Lys Glu Ala Ile Ser Gln Thr Ser Lys Gly
1010 1015 1020
Leu Asn Thr Val Ala His Ala Leu Thr Lys Val Gln Glu Val Val
1025 1030 1035
Asn Ser Gln Gly Ala Ala Leu Thr Gln Leu Thr Val Gln Leu Gln
1040 1045 1050
His Asn Phe Gln Ala Ile Ser Ser Ser Ile Asp Asp Ile Tyr Ser
1055 1060 1065
Arg Leu Asp Ile Leu Ser Ala Asp Val Gln Val Asp Arg Leu Ile
1070 1075 1080
Thr Gly Arg Leu Ser Ala Leu Asn Ala Phe Val Ala Gln Thr Leu
1085 1090 1095
Thr Lys Tyr Thr Glu Val Gln Ala Ser Arg Lys Leu Ala Gln Gln
1100 1105 1110
Lys Val Asn Glu Cys Val Lys Ser Gln Ser Gln Arg Tyr Gly Phe
1115 1120 1125
Cys Gly Gly Asp Gly Glu His Ile Phe Ser Leu Val Gln Ala Ala
1130 1135 1140
Pro Gln Gly Leu Leu Phe Leu His Thr Val Leu Val Pro Ser Asp
1145 1150 1155
Phe Val Asp Val Ile Ala Ile Ala Gly Leu Cys Val Asn Asp Glu
1160 1165 1170
Ile Ala Leu Thr Leu Arg Glu Pro Gly Leu Val Leu Phe Thr His
1175 1180 1185
Glu Leu Gln Asn His Thr Ala Thr Glu Tyr Phe Val Ser Ser Arg
1190 1195 1200
Arg Met Phe Glu Pro Arg Lys Pro Thr Val Ser Asp Phe Val Gln
1205 1210 1215
Ile Glu Ser Cys Val Val Thr Tyr Val Asn Leu Thr Arg Asp Gln
1220 1225 1230
Leu Pro Asp Val Ile Pro Asp Tyr Ile Asp Val Asn Lys Thr Leu
1235 1240 1245
Asp Glu Ile Leu Ala Ser Leu Pro Asn Arg Thr Gly Pro Ser Leu
1250 1255 1260
Pro Leu Asp Val Phe Asn Ala Thr Tyr Leu Asn Leu Thr Gly Glu
1265 1270 1275
Ile Ala Asp Leu Glu Gln Arg Ser Glu Ser Leu Arg Asn Thr Thr
1280 1285 1290
Glu Glu Leu Gln Ser Leu Ile Tyr Asn Ile Asn Asn Thr Leu Val
1295 1300 1305
Asp Leu Glu Trp Leu Asn Arg Val Glu Thr Tyr Ile Lys Trp Pro
1310 1315 1320
Trp Trp Val Trp Leu Ile Ile Phe Ile Val Leu Ile Phe Val Val
1325 1330 1335
Ser Leu Leu Val Phe Arg Asn Arg Asn Arg Gln Tyr
1340 1345 1350

Claims (52)

1.一种免疫原性组合物，其包含一种或多种猪丁型冠状病毒(PDCoV)抗原和佐剂，其中在所述猪丁型冠状病毒(PDCoV)中的是以下任何PDCoV：
