CN112375784A - 制备重组新型冠状病毒Spike蛋白的方法 - Google Patents

制备重组新型冠状病毒Spike蛋白的方法 Download PDF

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CN112375784A
CN112375784A CN202110020323.7A CN202110020323A CN112375784A CN 112375784 A CN112375784 A CN 112375784A CN 202110020323 A CN202110020323 A CN 202110020323A CN 112375784 A CN112375784 A CN 112375784A
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秦丽丽
宋洁
王芳琪
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Beijing Baipusai Biotechnology Co Ltd
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Abstract

本发明提供了一种制备重组新型冠状病毒Spike蛋白的方法。本发明首先提供以下多肽作为信号肽在制备重组新型冠状病毒Spike蛋白中的应用：SEQ ID No. 10所示氨基酸序列组成的多肽。本发明采用特定信号肽，构建含有编码重组新型冠状病毒Spike蛋白的多核苷酸的表达载体，转染哺乳动物细胞以分泌表达重组新型冠状病毒Spike蛋白，可显著提高Spike蛋白在HEK293细胞中的分泌表达水平。

Description

制备重组新型冠状病毒Spike蛋白的方法
技术领域
本发明是关于一种制备重组新型冠状病毒Spike蛋白的方法，更具体地说，是关于一种利用特定信号肽在哺乳动物细胞中高效分泌表达新型冠状病毒Spike蛋白的方法。
背景技术
新型冠状病毒(即SARS-CoV-2，或称为COVID-19)，是2020年在全球大规模爆发的冠状病毒新毒株，与SARS冠状病毒同为β属冠状病毒。为尽快得到新型冠状病毒的有效防治手段，各国科研单位正夜以继日开发各种抗病毒药物和治疗方案。
Spike蛋白表达于冠状病毒的表面，在识别并结合宿主细胞表面受体以及介导病毒包膜与细胞膜融合的过程中起到关键性作用。在感染过程中，Spike蛋白被宿主蛋白酶（如TMPRSS2）裂解为N末端S1亚基和C末端S2亚基，并使构象从融合前转变为融合后状态。S1和S2分别介导受体结合和膜融合。由N末端结构域（NTD）和受体结合结构域（RBD）组成的S1在确定组织嗜性和宿主范围方面至关重要。通常，针对新型冠状病毒的中和抗体往往靶向Spike蛋白的RBD，并阻断RBD与受体ACE2之间的结合。最近，中国军事医学科学院陈薇、李建民、西湖大学周强等研究人员合作发现一个结合新型冠状病毒NTD结构域的中和性人源抗体。因此，抗新型冠状病毒的疗法可以通过抗原表位的分布来指导，不仅限于Spike蛋白的RBD，还包括整个Spike蛋白。
作为主要抗病毒药物靶点的Spike蛋白，是一个三聚体蛋白。由于其分子量较大，且其prefusion状态下是处于亚稳定的三聚状态，导致其全长胞外区很难获得高水平的重组表达，严重限制了其在抗病毒药物开发和诊断领域的应用。
发明内容
本发明的一个目的在于提供一种制备重组新型冠状病毒Spike蛋白的方法。
本案发明人在研究中发现一种信号肽可以有效提高新型冠状病毒Spike蛋白在HEK293细胞中分泌表达，能够降低利用HEK293细胞生产Spike重组蛋白的成本，并实现短期内快速满足抗病毒药物开发和新型冠状病毒诊断对Spike蛋白的大量需求。
根据本发明的具体实施方案，一方面，本发明提供了以下多肽作为信号肽在制备重组新型冠状病毒Spike蛋白中的应用：
SEQ ID No. 10所示氨基酸序列组成的多肽。
另一方面，本发明提供了一种制备重组新型冠状病毒Spike蛋白的方法，该方法包括：
构建含有编码重组新型冠状病毒Spike蛋白的多核苷酸的表达载体，转染哺乳动物细胞以分泌表达重组新型冠状病毒Spike蛋白；
其中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列中包括了编码信号肽的核苷酸序列，所述信号肽为：SEQ ID No. 10所示氨基酸序列组成的多肽。
本发明中，所述SEQ ID No. 10所示氨基酸序列如下：
MAAPALSWRLPLLILLLPLATSWASA。
新型冠状病毒Spike蛋白的氨基酸序列如SEQ ID No. 1所示，其自身信号肽的氨基酸序列如SEQ ID No. 2所示。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列除包括编码信号肽的核苷酸序列外，还包括编码目标蛋白（即新型冠状病毒Spike蛋白）的核苷酸序列，具体而言，包括编码SEQ ID No. 1所示氨基酸序列的核苷酸序列。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，优选在新冠病毒Spike蛋白的C末端均引入一段T4 fibritin三聚体基序用于提高Spike蛋白的三聚稳定性。从而，本发明的制备重组新型冠状病毒Spike蛋白的方法中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列还包括：编码设计在重组新型冠状病毒Spike蛋白的C端的T4 fibritin三聚体基序的核苷酸序列，所述T4 fibritin三聚体基序包括SEQ ID No. 12所示氨基酸序列。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，进一步优选在T4 fibritin三聚体基序的C末端引入一段poly His标签用于蛋白的分离纯化。
根据本发明的一些具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列包括：编码SEQ IDNo. 22所示氨基酸序列的核苷酸序列。
根据本发明的一些更具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列如SEQ ID No. 32所示。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，所述哺乳动物细胞为人胚肾293细胞(human embryonic kidney293,HEK293）。利用人胚肾293细胞生产重组蛋白的系统已经建立了较长时间。与低等真核细胞表达系统或者原核表达系统相比，HEK293细胞表达系统能够指导新型冠状病毒Spike蛋白质的正确折叠、提供复杂的N型糖基化和准确的O型糖基化等多种翻译后加工修饰功能，因而表达的Spike重组蛋白在分子结构、理化特性和生物学功能方面最接近于新型冠状病毒感染人体后病人体内的Spike蛋白，因而HEK293细胞是新型冠状病毒Spike蛋白重组表达的最佳宿主。然而，现有技术中，HEK293表达系统的一个突出特点是蛋白表达水平较低。而本发明中惊奇地发现，采用本发明特定的信号肽在HEK293表达系统中分泌表达重组新型冠状病毒Spike蛋白，具有出乎意料的表达水平。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，所述表达载体可以为带有以下启动子的表达载体：EF-1α（human elongation factor-1α）启动子、hCMV（human cytomegalovirus）启动子、SV40（Simian vacuolating virus40）晚期启动子等。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法中，所述表达载体为pcDAN3.1(+)。
根据本发明的具体实施方案，本发明的制备重组新型冠状病毒Spike蛋白的方法，其还包括纯化所表达的重组新型冠状病毒Spike蛋白的过程。
另一方面，本发明还提供了一种用于制备重组新型冠状病毒Spike蛋白的表达载体，其为上述本发明的制备重组新型冠状病毒Spike蛋白的方法中所用到的含有编码重组新型冠状病毒Spike蛋白的多核苷酸的表达载体。
另一方面，本发明还提供了一种用于制备重组新型冠状病毒Spike蛋白的宿主细胞，其是由上述本发明所述的表达载体转染哺乳动物细胞得到的能分泌表达重组新型冠状病毒Spike蛋白的细胞。
本发明中的编码上述氨基酸序列的基因均可通过常规的合成方法进行制备。
综上所述，本发明提供了一种制备重组新型冠状病毒Spike蛋白的方法，其中采用特定的信号肽，使得新型冠状病毒Spike蛋白在HEK293细胞中高效分泌表达，降低生产成本；另一方面极大的满足了新型冠状病毒Spike蛋白作为抗病毒药物开发和新冠诊断原材料的大量需求。
附图说明
图1为表达载体pcDNA3.1图谱。
图2为重组质粒构建示意图。
图3显示不同信号肽效果对比结果。
具体实施方式
为了更清楚地理解本发明，现参照下列实施例及附图进一步描述本发明。实施例仅用于解释而不以任何方式限制本发明。实施例中未注明具体条件的实验方法为所属领域熟知的常规方法和常规条件，或按照制造商所建议的条件。
实施例1、重组表达载体的构建
本实施例中，筛选了10种信号肽，验证其应用于制备重组新型冠状病毒Spike蛋白（氨基酸序列如序列表SEQ ID No. 1所示）的分泌表达情况。
其中，新型冠状病毒Spike蛋白自身信号肽氨基酸序列如序列表SEQ ID No. 2所示。5种已知可以促进重组蛋白高分泌表达的信号肽（下文简称高表达信号肽），分别为Human IL-2信号肽、Mouse IgG kappa信号肽、Human tPA信号肽、流感病毒HA信号肽、HumanIgG kappa信号肽，其对应的氨基酸序列如序列表SEQ ID No. 3- SEQ ID No. 7所示。其余4种信号肽对应的氨基酸序列如序列表SEQ ID No. 8- SEQ ID No. 11所示。
Figure 801724DEST_PATH_IMAGE001
Figure 718864DEST_PATH_IMAGE002
首先，构建使用筛选信号肽(SEQ ID No. 2-11)的重组pHEK-S1~ pHEK-S10表达载体：由上海生物工程有限公司合成编码SEQ ID No. 14~ SEQ ID No. 23所示氨基酸序列的如SEQ ID No. 24~ SEQ ID No. 33的多核苷酸（SEQ ID No. 14~ SEQ ID No. 23所示氨基酸序列中，于Spike蛋白的C末端均引入一段T4 fibritin 三聚体基序用于提高Spike蛋白的三聚稳定性，T4 fibritin 三聚体基序氨基酸序列如SEQ ID No. 12所示； T4 fibritin三聚体基序的C末端均引入一段poly His标签用于蛋白的分离纯化，poly His标签氨基酸序列如SEQ ID No. 13所示），5’末端添加限制性内切酶BamH Ⅰ酶切位点、Kozak序列；3’末端添加终止密码子TAA和限制性内切酶Xho Ⅰ酶切位点（图2）。合成产物用限制性内切酶BamH Ⅰ和XhoⅠ双酶切，然后进行琼脂糖凝胶电泳，切胶回收目的片段。将目的片段分别与同样经限制性内切酶BamHⅠ和XhoⅠ双酶切的pcDNA3.1(+)连接，构建重组真核表达载体pHEK-S1/ pHEK-S2/ pHEK-S3/ pHEK-S4/ pHEK-S5/ pHEK-S6/ pHEK-S7/ pHEK-S8/ pHEK-S9/pHEK-S10，分别转化大肠杆菌E.coli DH5α，37℃培养16h后挑取单菌落进行质粒提取，将提取后的质粒用限制性内切酶BamH Ⅰ和Xho Ⅰ双酶切后进行琼脂糖凝胶电泳鉴定，将阳性克隆送上海生物工程有限公司进行测序，选择测序正确的克隆进行质粒的放大提取，提取后的质粒无菌过滤后，-20℃保存备用。表达载体pcDNA3.1图谱参见图1，重组质粒构建参见图2。
实施例2、重组表达载体转染HEK293细胞并表达新型冠状病毒Spike蛋白
使用PEI转染试剂将构建好的重组质粒pHEK-S1~ pHEK-S10分别转染HEK293细胞，进行新型冠状病毒Spike蛋白的重组表达。转染前一天将HEK293细胞按照1×106/mL传代，37℃培养。转染当天进行计数，调整细胞密度为2×106/mL，活率95%以上。
根据细胞密度计算转染复合物中各组分的用量:质粒剂量与细胞的数量对应关系为1×106cells对应1 ug质粒，PEI剂量与质粒剂量的对应关系为 PEI的质量为DNA质量的3倍。
根据上述计算得到的组分用量配置转染复合物：将50 ug浓度为200 ug/mL的重组质粒加入CD 293 TGE培养基至0.5 mL（A液），150 ug浓度为1 mg/mL的 PEI转染试剂加入CD293 TGE培养基至0.5ml（B液），分别混匀后，将B液缓慢加入至A液中，混匀室温静置10-15min后，将转染混合液缓慢滴加入10 mL HEK293细胞中。37℃，5％ CO2 ，135 rpm培养96 h后，收集上清。
实施例3、Spike蛋白的分离纯化
使用His-tag亲和层析的方法对Spike重组蛋白进行分离纯化，比较10种信号肽对提高Spike蛋白分泌表达的效果。 1000×g，30 min离心收集上述细胞培养上清，将上清过滤后分别结合预先用PBS缓冲液平衡的GE HisTrap excel层析柱，然后分别用含有30/100/500mM咪唑的PBS洗脱目标蛋白，每个浓度洗脱体积为5 CV，收集蛋白洗脱液，并进行SDS-PAGE分析。
实施例4、本发明信号肽对验证蛋白分泌表达效果分析
同等条件下，分别使用包含本发明信号肽在内的9中已知高表达信号肽和新型冠状病毒Spike蛋白自身信号肽比较验证新型冠状病毒Spike蛋白在HEK293细胞中的分泌表达水平，SDS-PAGE结果（图3）显示：
1）相比Spike蛋白自身信号肽，本发明的信号肽可使得Spike蛋白的分泌表达量提高6倍以上；
2）相比Spike蛋白自身信号肽，其他8种信号肽可使得Spike蛋白提高1-4倍。
实验数据表明, 使用本发明信号肽与Spike蛋白自身信号肽及其他8种高表达信号肽相比，在HEK293细胞中的Spike蛋白表达量获得大幅提升。使用本发明信号肽，相比蛋白自身信号肽，Spike蛋白表达量提高6倍以上。
上述结果表明，本发明的信号肽特别适用于新型冠状病毒Spike蛋白在HEK293细胞中的分泌表达，可大幅降低生产成本，满足Spike蛋白作为抗毒药物开发和新冠诊断原材料的大量需求。
SEQUENCE LISTING
<110> 北京百普赛斯生物科技股份有限公司
<120> 制备重组新型冠状病毒Spike蛋白的方法
<130> GAI20CN7739
<160> 33
<170> PatentIn version 3.5
<210> 1
<211> 1197
<212> PRT
<213> SARS-CoV-2
<400> 1
Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser
1 5 10 15
Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val
20 25 30
Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr
35 40 45
Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe
50 55 60
Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr
65 70 75 80
Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp
85 90 95
Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val
100 105 110
Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val
115 120 125
Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val
130 135 140
Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe
145 150 155 160
Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu
165 170 175
Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His
180 185 190
Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu
195 200 205
Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln
210 215 220
Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser
225 230 235 240
Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln
245 250 255
Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp
260 265 270
Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu
275 280 285
Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg
290 295 300
Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu
305 310 315 320
Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr
325 330 335
Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val
340 345 350
Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser
355 360 365
Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser
370 375 380
Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr
385 390 395 400
Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly
405 410 415
Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly
420 425 430
Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro
435 440 445
Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro
450 455 460
Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr
465 470 475 480
Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val
485 490 495
Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro
500 505 510
Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe
515 520 525
Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe
530 535 540
Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala
545 550 555 560
Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser
565 570 575
Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln
580 585 590
Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala
595 600 605
Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly
610 615 620
Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His
625 630 635 640
Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys
645 650 655
Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala Ala Ala Ser Val
660 665 670
Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn
675 680 685
Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr
690 695 700
Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser
705 710 715 720
Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn
725 730 735
Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu
740 745 750
Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala
755 760 765
Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly
770 775 780
Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg
785 790 795 800
Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala
805 810 815
Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg
820 825 830
Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro
835 840 845
Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala
850 855 860
Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln
865 870 875 880
Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val
885 890 895
Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe
900 905 910
Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser
915 920 925
Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu
930 935 940
Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser
945 950 955 960
Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val
965 970 975
Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr
980 985 990
Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
995 1000 1005
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1010 1015 1020
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1025 1030 1035
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1040 1045 1050
Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp
1055 1060 1065
Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
1070 1075 1080
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile
1085 1090 1095
Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val
1100 1105 1110
Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu
1115 1120 1125
Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His
1130 1135 1140
Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala
1145 1150 1155
Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu Val
1160 1165 1170
Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu Gly
1175 1180 1185
Lys Tyr Glu Gln Tyr Ile Lys Trp Pro
1190 1195
<210> 2
<211> 15
<212> PRT
<213> SARS-CoV-2
<400> 2
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys
1 5 10 15
<210> 3
<211> 20
<212> PRT
<213> Homo sapiens
<400> 3
Met Tyr Arg Met Gln Leu Leu Ser Cys Ile Ala Leu Ser Leu Ala Leu
1 5 10 15
Val Thr Asn Ser
20
<210> 4
<211> 20
<212> PRT
<213> Mouse
<400> 4
Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro
1 5 10 15
Gly Ser Thr Gly
20
<210> 5
<211> 23
<212> PRT
<213> Homo sapiens
<400> 5
Met Asp Ala Met Lys Arg Gly Leu Cys Cys Val Leu Leu Leu Cys Gly
1 5 10 15
Ala Val Phe Val Ser Pro Ser
20
<210> 6
<211> 16
<212> PRT
<213> Influenza virus
<400> 6
Met Lys Thr Ile Ile Ala Leu Ser Tyr Ile Phe Cys Leu Val Leu Gly
1 5 10 15
<210> 7
<211> 20
<212> PRT
<213> Homo sapiens
<400> 7
Met Glu Leu Gly Leu Ser Trp Val Phe Leu Val Ala Ile Leu Glu Gly
1 5 10 15
Val Gln Cys Glu
20
<210> 8
<211> 24
<212> PRT
<213> Homo sapiens
<400> 8
Met Pro Met Gly Ser Leu Gln Pro Leu Ala Thr Leu Tyr Leu Leu Gly
1 5 10 15
Met Leu Val Ala Ser Cys Leu Gly
20
<210> 9
<211> 30
<212> PRT
<213> Bovine
<400> 9
Met Asp Ser Lys Gly Ser Ser Gln Lys Gly Ser Arg Leu Leu Leu Leu
1 5 10 15
Leu Val Val Ser Asn Leu Leu Leu Cys Gln Gly Val Val Ser
20 25 30
<210> 10
<211> 26
<212> PRT
<213> Homo sapiens
<400> 10
Met Ala Ala Pro Ala Leu Ser Trp Arg Leu Pro Leu Leu Ile Leu Leu
1 5 10 15
Leu Pro Leu Ala Thr Ser Trp Ala Ser Ala
20 25
<210> 11
<211> 17
<212> PRT
<213> Homo sapiens
<400> 11
Met Arg Ala Trp Ile Phe Phe Leu Leu Cys Leu Ala Gly Arg Ala Leu
1 5 10 15
Ala
<210> 12
<211> 26
<212> PRT
<213> Artificial Sequence
<220>
<223> T4 fibritin三聚体基序
<400> 12
Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp
1 5 10 15
Gly Glu Trp Val Leu Leu Ser Thr Phe Leu
20 25
<210> 13
<211> 6
<212> PRT
<213> Artificial Sequence
<220>
<223> poly His标签
<400> 13
His His His His His His
1 5
<210> 14
<211> 1244
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 14
Met Phe Val Phe Leu Val Leu Leu Pro Leu Val Ser Ser Gln Cys Val
1 5 10 15
Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe
20 25 30
Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu
35 40 45
His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp
50 55 60
Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp
65 70 75 80
Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu
85 90 95
Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser
100 105 110
Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile
115 120 125
Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr
130 135 140
Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr
145 150 155 160
Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu
165 170 175
Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe
180 185 190
Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr
195 200 205
Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu
210 215 220
Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr
225 230 235 240
Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser
245 250 255
Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro
260 265 270
Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala
275 280 285
Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys
290 295 300
Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val
305 310 315 320
Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys
325 330 335
Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala
340 345 350
Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu
355 360 365
Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro
370 375 380
Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe
385 390 395 400
Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly
405 410 415
Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys
420 425 430
Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn
435 440 445
Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe
450 455 460
Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys
465 470 475 480
Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly
485 490 495
Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val
500 505 510
Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys
515 520 525
Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn
530 535 540
Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu
545 550 555 560
Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val
565 570 575
Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe
580 585 590
Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val
595 600 605
Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile
610 615 620
His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser
625 630 635 640
Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val
645 650 655
Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala
660 665 670
Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala Ala Ala Ser Val Ala
675 680 685
Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser
690 695 700
Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile
705 710 715 720
Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val
725 730 735
Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu
740 745 750
Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr
755 760 765
Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln
770 775 780
Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe
785 790 795 800
Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser
805 810 815
Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly
820 825 830
Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp
835 840 845
Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu
850 855 860
Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly
865 870 875 880
Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile
885 890 895
Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr
900 905 910
Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn
915 920 925
Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala
930 935 940
Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn
945 950 955 960
Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val
965 970 975
Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln
980 985 990
Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
995 1000 1005
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1010 1015 1020
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1025 1030 1035
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1040 1045 1050
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1055 1060 1065
Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp
1070 1075 1080
Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
1085 1090 1095
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile
1100 1105 1110
Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val
1115 1120 1125
Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu
1130 1135 1140
Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His
1145 1150 1155
Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala
1160 1165 1170
Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu Val
1175 1180 1185
Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu Gly
1190 1195 1200
Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr Ile Pro Glu Ala Pro
1205 1210 1215
Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu
1220 1225 1230
Leu Ser Thr Phe Leu His His His His His His
1235 1240
<210> 15
<211> 1249
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 15
Met Tyr Arg Met Gln Leu Leu Ser Cys Ile Ala Leu Ser Leu Ala Leu
1 5 10 15
Val Thr Asn Ser Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala
20 25 30
Tyr Thr Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe
35 40 45
Arg Ser Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe
50 55 60
Ser Asn Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly
65 70 75 80
Thr Lys Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr
85 90 95
Phe Ala Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly
100 105 110
Thr Thr Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala
115 120 125