a)其由SEQ ID NO：1、5或9编码，和/或包含SEQ ID NO：1、5或9的序列，和/或包含SEQID NO：1、5或9的RNA等价物；
b)其序列与SEQ ID NO：1至少99％相同，和/或与SEQ ID NO：1、5或9的RNA等价物至少99％相同；
c)其刺突蛋白由SEQ ID NO：3、7、11、17或27的核酸序列编码；
d)其刺突蛋白由与SEQ ID NO：3、7、11、17或27至少90％相同的核酸序列编码；
e)其由SEQ ID NO：2、6或10编码；或
f)其序列与SEQ ID NO：2、6或10至少99％相同。
2.根据权利要求1的免疫原性组合物，其中所述佐剂是水包油型乳状液。
3.权利要求1或2的免疫原性组合物，其中所述免疫原性组合物是重组抗原或灭活的完整猪丁型冠状病毒(PDCoV)抗原。
4.权利要求1-3中任一项的免疫原性组合物，其中所述抗原是灭活的完整PDCoV抗原。
5.权利要求4的免疫原性组合物，其中所述PDCoV抗原是化学灭活的。
6.权利要求5的免疫原性组合物，其中所述PDCoV抗原通过用化学灭活剂处理而化学灭活，所述化学灭活剂包括选自乙烯亚胺、二乙烯亚胺、乙酰乙烯亚胺及其混合物的化合物。
7.权利要求6的免疫原性组合物，其中所述PDCoV通过用二乙烯亚胺处理而化学灭活。
8.权利要求1-7中任一项的免疫原性组合物，其中所述佐剂是基于水包油型乳状液的佐剂。
9.权利要求4的免疫原性组合物，其中灭活的猪丁型冠状病毒(PDCoV)包含SEQ ID NO：1、5或9，和/或包含SEQ ID NO：1、5或9的RNA等价物。
10.权利要求1-3任一项的免疫原性组合物，其中所述PDCoV抗原是重组抗原。
11.权利要求10的免疫原性组合物，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：
a)编码猪丁型冠状病毒(PDCoV)刺突蛋白的抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：4、8、12、18或28具有至少90％的同源性；
b)包含a)的分离核酸的重组载体；
c)由a)的核酸编码的重组PDCoV刺突蛋白；和
d)其任何组合。
12.权利要求10或11的免疫原性组合物，其中此类免疫原性组合物包含药学上可接受的载体和/或赋形剂。
13.权利要求10-12中任一项的免疫原性组合物，其中所述水包油型乳状液是基于水包油型乳状液的佐剂。
14.权利要求1-13中任一项的免疫原性组合物，其中所述免疫原性组合物进一步包含一种或多种另外的抗原。
15.权利要求14的免疫原性组合物，其中所述另外的抗原是猪流行性腹泻病毒(PEDV)的抗原，其中所述猪流行性腹泻病毒(PEDV)是以下任何PEDV：
a)其由SEQ ID NO：29编码，和/或包含SEQ ID NO：29或33的序列，和/或包含SEQ IDNO：29或33的RNA等价物；
b)其序列与SEQ ID NO：29或33至少99％相同，和/或与SEQ ID NO：29或33的RNA等价物至少99％相同；
c)其刺突蛋白由SEQ ID NO：30、34、46或52的核酸序列编码；
d)其刺突蛋白由与SEQ ID NO：30、34、46或52至少90％相同的核酸序列编码；
e)其由SEQ ID NO：32或36编码；或
f)其由与SEQ ID NO：32或36至少99％相同的序列编码。
16.权利要求15的免疫原性组合物，其中所述另外的PEDV抗原是重组抗原或灭活的完整猪流行性腹泻病毒(PEDV)。
17.权利要求16的免疫原性组合物，其中所述抗原是灭活的完整猪流行性腹泻病毒(PEDV)。
18.权利要求16的免疫原性组合物，其中所述抗原是重组PEDV抗原。
19.权利要求16或18中任一项的免疫原性组合物，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：
a)编码猪流行性腹泻病毒(PEDV)刺突蛋白的抗原的分离核酸，其中所述抗原与SEQ IDNO：31、35、47或52具有至少90％的同源性；
b)包含a)的分离核酸的重组载体；
c)由a)的核酸编码的重组PEDV刺突蛋白；和
d)其任何组合。
20.权利要求14、15、16或18中任一项的免疫原性组合物，其中所述重组抗原包含选自以下的一种或多种免疫原性组分：猪流行性腹泻病毒(PEDV)的结构蛋白M、E或N。
21.权利要求19的免疫原性组合物，其中所述免疫原性组分是所述分离的核酸。
22.权利要求19的免疫原性组合物，其中所述免疫原性组分是所述重组载体。
23.权利要求19的免疫原性组合物，其中所述免疫原性组分是所述重组猪流行性腹泻病毒(PEDV)刺突蛋白。
24.权利要求19的免疫原性组合物，其中所述免疫原性组分是组合。
25.一种用于减少与猪丁型冠状病毒(PDCoV)相关的疾病的临床症状的方法，其包括给猪施用权利要求1-24中任一项的免疫原性组合物。
26.一种用于减少与猪丁型冠状病毒(PDCoV)相关的疾病的临床症状的方法，其包括给猪施用权利要求4的免疫原性组合物。
27.一种用于减少与猪丁型冠状病毒(PDCoV)相关的疾病的临床症状的方法，其包括给猪施用权利要求10的免疫原性组合物。
28.权利要求25的方法，其中此类免疫原性组合物包含选自以下的一种或多种免疫原性组分：
a)其由SEQ ID NO：1、5或9编码，和/或包含SEQ ID NO：1的序列，和/或包含SEQ ID NO：1、5或9的RNA等价物；
b)其序列与SEQ ID NO：1、5或9至少99％相同，和/或与SEQ ID NO：1、5或9的RNA等价物至少99％相同；
c)其刺突蛋白由SEQ ID NO：3、7、11、17或27的核酸序列编码；
d)其刺突蛋白由与SEQ ID NO：3、7、11、17或27至少90％相同的核酸序列编码；
e)其由SEQ ID NO：2、6或10编码；和
f)其序列与SEQ ID NO：2、6或10至少99％相同。
29.权利要求26的方法，其中此类免疫原性组合物包含选自以下的一种或多种免疫原性组分：
a)其由SEQ ID NO：1、5或9的序列编码，或者包含SEQ ID NO：1、5或9的序列；
b)其序列与SEQ ID NO：1、5或9至少99％相同；
c)其刺突蛋白由SEQ ID NO：3、7、11、17或27的核酸序列编码；和
d)其刺突蛋白由与SEQ ID NO：3、7、11、17或27至少90％相同的核酸序列编码。
30.权利要求27的方法，其中此类重组抗原包含选自以下的一种或多种免疫原性组分：
a)编码猪流行性腹泻病毒、猪丁型冠状病毒(PDCoV)刺突蛋白抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：4、8、12、18或28具有至少90％的同源性；
b)包含a)的分离核酸的重组载体；
c)由a)的核酸编码的重组猪丁型冠状病毒(PDCoV)刺突蛋白；和
d)其任何组合。
31.一种用于减少与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的临床症状的方法，其包括给猪施用权利要求15-24中任一项的免疫原性组合物。
32.一种减少与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的临床症状的方法，其包括给此类猪施用权利要求14-17中任一项的免疫原性组合物。
33.一种用于减少与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的临床体征的方法，其包括给此类猪施用权利要求14、15、16或18-24中任一项的免疫原性组合物。
34.一种用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)相关的疾病的免疫原性应答的试剂盒，其包含：
a)能够给猪施用免疫原性组合物的分配器；和
b)根据权利要求1-14中任一项的免疫原性组合物。
35.一种用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)相关的疾病的免疫原性应答的试剂盒，其包含：
a)能够给猪施用免疫原性的分配器；和