Thr Asn Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro
130 135 140
Phe Leu Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser
145 150 155 160
Glu Phe Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val
165 170 175
Ser Gln Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys
180 185 190
Asn Leu Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile
195 200 205
Tyr Ser Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly
210 215 220
Phe Ser Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile
225 230 235 240
Thr Arg Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro
245 250 255
Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val
260 265 270
Gly Tyr Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly
275 280 285
Thr Ile Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr
290 295 300
Lys Cys Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr
305 310 315 320
Ser Asn Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn
325 330 335
Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe
340 345 350
Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala
355 360 365
Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys
370 375 380
Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val
385 390 395 400
Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala
405 410 415
Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp
420 425 430
Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser
435 440 445
Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser
450 455 460
Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala
465 470 475 480
Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro
485 490 495
Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro
500 505 510
Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr
515 520 525
Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val
530 535 540
Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser
545 550 555 560
Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp
565 570 575
Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile
580 585 590
Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn
595 600 605
Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
610 615 620
Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val
625 630 635 640
Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile
645 650 655
Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly
660 665 670
Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala
675 680 685
Ala Ala Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu
690 695 700
Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro
705 710 715 720
Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met
725 730 735
Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr
740 745 750
Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu
755 760 765
Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln
770 775 780
Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys
785 790 795 800
Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys
805 810 815
Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr
820 825 830
Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp
835 840 845
Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr
850 855 860
Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser
865 870 875 880
Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly
885 890 895
Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn
900 905 910
Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile
915 920 925
Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser
930 935 940
Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn
945 950 955 960
Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly
965 970 975
Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val
980 985 990
Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser
995 1000 1005
Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
1010 1015 1020
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1025 1030 1035
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1040 1045 1050
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1055 1060 1065
His Val Thr Tyr Val Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro
1070 1075 1080
Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val
1085 1090 1095
Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe
1100 1105 1110
Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly
1115 1120 1125
Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp
1130 1135 1140
Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys
1145 1150 1155
Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile
1160 1165 1170
Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp
1175 1180 1185
Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp
1190 1195 1200
Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr
1205 1210 1215
Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp
1220 1225 1230
Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His His His
1235 1240 1245
His
<210> 16
<211> 1249
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 16
Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro
1 5 10 15
Gly Ser Thr Gly Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala
20 25 30
Tyr Thr Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe
35 40 45
Arg Ser Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe
50 55 60
Ser Asn Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly
65 70 75 80
Thr Lys Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr
85 90 95
Phe Ala Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly
100 105 110
Thr Thr Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala
115 120 125
Thr Asn Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro
130 135 140
Phe Leu Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser
145 150 155 160
Glu Phe Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val
165 170 175
Ser Gln Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys
180 185 190
Asn Leu Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile
195 200 205
Tyr Ser Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly
210 215 220
Phe Ser Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile
225 230 235 240
Thr Arg Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro
245 250 255
Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val
260 265 270
Gly Tyr Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly
275 280 285
Thr Ile Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr
290 295 300
Lys Cys Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr
305 310 315 320
Ser Asn Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn
325 330 335
Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe
340 345 350
Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala
355 360 365
Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys
370 375 380
Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val
385 390 395 400
Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala
405 410 415
Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp
420 425 430
Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser
435 440 445
Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser
450 455 460
Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala
465 470 475 480
Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro
485 490 495
Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro
500 505 510
Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr
515 520 525
Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val
530 535 540
Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser
545 550 555 560
Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp
565 570 575
Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile
580 585 590
Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn
595 600 605
Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
610 615 620
Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val
625 630 635 640
Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile
645 650 655
Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly
660 665 670
Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala
675 680 685
Ala Ala Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu
690 695 700
Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro
705 710 715 720
Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met
725 730 735
Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr
740 745 750
Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu
755 760 765
Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln
770 775 780
Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys
785 790 795 800
Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys
805 810 815
Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr
820 825 830
Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp
835 840 845
Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr
850 855 860
Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser
865 870 875 880
Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly
885 890 895
Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn
900 905 910
Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile
915 920 925
Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser
930 935 940
Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn
945 950 955 960
Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly
965 970 975
Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val
980 985 990
Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser
995 1000 1005
Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
1010 1015 1020
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1025 1030 1035
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1040 1045 1050
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1055 1060 1065