b)根据权利要求4-9中任一项的免疫原性组合物。
36.一种用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)相关的疾病的免疫原性应答的试剂盒，其包含：
a)能够给猪施用免疫原性组合物的分配器；和
b)根据权利要求10-14中任一项的免疫原性组合物。
37.一种用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的免疫原性应答的试剂盒，其包含：
a)能够给猪施用免疫原性组合物的分配器；和
b)根据权利要求15-24中任一项的免疫原性组合物。
38.一种用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的免疫原性应答的试剂盒，其包含：
a)能够给猪施用免疫原性组合物的分配器；和
b)根据权利要求15-17中任一项的免疫原性组合物。
39.一种用于在猪中诱导针对与猪丁型冠状病毒(PDCoV)和猪流行性腹泻病毒(PEDV)相关的疾病的免疫原性应答的试剂盒，其包含：
a)能够给猪施用免疫原性组合物的分配器；和
b)根据权利要求18-24中任一项的免疫原性组合物。
40.一种产生根据权利要求4-9中任一项的猪丁型冠状病毒(PDCoV)免疫原性组合物的方法，其包括：
a)用猪丁型冠状病毒(PDCoV)接种猪睾丸细胞；
b)温育所述接种的猪睾丸细胞；
c)从所述温育的细胞中收获猪丁型冠状病毒(PDCoV)；和
d)用化学灭活剂，优选用选自乙烯亚胺、二乙烯亚胺、乙酰乙烯亚胺或其混合物的化合物处理所述收获的细胞，以形成灭活的猪丁型冠状病毒(PDCoV)抗原。
41.权利要求40的方法，其中所述猪丁型冠状病毒(PDCoV)包含：
a)由SEQ ID NO：1、5或9的序列编码，或者包含SEQ ID NO：1、5或9的序列；
b)与SEQ ID NO：1、5或9至少99％相同的序列；
c)由SEQ ID NO：3、7、11、17或27的核酸序列编码的刺突蛋白；
d)由与SEQ ID NO：3、7、11、17或27至少90％相同的核酸序列编码的刺突蛋白。
42.权利要求41的方法，其中所述猪丁型冠状病毒(PDCoV)包含SEQ ID NO：1、5或9，和/或SEQ ID NO：1、5或9的RNA等价物。
43.权利要求40的方法，其中所述猪睾丸细胞是AI-ST细胞。
44.权利要求43的方法，其中所述化学灭活剂包括二乙烯亚胺。
45.权利要求40的方法，其进一步包括给所述猪丁型冠状病毒(PDCoV)抗原添加基于水包油型乳状液的佐剂
46.一种产生免疫原性组合物的方法，所述免疫原性组合物包含权利要求10-24中任一项的重组猪丁型冠状病毒(PDCoV)抗原，所述方法包括：
a)在宿主细胞中表达猪丁型冠状病毒(PDCoV)的抗原；
b)收获所述猪丁型冠状病毒(PDCoV)细胞的抗原；和
c)将基于水包油型乳状液的佐剂加入步骤b)的所述猪丁型冠状病毒(PDCoV)抗原中。
47.权利要求46的方法，其中所述猪丁型冠状病毒(PDCoV)抗原包含：
a)编码猪丁型冠状病毒(PDCoV)刺突蛋白的抗原的分离核酸，其中所述重组刺突多肽与SEQ ID NO：4、8、12、18或28具有至少90％的同源性；
b)包含a)的分离核酸的重组载体；
c)由a)的核酸编码的重组猪丁型冠状病毒(PDCoV)刺突蛋白；和
d)其任何组合。
48.权利要求46的方法，其中所述猪丁型冠状病毒(PDCoV)的抗原由重组杆状病毒载体表达。
49.权利要求48的方法，其中所述猪丁型冠状病毒(PDCoV)的抗原在昆虫细胞中表达。
50.权利要求46的方法，其中所述基于水包油的佐剂是基于水包油型乳状液的佐剂。
51.权利要求1-14中任一项的免疫原性组合物，其用于减少与猪丁型冠状病毒(PDCoV)相关的疾病的症状。
52.权利要求15-24中任一项的免疫原性组合物，其用于减少与猪丁型冠状病毒(PDCoV)和/或猪流行病毒(PEDV)相关的疾病的症状。
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