His Val Thr Tyr Val Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro
1070 1075 1080
Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val
1085 1090 1095
Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe
1100 1105 1110
Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly
1115 1120 1125
Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp
1130 1135 1140
Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys
1145 1150 1155
Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile
1160 1165 1170
Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp
1175 1180 1185
Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp
1190 1195 1200
Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr
1205 1210 1215
Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp
1220 1225 1230
Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His His His
1235 1240 1245
His
<210> 17
<211> 1252
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 17
Met Asp Ala Met Lys Arg Gly Leu Cys Cys Val Leu Leu Leu Cys Gly
1 5 10 15
Ala Val Phe Val Ser Pro Ser Val Asn Leu Thr Thr Arg Thr Gln Leu
20 25 30
Pro Pro Ala Tyr Thr Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp
35 40 45
Lys Val Phe Arg Ser Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu
50 55 60
Pro Phe Phe Ser Asn Val Thr Trp Phe His Ala Ile His Val Ser Gly
65 70 75 80
Thr Asn Gly Thr Lys Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp
85 90 95
Gly Val Tyr Phe Ala Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp
100 105 110
Ile Phe Gly Thr Thr Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val
115 120 125
Asn Asn Ala Thr Asn Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys
130 135 140
Asn Asp Pro Phe Leu Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp
145 150 155 160
Met Glu Ser Glu Phe Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe
165 170 175
Glu Tyr Val Ser Gln Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly
180 185 190
Asn Phe Lys Asn Leu Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr
195 200 205
Phe Lys Ile Tyr Ser Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu
210 215 220
Pro Gln Gly Phe Ser Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly
225 230 235 240
Ile Asn Ile Thr Arg Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr
245 250 255
Leu Thr Pro Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala
260 265 270
Tyr Tyr Val Gly Tyr Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn
275 280 285
Glu Asn Gly Thr Ile Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu
290 295 300
Ser Glu Thr Lys Cys Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile
305 310 315 320
Tyr Gln Thr Ser Asn Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg
325 330 335
Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala
340 345 350
Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn
355 360 365
Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr
370 375 380
Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe
385 390 395 400
Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg
405 410 415
Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys
420 425 430
Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn
435 440 445
Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe
450 455 460
Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile
465 470 475 480
Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys
485 490 495
Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly
500 505 510
Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala
515 520 525
Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn
530 535 540
Lys Cys Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu
545 550 555 560
Thr Glu Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp
565 570 575
Ile Ala Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile
580 585 590
Leu Asp Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro
595 600 605
Gly Thr Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn
610 615 620
Cys Thr Glu Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr
625 630 635 640
Trp Arg Val Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly
645 650 655
Cys Leu Ile Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile
660 665 670
Pro Ile Gly Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser
675 680 685
Pro Arg Ala Ala Ala Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr
690 695 700
Met Ser Leu Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile
705 710 715 720
Ala Ile Pro Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro
725 730 735
Val Ser Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly
740 745 750
Asp Ser Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys
755 760 765
Thr Gln Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys
770 775 780
Asn Thr Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro
785 790 795 800
Pro Ile Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp
805 810 815
Pro Ser Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn
820 825 830
Lys Val Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys
835 840 845
Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn
850 855 860
Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln
865 870 875 880
Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe
885 890 895
Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr
900 905 910
Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln
915 920 925
Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp
930 935 940
Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val
945 950 955 960
Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser
965 970 975
Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu
980 985 990
Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg
995 1000 1005
Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala
1010 1015 1020
Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser
1025 1030 1035
Glu Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys
1040 1045 1050
Gly Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val
1055 1060 1065
Val Phe Leu His Val Thr Tyr Val Pro Ala Glu Lys Asn Phe Thr
1070 1075 1080
Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg
1085 1090 1095
Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln
1100 1105 1110
Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe
1115 1120 1125
Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr
1130 1135 1140
Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu
1145 1150 1155
Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu
1160 1165 1170
Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys
1175 1180 1185
Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser
1190 1195 1200
Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys
1205 1210 1215
Trp Pro Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val
1220 1225 1230
Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His
1235 1240 1245
His His His His
1250
<210> 18
<211> 1245
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 18
Met Lys Thr Ile Ile Ala Leu Ser Tyr Ile Phe Cys Leu Val Leu Gly
1 5 10 15
Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser
20 25 30
Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val
35 40 45
Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr
50 55 60
Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe
65 70 75 80
Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr
85 90 95
Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp
100 105 110
Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val
115 120 125
Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val
130 135 140
Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val
145 150 155 160
Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe
165 170 175
Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu
180 185 190
Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His
195 200 205
Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu
210 215 220
Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln
225 230 235 240
Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser
245 250 255
Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln
260 265 270
Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp
275 280 285
Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu
290 295 300
Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg
305 310 315 320
Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu
325 330 335
Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr
340 345 350
Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val
355 360 365
Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser
370 375 380
Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser
385 390 395 400
Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr
405 410 415
Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly
420 425 430
Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly
435 440 445
Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro
450 455 460
Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro
465 470 475 480
Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr
485 490 495
Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val
500 505 510
Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro
515 520 525
Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe
530 535 540
Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe
545 550 555 560
Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala
565 570 575
Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser
580 585 590
Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln
595 600 605
Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala
610 615 620
Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly
625 630 635 640
Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His
645 650 655
Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys
660 665 670
Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala Ala Ala Ser Val
675 680 685
Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn
690 695 700
Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr
705 710 715 720
Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser
725 730 735
Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn
740 745 750
Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu
755 760 765
Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala
770 775 780
Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly
785 790 795 800
Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg
805 810 815
Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala
820 825 830
Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg
835 840 845
Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro
850 855 860
Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala
865 870 875 880
Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln
885 890 895
Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val
900 905 910
Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe
915 920 925
Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser
930 935 940
Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu
945 950 955 960
Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser
965 970 975
Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val
980 985 990
Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr
995 1000 1005
Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala
1010 1015 1020
Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser
1025 1030 1035
Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe
1040 1045 1050
Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr
1055 1060 1065
Val Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His
1070 1075 1080
Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn
1085 1090 1095
Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln
1100 1105 1110
Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val
1115 1120 1125
Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro
1130 1135 1140
Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn
1145 1150 1155
His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn
1160 1165 1170
Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu
1175 1180 1185
Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu
1190 1195 1200
Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr Ile Pro Glu Ala
1205 1210 1215
Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val
1220 1225 1230
Leu Leu Ser Thr Phe Leu His His His His His His
1235 1240 1245
<210> 19
<211> 1249
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 19
Met Glu Leu Gly Leu Ser Trp Val Phe Leu Val Ala Ile Leu Glu Gly
1 5 10 15
Val Gln Cys Glu Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala
20 25 30
Tyr Thr Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe
35 40 45
Arg Ser Ser Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe
50 55 60
Ser Asn Val Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly
65 70 75 80
Thr Lys Arg Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr
85 90 95
Phe Ala Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly
100 105 110
Thr Thr Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala
115 120 125
Thr Asn Val Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro
130 135 140
Phe Leu Gly Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser
145 150 155 160
Glu Phe Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val
165 170 175
Ser Gln Pro Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys
180 185 190
Asn Leu Arg Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile
195 200 205
Tyr Ser Lys His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly
210 215 220
Phe Ser Ala Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile
225 230 235 240
Thr Arg Phe Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro
245 250 255
Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val
260 265 270
Gly Tyr Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly
275 280 285
Thr Ile Thr Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr
290 295 300
Lys Cys Thr Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr
305 310 315 320
Ser Asn Phe Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn
325 330 335
Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe
340 345 350
Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala
355 360 365
Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys
370 375 380
Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val
385 390 395 400
Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala
405 410 415
Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp
420 425 430
Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser
435 440 445
Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser
450 455 460
Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala
465 470 475 480
Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro
485 490 495
Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro
500 505 510
Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr
515 520 525
Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val
530 535 540
Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser
545 550 555 560
Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp
565 570 575
Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile
580 585 590
Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn
595 600 605
Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu
610 615 620
Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val
625 630 635 640
Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile
645 650 655
Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly
660 665 670
Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala
675 680 685
Ala Ala Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu
690 695 700
Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro
705 710 715 720
Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met
725 730 735
Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr
740 745 750
Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu
755 760 765
Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln
770 775 780
Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys
785 790 795 800
Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys
805 810 815
Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr
820 825 830
Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp
835 840 845
Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr
850 855 860
Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser
865 870 875 880
Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly
885 890 895
Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn
900 905 910
Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile
915 920 925
Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser
930 935 940
Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn
945 950 955 960
Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly
965 970 975
Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val
980 985 990
Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser
995 1000 1005
Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile
1010 1015 1020
Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val
1025 1030 1035
Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His
1040 1045 1050
Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu
1055 1060 1065
His Val Thr Tyr Val Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro
1070 1075 1080
Ala Ile Cys His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val
1085 1090 1095
Phe Val Ser Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe
1100 1105 1110
Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly
1115 1120 1125
Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp
1130 1135 1140
Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys
1145 1150 1155
Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile
1160 1165 1170
Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp
1175 1180 1185
Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp
1190 1195 1200
Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr
1205 1210 1215
Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp
1220 1225 1230
Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His His His His His
1235 1240 1245
His
<210> 20
<211> 1253
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 20
Met Pro Met Gly Ser Leu Gln Pro Leu Ala Thr Leu Tyr Leu Leu Gly
1 5 10 15
Met Leu Val Ala Ser Cys Leu Gly Val Asn Leu Thr Thr Arg Thr Gln
20 25 30
Leu Pro Pro Ala Tyr Thr Asn Ser Phe Thr Arg Gly Val Tyr Tyr Pro
35 40 45
Asp Lys Val Phe Arg Ser Ser Val Leu His Ser Thr Gln Asp Leu Phe
50 55 60
Leu Pro Phe Phe Ser Asn Val Thr Trp Phe His Ala Ile His Val Ser
65 70 75 80
Gly Thr Asn Gly Thr Lys Arg Phe Asp Asn Pro Val Leu Pro Phe Asn
85 90 95
Asp Gly Val Tyr Phe Ala Ser Thr Glu Lys Ser Asn Ile Ile Arg Gly
100 105 110
Trp Ile Phe Gly Thr Thr Leu Asp Ser Lys Thr Gln Ser Leu Leu Ile
115 120 125
Val Asn Asn Ala Thr Asn Val Val Ile Lys Val Cys Glu Phe Gln Phe
130 135 140
Cys Asn Asp Pro Phe Leu Gly Val Tyr Tyr His Lys Asn Asn Lys Ser
145 150 155 160
Trp Met Glu Ser Glu Phe Arg Val Tyr Ser Ser Ala Asn Asn Cys Thr
165 170 175
Phe Glu Tyr Val Ser Gln Pro Phe Leu Met Asp Leu Glu Gly Lys Gln
180 185 190
Gly Asn Phe Lys Asn Leu Arg Glu Phe Val Phe Lys Asn Ile Asp Gly
195 200 205
Tyr Phe Lys Ile Tyr Ser Lys His Thr Pro Ile Asn Leu Val Arg Asp
210 215 220
Leu Pro Gln Gly Phe Ser Ala Leu Glu Pro Leu Val Asp Leu Pro Ile
225 230 235 240
Gly Ile Asn Ile Thr Arg Phe Gln Thr Leu Leu Ala Leu His Arg Ser
245 250 255
Tyr Leu Thr Pro Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly Ala Ala
260 265 270
Ala Tyr Tyr Val Gly Tyr Leu Gln Pro Arg Thr Phe Leu Leu Lys Tyr
275 280 285
Asn Glu Asn Gly Thr Ile Thr Asp Ala Val Asp Cys Ala Leu Asp Pro
290 295 300
Leu Ser Glu Thr Lys Cys Thr Leu Lys Ser Phe Thr Val Glu Lys Gly
305 310 315 320
Ile Tyr Gln Thr Ser Asn Phe Arg Val Gln Pro Thr Glu Ser Ile Val
325 330 335
Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val Phe Asn
340 345 350
Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg Ile Ser
355 360 365
Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser Phe Ser
370 375 380
Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp Leu Cys
385 390 395 400
Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp Glu Val
405 410 415
Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr Asn Tyr
420 425 430
Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn Ser Asn
435 440 445
Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu
450 455 460
Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser Thr Glu
465 470 475 480
Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly Phe Asn
485 490 495
Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn Gly Val
500 505 510
Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu Leu His
515 520 525
Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu Val Lys
530 535 540
Asn Lys Cys Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr Gly Val
545 550 555 560
Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe Gly Arg
565 570 575
Asp Ile Ala Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr Leu Glu
580 585 590
Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val Ile Thr
595 600 605
Pro Gly Thr Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln Asp Val
610 615 620
Asn Cys Thr Glu Val Pro Val Ala Ile His Ala Asp Gln Leu Thr Pro
625 630 635 640
Thr Trp Arg Val Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr Arg Ala
645 650 655
Gly Cys Leu Ile Gly Ala Glu His Val Asn Asn Ser Tyr Glu Cys Asp
660 665 670
Ile Pro Ile Gly Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln Thr Asn
675 680 685
Ser Pro Arg Ala Ala Ala Ser Val Ala Ser Gln Ser Ile Ile Ala Tyr
690 695 700
Thr Met Ser Leu Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn Asn Ser
705 710 715 720
Ile Ala Ile Pro Thr Asn Phe Thr Ile Ser Val Thr Thr Glu Ile Leu
725 730 735
Pro Val Ser Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr Ile Cys
740 745 750
Gly Asp Ser Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly Ser Phe
755 760 765
Cys Thr Gln Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu Gln Asp
770 775 780
Lys Asn Thr Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr Lys Thr
785 790 795 800
Pro Pro Ile Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile Leu Pro
805 810 815
Asp Pro Ser Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu Leu Phe
820 825 830
Asn Lys Val Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr Gly Asp
835 840 845
Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln Lys Phe
850 855 860
Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met Ile Ala
865 870 875 880
Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly Trp Thr
885 890 895
Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln Met Ala
900 905 910
Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr Glu Asn
915 920 925
Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys Ile Gln
930 935 940
Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln Asp Val
945 950 955 960
Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln Leu Ser
965 970 975
Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu Ser Arg
980 985 990
Leu Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile Thr Gly
995 1000 1005
Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu Ile Arg
1010 1015 1020
Ala Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr Lys Met
1025 1030 1035
Ser Glu Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe Cys Gly
1040 1045 1050
Lys Gly Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro His Gly
1055 1060 1065
Val Val Phe Leu His Val Thr Tyr Val Pro Ala Glu Lys Asn Phe
1070 1075 1080
Thr Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His Phe Pro
1085 1090 1095
Arg Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe Val Thr
1100 1105 1110
Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp Asn Thr
1115 1120 1125
Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val Asn Asn
1130 1135 1140
Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe Lys Glu
1145 1150 1155
Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp Val Asp
1160 1165 1170
Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn Ile Gln
1175 1180 1185
Lys Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu Asn Glu
1190 1195 1200
Ser Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln Tyr Ile
1205 1210 1215
Lys Trp Pro Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln Ala Tyr
1220 1225 1230
Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe Leu His
1235 1240 1245
His His His His His
1250
<210> 21
<211> 1259
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 21
Met Asp Ser Lys Gly Ser Ser Gln Lys Gly Ser Arg Leu Leu Leu Leu
1 5 10 15
Leu Val Val Ser Asn Leu Leu Leu Cys Gln Gly Val Val Ser Val Asn
20 25 30
Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe Thr
35 40 45
Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu His
50 55 60
Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp Phe
65 70 75 80
His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp Asn
85 90 95
Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu Lys
100 105 110
Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser Lys
115 120 125
Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile Lys
130 135 140
Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr Tyr
145 150 155 160
His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr Ser
165 170 175
Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu Met
180 185 190
Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe Val
195 200 205
Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr Pro
210 215 220
Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu Pro
225 230 235 240
Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr Leu
245 250 255
Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser Gly
260 265 270
Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro Arg
275 280 285
Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala Val
290 295 300
Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys Ser
305 310 315 320
Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val Gln
325 330 335
Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro
340 345 350
Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp
355 360 365
Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr
370 375 380
Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr
385 390 395 400
Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val
405 410 415
Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys
420 425 430
Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val
435 440 445
Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr
450 455 460
Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu
465 470 475 480
Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn
485 490 495
Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe
500 505 510
Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu
515 520 525
Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys
530 535 540
Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn Gly
545 550 555 560
Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro
565 570 575
Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val Arg
580 585 590
Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly
595 600 605
Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val Ala
610 615 620
Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile His
625 630 635 640
Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser Asn
645 650 655
Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val Asn
660 665 670
Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala Ser
675 680 685
Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala Ala Ala Ser Val Ala Ser
690 695 700
Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser Val
705 710 715 720
Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile Ser
725 730 735
Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val Asp
740 745 750
Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu Leu
755 760 765
Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr Gly
770 775 780
Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln Val
785 790 795 800
Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe Asn
805 810 815
Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser Phe
820 825 830
Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly Phe
835 840 845
Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu
850 855 860
Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu Leu
865 870 875 880
Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr
885 890 895
Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro
900 905 910
Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln
915 920 925
Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser
930 935 940
Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu
945 950 955 960
Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn Thr
965 970 975
Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val Leu
980 985 990
Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln Ile
995 1000 1005
Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val
1010 1015 1020
Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn
1025 1030 1035
Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys
1040 1045 1050
Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro
1055 1060 1065
Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr Tyr Val
1070 1075 1080
Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp
1085 1090 1095
Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly
1100 1105 1110
Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile
1115 1120 1125
Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val
1130 1135 1140
Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu
1145 1150 1155
Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His
1160 1165 1170
Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala
1175 1180 1185
Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu Val
1190 1195 1200
Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu Gly
1205 1210 1215
Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr Ile Pro Glu Ala Pro
1220 1225 1230
Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu
1235 1240 1245
Leu Ser Thr Phe Leu His His His His His His
1250 1255
<210> 22
<211> 1255
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 22
Met Ala Ala Pro Ala Leu Ser Trp Arg Leu Pro Leu Leu Ile Leu Leu
1 5 10 15
Leu Pro Leu Ala Thr Ser Trp Ala Ser Ala Val Asn Leu Thr Thr Arg
20 25 30
Thr Gln Leu Pro Pro Ala Tyr Thr Asn Ser Phe Thr Arg Gly Val Tyr
35 40 45
Tyr Pro Asp Lys Val Phe Arg Ser Ser Val Leu His Ser Thr Gln Asp
50 55 60
Leu Phe Leu Pro Phe Phe Ser Asn Val Thr Trp Phe His Ala Ile His
65 70 75 80
Val Ser Gly Thr Asn Gly Thr Lys Arg Phe Asp Asn Pro Val Leu Pro
85 90 95
Phe Asn Asp Gly Val Tyr Phe Ala Ser Thr Glu Lys Ser Asn Ile Ile
100 105 110
Arg Gly Trp Ile Phe Gly Thr Thr Leu Asp Ser Lys Thr Gln Ser Leu
115 120 125
Leu Ile Val Asn Asn Ala Thr Asn Val Val Ile Lys Val Cys Glu Phe
130 135 140
Gln Phe Cys Asn Asp Pro Phe Leu Gly Val Tyr Tyr His Lys Asn Asn
145 150 155 160
Lys Ser Trp Met Glu Ser Glu Phe Arg Val Tyr Ser Ser Ala Asn Asn
165 170 175
Cys Thr Phe Glu Tyr Val Ser Gln Pro Phe Leu Met Asp Leu Glu Gly
180 185 190
Lys Gln Gly Asn Phe Lys Asn Leu Arg Glu Phe Val Phe Lys Asn Ile
195 200 205
Asp Gly Tyr Phe Lys Ile Tyr Ser Lys His Thr Pro Ile Asn Leu Val
210 215 220
Arg Asp Leu Pro Gln Gly Phe Ser Ala Leu Glu Pro Leu Val Asp Leu
225 230 235 240
Pro Ile Gly Ile Asn Ile Thr Arg Phe Gln Thr Leu Leu Ala Leu His
245 250 255
Arg Ser Tyr Leu Thr Pro Gly Asp Ser Ser Ser Gly Trp Thr Ala Gly
260 265 270
Ala Ala Ala Tyr Tyr Val Gly Tyr Leu Gln Pro Arg Thr Phe Leu Leu
275 280 285
Lys Tyr Asn Glu Asn Gly Thr Ile Thr Asp Ala Val Asp Cys Ala Leu
290 295 300
Asp Pro Leu Ser Glu Thr Lys Cys Thr Leu Lys Ser Phe Thr Val Glu
305 310 315 320
Lys Gly Ile Tyr Gln Thr Ser Asn Phe Arg Val Gln Pro Thr Glu Ser
325 330 335
Ile Val Arg Phe Pro Asn Ile Thr Asn Leu Cys Pro Phe Gly Glu Val
340 345 350
Phe Asn Ala Thr Arg Phe Ala Ser Val Tyr Ala Trp Asn Arg Lys Arg
355 360 365
Ile Ser Asn Cys Val Ala Asp Tyr Ser Val Leu Tyr Asn Ser Ala Ser
370 375 380
Phe Ser Thr Phe Lys Cys Tyr Gly Val Ser Pro Thr Lys Leu Asn Asp
385 390 395 400
Leu Cys Phe Thr Asn Val Tyr Ala Asp Ser Phe Val Ile Arg Gly Asp
405 410 415
Glu Val Arg Gln Ile Ala Pro Gly Gln Thr Gly Lys Ile Ala Asp Tyr
420 425 430
Asn Tyr Lys Leu Pro Asp Asp Phe Thr Gly Cys Val Ile Ala Trp Asn
435 440 445
Ser Asn Asn Leu Asp Ser Lys Val Gly Gly Asn Tyr Asn Tyr Leu Tyr
450 455 460
Arg Leu Phe Arg Lys Ser Asn Leu Lys Pro Phe Glu Arg Asp Ile Ser
465 470 475 480
Thr Glu Ile Tyr Gln Ala Gly Ser Thr Pro Cys Asn Gly Val Glu Gly
485 490 495
Phe Asn Cys Tyr Phe Pro Leu Gln Ser Tyr Gly Phe Gln Pro Thr Asn
500 505 510
Gly Val Gly Tyr Gln Pro Tyr Arg Val Val Val Leu Ser Phe Glu Leu
515 520 525
Leu His Ala Pro Ala Thr Val Cys Gly Pro Lys Lys Ser Thr Asn Leu
530 535 540
Val Lys Asn Lys Cys Val Asn Phe Asn Phe Asn Gly Leu Thr Gly Thr
545 550 555 560
Gly Val Leu Thr Glu Ser Asn Lys Lys Phe Leu Pro Phe Gln Gln Phe
565 570 575
Gly Arg Asp Ile Ala Asp Thr Thr Asp Ala Val Arg Asp Pro Gln Thr
580 585 590
Leu Glu Ile Leu Asp Ile Thr Pro Cys Ser Phe Gly Gly Val Ser Val
595 600 605
Ile Thr Pro Gly Thr Asn Thr Ser Asn Gln Val Ala Val Leu Tyr Gln
610 615 620
Asp Val Asn Cys Thr Glu Val Pro Val Ala Ile His Ala Asp Gln Leu
625 630 635 640
Thr Pro Thr Trp Arg Val Tyr Ser Thr Gly Ser Asn Val Phe Gln Thr
645 650 655
Arg Ala Gly Cys Leu Ile Gly Ala Glu His Val Asn Asn Ser Tyr Glu
660 665 670
Cys Asp Ile Pro Ile Gly Ala Gly Ile Cys Ala Ser Tyr Gln Thr Gln
675 680 685
Thr Asn Ser Pro Arg Ala Ala Ala Ser Val Ala Ser Gln Ser Ile Ile
690 695 700
Ala Tyr Thr Met Ser Leu Gly Ala Glu Asn Ser Val Ala Tyr Ser Asn
705 710 715 720
Asn Ser Ile Ala Ile Pro Thr Asn Phe Thr Ile Ser Val Thr Thr Glu
725 730 735
Ile Leu Pro Val Ser Met Thr Lys Thr Ser Val Asp Cys Thr Met Tyr
740 745 750
Ile Cys Gly Asp Ser Thr Glu Cys Ser Asn Leu Leu Leu Gln Tyr Gly
755 760 765
Ser Phe Cys Thr Gln Leu Asn Arg Ala Leu Thr Gly Ile Ala Val Glu
770 775 780
Gln Asp Lys Asn Thr Gln Glu Val Phe Ala Gln Val Lys Gln Ile Tyr
785 790 795 800
Lys Thr Pro Pro Ile Lys Asp Phe Gly Gly Phe Asn Phe Ser Gln Ile
805 810 815
Leu Pro Asp Pro Ser Lys Pro Ser Lys Arg Ser Phe Ile Glu Asp Leu
820 825 830
Leu Phe Asn Lys Val Thr Leu Ala Asp Ala Gly Phe Ile Lys Gln Tyr
835 840 845
Gly Asp Cys Leu Gly Asp Ile Ala Ala Arg Asp Leu Ile Cys Ala Gln
850 855 860
Lys Phe Asn Gly Leu Thr Val Leu Pro Pro Leu Leu Thr Asp Glu Met
865 870 875 880
Ile Ala Gln Tyr Thr Ser Ala Leu Leu Ala Gly Thr Ile Thr Ser Gly
885 890 895
Trp Thr Phe Gly Ala Gly Ala Ala Leu Gln Ile Pro Phe Ala Met Gln
900 905 910
Met Ala Tyr Arg Phe Asn Gly Ile Gly Val Thr Gln Asn Val Leu Tyr
915 920 925
Glu Asn Gln Lys Leu Ile Ala Asn Gln Phe Asn Ser Ala Ile Gly Lys
930 935 940
Ile Gln Asp Ser Leu Ser Ser Thr Ala Ser Ala Leu Gly Lys Leu Gln
945 950 955 960
Asp Val Val Asn Gln Asn Ala Gln Ala Leu Asn Thr Leu Val Lys Gln
965 970 975
Leu Ser Ser Asn Phe Gly Ala Ile Ser Ser Val Leu Asn Asp Ile Leu
980 985 990
Ser Arg Leu Asp Lys Val Glu Ala Glu Val Gln Ile Asp Arg Leu Ile
995 1000 1005
Thr Gly Arg Leu Gln Ser Leu Gln Thr Tyr Val Thr Gln Gln Leu
1010 1015 1020
Ile Arg Ala Ala Glu Ile Arg Ala Ser Ala Asn Leu Ala Ala Thr
1025 1030 1035
Lys Met Ser Glu Cys Val Leu Gly Gln Ser Lys Arg Val Asp Phe
1040 1045 1050
Cys Gly Lys Gly Tyr His Leu Met Ser Phe Pro Gln Ser Ala Pro
1055 1060 1065
His Gly Val Val Phe Leu His Val Thr Tyr Val Pro Ala Glu Lys
1070 1075 1080
Asn Phe Thr Thr Ala Pro Ala Ile Cys His Asp Gly Lys Ala His
1085 1090 1095
Phe Pro Arg Glu Gly Val Phe Val Ser Asn Gly Thr His Trp Phe
1100 1105 1110
Val Thr Gln Arg Asn Phe Tyr Glu Pro Gln Ile Ile Thr Thr Asp
1115 1120 1125
Asn Thr Phe Val Ser Gly Asn Cys Asp Val Val Ile Gly Ile Val
1130 1135 1140
Asn Asn Thr Val Tyr Asp Pro Leu Gln Pro Glu Leu Asp Ser Phe
1145 1150 1155
Lys Glu Glu Leu Asp Lys Tyr Phe Lys Asn His Thr Ser Pro Asp
1160 1165 1170
Val Asp Leu Gly Asp Ile Ser Gly Ile Asn Ala Ser Val Val Asn
1175 1180 1185
Ile Gln Lys Glu Ile Asp Arg Leu Asn Glu Val Ala Lys Asn Leu
1190 1195 1200
Asn Glu Ser Leu Ile Asp Leu Gln Glu Leu Gly Lys Tyr Glu Gln
1205 1210 1215
Tyr Ile Lys Trp Pro Tyr Ile Pro Glu Ala Pro Arg Asp Gly Gln
1220 1225 1230
Ala Tyr Val Arg Lys Asp Gly Glu Trp Val Leu Leu Ser Thr Phe
1235 1240 1245
Leu His His His His His His
1250 1255
<210> 23
<211> 1246
<212> PRT
<213> Artificial Sequence
<220>
<223> 重组Spike蛋白
<400> 23
Met Arg Ala Trp Ile Phe Phe Leu Leu Cys Leu Ala Gly Arg Ala Leu
1 5 10 15
Ala Val Asn Leu Thr Thr Arg Thr Gln Leu Pro Pro Ala Tyr Thr Asn
20 25 30
Ser Phe Thr Arg Gly Val Tyr Tyr Pro Asp Lys Val Phe Arg Ser Ser
35 40 45
Val Leu His Ser Thr Gln Asp Leu Phe Leu Pro Phe Phe Ser Asn Val
50 55 60
Thr Trp Phe His Ala Ile His Val Ser Gly Thr Asn Gly Thr Lys Arg
65 70 75 80
Phe Asp Asn Pro Val Leu Pro Phe Asn Asp Gly Val Tyr Phe Ala Ser
85 90 95
Thr Glu Lys Ser Asn Ile Ile Arg Gly Trp Ile Phe Gly Thr Thr Leu
100 105 110
Asp Ser Lys Thr Gln Ser Leu Leu Ile Val Asn Asn Ala Thr Asn Val
115 120 125
Val Ile Lys Val Cys Glu Phe Gln Phe Cys Asn Asp Pro Phe Leu Gly
130 135 140
Val Tyr Tyr His Lys Asn Asn Lys Ser Trp Met Glu Ser Glu Phe Arg
145 150 155 160
Val Tyr Ser Ser Ala Asn Asn Cys Thr Phe Glu Tyr Val Ser Gln Pro
165 170 175
Phe Leu Met Asp Leu Glu Gly Lys Gln Gly Asn Phe Lys Asn Leu Arg
180 185 190
Glu Phe Val Phe Lys Asn Ile Asp Gly Tyr Phe Lys Ile Tyr Ser Lys
195 200 205
His Thr Pro Ile Asn Leu Val Arg Asp Leu Pro Gln Gly Phe Ser Ala
210 215 220
Leu Glu Pro Leu Val Asp Leu Pro Ile Gly Ile Asn Ile Thr Arg Phe
225 230 235 240
Gln Thr Leu Leu Ala Leu His Arg Ser Tyr Leu Thr Pro Gly Asp Ser
245 250 255
Ser Ser Gly Trp Thr Ala Gly Ala Ala Ala Tyr Tyr Val Gly Tyr Leu
260 265 270
Gln Pro Arg Thr Phe Leu Leu Lys Tyr Asn Glu Asn Gly Thr Ile Thr
275 280 285
Asp Ala Val Asp Cys Ala Leu Asp Pro Leu Ser Glu Thr Lys Cys Thr
290 295 300
Leu Lys Ser Phe Thr Val Glu Lys Gly Ile Tyr Gln Thr Ser Asn Phe
305 310 315 320
Arg Val Gln Pro Thr Glu Ser Ile Val Arg Phe Pro Asn Ile Thr Asn
325 330 335
Leu Cys Pro Phe Gly Glu Val Phe Asn Ala Thr Arg Phe Ala Ser Val
340 345 350
Tyr Ala Trp Asn Arg Lys Arg Ile Ser Asn Cys Val Ala Asp Tyr Ser
355 360 365
Val Leu Tyr Asn Ser Ala Ser Phe Ser Thr Phe Lys Cys Tyr Gly Val
370 375 380
Ser Pro Thr Lys Leu Asn Asp Leu Cys Phe Thr Asn Val Tyr Ala Asp
385 390 395 400
Ser Phe Val Ile Arg Gly Asp Glu Val Arg Gln Ile Ala Pro Gly Gln
405 410 415
Thr Gly Lys Ile Ala Asp Tyr Asn Tyr Lys Leu Pro Asp Asp Phe Thr
420 425 430
Gly Cys Val Ile Ala Trp Asn Ser Asn Asn Leu Asp Ser Lys Val Gly
435 440 445
Gly Asn Tyr Asn Tyr Leu Tyr Arg Leu Phe Arg Lys Ser Asn Leu Lys
450 455 460
Pro Phe Glu Arg Asp Ile Ser Thr Glu Ile Tyr Gln Ala Gly Ser Thr
465 470 475 480
Pro Cys Asn Gly Val Glu Gly Phe Asn Cys Tyr Phe Pro Leu Gln Ser
485 490 495
Tyr Gly Phe Gln Pro Thr Asn Gly Val Gly Tyr Gln Pro Tyr Arg Val
500 505 510
Val Val Leu Ser Phe Glu Leu Leu His Ala Pro Ala Thr Val Cys Gly
515 520 525
Pro Lys Lys Ser Thr Asn Leu Val Lys Asn Lys Cys Val Asn Phe Asn
530 535 540
Phe Asn Gly Leu Thr Gly Thr Gly Val Leu Thr Glu Ser Asn Lys Lys
545 550 555 560
Phe Leu Pro Phe Gln Gln Phe Gly Arg Asp Ile Ala Asp Thr Thr Asp
565 570 575
Ala Val Arg Asp Pro Gln Thr Leu Glu Ile Leu Asp Ile Thr Pro Cys
580 585 590
Ser Phe Gly Gly Val Ser Val Ile Thr Pro Gly Thr Asn Thr Ser Asn
595 600 605
Gln Val Ala Val Leu Tyr Gln Asp Val Asn Cys Thr Glu Val Pro Val
610 615 620
Ala Ile His Ala Asp Gln Leu Thr Pro Thr Trp Arg Val Tyr Ser Thr
625 630 635 640
Gly Ser Asn Val Phe Gln Thr Arg Ala Gly Cys Leu Ile Gly Ala Glu
645 650 655
His Val Asn Asn Ser Tyr Glu Cys Asp Ile Pro Ile Gly Ala Gly Ile
660 665 670
Cys Ala Ser Tyr Gln Thr Gln Thr Asn Ser Pro Arg Ala Ala Ala Ser
675 680 685
Val Ala Ser Gln Ser Ile Ile Ala Tyr Thr Met Ser Leu Gly Ala Glu
690 695 700
Asn Ser Val Ala Tyr Ser Asn Asn Ser Ile Ala Ile Pro Thr Asn Phe
705 710 715 720
Thr Ile Ser Val Thr Thr Glu Ile Leu Pro Val Ser Met Thr Lys Thr
725 730 735
Ser Val Asp Cys Thr Met Tyr Ile Cys Gly Asp Ser Thr Glu Cys Ser
740 745 750
Asn Leu Leu Leu Gln Tyr Gly Ser Phe Cys Thr Gln Leu Asn Arg Ala
755 760 765
Leu Thr Gly Ile Ala Val Glu Gln Asp Lys Asn Thr Gln Glu Val Phe
770 775 780
Ala Gln Val Lys Gln Ile Tyr Lys Thr Pro Pro Ile Lys Asp Phe Gly
785 790 795 800
Gly Phe Asn Phe Ser Gln Ile Leu Pro Asp Pro Ser Lys Pro Ser Lys
805 810 815
Arg Ser Phe Ile Glu Asp Leu Leu Phe Asn Lys Val Thr Leu Ala Asp
820 825 830
Ala Gly Phe Ile Lys Gln Tyr Gly Asp Cys Leu Gly Asp Ile Ala Ala
835 840 845
Arg Asp Leu Ile Cys Ala Gln Lys Phe Asn Gly Leu Thr Val Leu Pro
850 855 860
Pro Leu Leu Thr Asp Glu Met Ile Ala Gln Tyr Thr Ser Ala Leu Leu
865 870 875 880
Ala Gly Thr Ile Thr Ser Gly Trp Thr Phe Gly Ala Gly Ala Ala Leu
885 890 895
Gln Ile Pro Phe Ala Met Gln Met Ala Tyr Arg Phe Asn Gly Ile Gly
900 905 910
Val Thr Gln Asn Val Leu Tyr Glu Asn Gln Lys Leu Ile Ala Asn Gln
915 920 925
Phe Asn Ser Ala Ile Gly Lys Ile Gln Asp Ser Leu Ser Ser Thr Ala
930 935 940
Ser Ala Leu Gly Lys Leu Gln Asp Val Val Asn Gln Asn Ala Gln Ala
945 950 955 960
Leu Asn Thr Leu Val Lys Gln Leu Ser Ser Asn Phe Gly Ala Ile Ser
965 970 975
Ser Val Leu Asn Asp Ile Leu Ser Arg Leu Asp Lys Val Glu Ala Glu
980 985 990
Val Gln Ile Asp Arg Leu Ile Thr Gly Arg Leu Gln Ser Leu Gln Thr
995 1000 1005
Tyr Val Thr Gln Gln Leu Ile Arg Ala Ala Glu Ile Arg Ala Ser
1010 1015 1020
Ala Asn Leu Ala Ala Thr Lys Met Ser Glu Cys Val Leu Gly Gln
1025 1030 1035
Ser Lys Arg Val Asp Phe Cys Gly Lys Gly Tyr His Leu Met Ser
1040 1045 1050
Phe Pro Gln Ser Ala Pro His Gly Val Val Phe Leu His Val Thr
1055 1060 1065
Tyr Val Pro Ala Glu Lys Asn Phe Thr Thr Ala Pro Ala Ile Cys
1070 1075 1080
His Asp Gly Lys Ala His Phe Pro Arg Glu Gly Val Phe Val Ser
1085 1090 1095
Asn Gly Thr His Trp Phe Val Thr Gln Arg Asn Phe Tyr Glu Pro
1100 1105 1110
Gln Ile Ile Thr Thr Asp Asn Thr Phe Val Ser Gly Asn Cys Asp
1115 1120 1125
Val Val Ile Gly Ile Val Asn Asn Thr Val Tyr Asp Pro Leu Gln
1130 1135 1140
Pro Glu Leu Asp Ser Phe Lys Glu Glu Leu Asp Lys Tyr Phe Lys
1145 1150 1155
Asn His Thr Ser Pro Asp Val Asp Leu Gly Asp Ile Ser Gly Ile
1160 1165 1170
Asn Ala Ser Val Val Asn Ile Gln Lys Glu Ile Asp Arg Leu Asn
1175 1180 1185
Glu Val Ala Lys Asn Leu Asn Glu Ser Leu Ile Asp Leu Gln Glu
1190 1195 1200
Leu Gly Lys Tyr Glu Gln Tyr Ile Lys Trp Pro Tyr Ile Pro Glu
1205 1210 1215
Ala Pro Arg Asp Gly Gln Ala Tyr Val Arg Lys Asp Gly Glu Trp
1220 1225 1230
Val Leu Leu Ser Thr Phe Leu His His His His His His
1235 1240 1245
<210> 24
<211> 3732
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 24
atgttcgtct tccttgtgtt gcttccgctc gtgtcatctc aatgtgtgaa tctgaccact 60
cggacgcagc ttcctccagc ttataccaat agcttcacca ggggggttta ttaccctgat 120
aaagttttca ggagctcagt gctgcattct acgcaggatc tgttcctccc tttcttcagt 180
aacgtgacgt ggtttcacgc tattcatgtc tctgggacca atgggacaaa gcggttcgac 240
aatccagtgc tcccatttaa tgatggcgtg tactttgcga gtactgaaaa gtctaatatt 300
atacggggat ggattttcgg gacaactttg gatagtaaga cccagagcct gcttattgtt 360
aacaatgcca caaatgttgt catcaaggtg tgtgagttcc agttctgcaa tgaccctttc 420
cttggtgtgt actatcacaa aaataacaag agctggatgg agtcagagtt tagagtgtac 480
tcatccgcaa ataactgcac atttgagtac gtgagccagc ctttcctgat ggatctggag 540
gggaagcagg ggaattttaa gaatctgaga gagttcgttt tcaaaaatat cgacggctac 600
ttcaagattt attctaagca cactcctatt aatcttgtgc gcgatttgcc ccaaggcttc 660
tctgcacttg agcccctcgt cgatctccct atcggcatta atattacgag attccagacg 720
ctcctggcgc tccaccggtc atatcttacg cctggggatt cttctagcgg ttggactgct 780
ggtgctgctg cttactacgt gggctacctc cagcctcgca ccttcttgct caagtacaat 840
gaaaacggga ctataacaga cgccgtggat tgcgcacttg accccctgag cgagacaaag 900
tgtaccctta aaagttttac ggtggagaag ggtatctatc agacatctaa ttttcgggtt 960
cagccaaccg agtctattgt gcgctttcca aatattacca atctgtgccc tttcggggag 1020
gtttttaacg ctacccggtt tgcatctgtt tacgcttgga acagaaaacg gattagcaat 1080
tgtgtggctg attattccgt cctgtacaac tctgcttcat tctccacctt taagtgctac 1140
ggggtcagcc caactaaact gaacgatttg tgttttacta acgtttacgc cgattccttc 1200
gtgatccggg gggatgaagt gcggcagata gctccaggcc aaaccggaaa gatagcagac 1260
tacaattata agctccctga cgattttact gggtgcgtaa tagcctggaa tagcaataac 1320
ctcgacagta aggtgggagg caactacaac tacctctata ggctctttag gaaaagcaac 1380
ttgaagcctt ttgagagaga tattagtacc gaaatctatc aagcagggtc tactccatgt 1440
aatggagttg aaggctttaa ttgctacttt ccgctgcagt cctatggttt tcagcctacg 1500
aacggagttg gttatcagcc ctacagggtt gtggtactgt ctttcgaact gcttcacgcc 1560
cctgcgaccg tatgtggacc aaaaaaatcc accaacttgg tgaaaaataa gtgcgttaat 1620
ttcaatttta atgggctcac aggaacagga gttctcactg agtctaataa gaaattcctg 1680
ccttttcaac agttcgggag ggatatcgct gacacgacag atgcggttcg ggaccctcag 1740
acattggaaa tactggacat tacgccctgc agtttcgggg gagtatccgt cataactccc 1800
gggacaaaca ccagtaatca ggtcgctgtt ctgtatcaag acgtaaattg caccgaagtg 1860
cccgtagcaa tccacgctga tcagcttacc cccacttgga gggtgtattc taccggaagt 1920
aatgtcttcc aaacgagagc gggctgcctg attggtgctg aacatgttaa caacagctat 1980
gaatgcgaca tcccaatagg cgccggaatt tgcgcgtcct accagacgca aactaattcc 2040
ccacgggctg ccgctagcgt agcttcacag agcattatag catatacaat gagcctgggc 2100
gctgagaaca gtgtggccta tagcaataac agtattgcta tccctactaa tttcacaatc 2160
agtgtgacga cagaaatact gccagtgagc atgaccaaaa ccagtgtcga ttgcaccatg 2220
tatatctgcg gcgactccac agaatgttcc aatctgctgt tgcagtacgg tagcttctgt 2280
acacagctca atcgggcact gacaggcatt gctgttgagc aagataaaaa cacccaggaa 2340
gtgttcgccc aggttaaaca gatctacaag acccccccta taaaagactt cgggggcttc 2400
aatttttctc agatcttgcc cgacccatca aagccgtcaa agcgctcttt catcgaagat 2460
cttttgttta ataaagtgac tctggccgac gctgggttca ttaagcagta tggggactgt 2520
ctcggagaca ttgctgcccg ggacttgatc tgcgcacaaa agtttaatgg actcacagtt 2580
ctgccaccac tgcttactga cgaaatgatc gcccaataca cgagtgctct gctcgcgggc 2640
acgattacca gtggctggac atttggagca ggcgccgctc ttcagatccc cttcgcaatg 2700
cagatggcat accggtttaa tggcattgga gtcacccaaa atgtcctcta tgagaaccaa 2760
aagctgatcg ctaaccagtt caactctgca attggaaaaa tccaggattc tctctcctca 2820
accgctagcg ccctgggaaa gcttcaggac gtggtgaacc agaatgcaca ggcccttaat 2880
accctggtga agcaactgtc tagcaacttt ggagcgatca gctcagtgct caatgatatt 2940
ctgagccggc tcgacaaggt ggaggcagaa gttcaaatag acagactgat aaccggtcgg 3000
ctgcagagcc tgcagacata cgtcacgcaa cagctcatcc gcgccgcaga aattcgagca 3060
agcgctaatc tcgcggcaac aaagatgagt gagtgcgtgc tcggacagag caaaagagtc 3120
gacttttgtg gcaagggata ccacctgatg agctttccac agtctgcccc acacggcgtg 3180
gtgttcttgc acgtcaccta cgtgccagct gagaaaaatt ttactaccgc tcccgctatt 3240
tgccacgatg gaaaagctca ctttccgcgg gagggagtct tcgtgagcaa cggtactcat 3300
tggttcgtca cacagcggaa cttctacgag ccacaaataa taactacaga taatactttc 3360
gtgagtggca actgtgacgt tgtgattgga atcgtaaata acacagtcta tgaccccctt 3420
caaccagagc tggatagttt caaggaggaa cttgataagt attttaaaaa ccatacctca 3480
cctgatgtcg atcttgggga tatcagcggc atcaacgcct ccgtggtcaa cattcaaaag 3540
gagatcgatc gactgaacga ggtcgccaag aacttgaacg agtccctgat cgacctgcaa 3600
gaactcggca agtatgaaca gtacattaag tggccctata tcccagaggc tccacgagat 3660
ggacaggcgt atgtccggaa ggacggtgag tgggtgcttc tgtccacttt tctccaccat 3720
caccaccacc at 3732
<210> 25
<211> 3747
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 25
atgtacagga tgcagctgct gtcttgcatt gccttgtcac tcgccctggt gacgaactcc 60
gtgaacctga ccacaaggac tcagctgcca cctgcctata ccaacagttt caccagaggc 120
gtttactacc cagataaagt gttcagaagt tccgttctgc atagtacaca ggacttgttc 180
ctgccattct tctccaacgt gacctggttt cacgcaatac acgtgagcgg tactaatggc 240
accaagcggt tcgataatcc agtgcttcca tttaatgacg gggtttactt tgctagcacc 300
gaaaaatcta atatcattag gggctggata tttggcacca cgcttgactc aaaaacacag 360
agcctgctca ttgtgaacaa cgccactaac gttgtgatca aggtgtgcga gtttcagttc 420
tgtaatgatc cttttttggg cgtttactat cataaaaaca ataagtcttg gatggaatcc 480
gaatttcggg tctactcaag cgctaataac tgtacgttcg aatatgtgtc acaacctttt 540
cttatggatt tggaagggaa acagggaaac tttaagaacc ttcgggaatt tgtcttcaag 600
aacattgacg gatatttcaa gatctactcc aagcacacgc caatcaactt ggttagggac 660
ctgccccaag ggttttctgc gttggaacca ttggtggacc ttccaatagg gatcaacatc 720
actaggttcc agactttgct ggccctccac cgctcctatt tgaccccggg tgactcatca 780
tcaggatgga cagccggcgc tgctgcgtac tatgtggggt acctccagcc ccgcactttc 840
ctcctgaagt ataacgagaa cggtacgatc accgacgctg tggactgcgc actggaccct 900
ctcagtgaga ctaagtgcac acttaaaagc ttcaccgtag aaaagggcat ctatcagaca 960
agcaatttcc gcgtgcagcc gacggaatct attgttcggt tcccgaacat cacgaacctg 1020
tgccccttcg gagaagtgtt taacgcgacc agatttgcaa gcgtctatgc gtggaacaga 1080
aagcggatta gcaactgcgt cgctgactac tctgttttgt ataattcagc tagcttctca 1140
acattcaagt gttacggggt gagccccact aagttgaacg acttgtgttt tacgaacgtg 1200
tatgccgata gcttcgtgat ccggggggat gaggtgaggc aaattgctcc cgggcagacc 1260
ggaaaaatcg cggactacaa ttacaaactg ccggacgact ttaccggttg tgtgatagcc 1320
tggaacagca acaaccttga tagcaaagtc ggaggcaact acaattacct gtaccggctt 1380
tttcgcaaaa gcaacctgaa acccttcgag agggatatct ccacagaaat ttaccaggct 1440
ggttcaacac cctgtaatgg cgtggagggc ttcaactgct attttcctct ccagtcctac 1500
ggcttccaac ccacaaacgg cgtgggatat caaccctatc gagtggtggt gcttagcttt 1560
gaactcctgc acgcccccgc tactgtttgt gggccaaaga agtcaaccaa tctcgtcaaa 1620
aataagtgcg tcaacttcaa ttttaatggc ctgaccggta caggagtgct gactgagagc 1680
aacaaaaaat tcctcccttt ccagcaattt ggcagggata tagcagatac aacagacgct 1740
gtgcgagatc cacagacact cgaaatcctg gacatcaccc cctgctcttt cggaggcgtc 1800
tcagttatta ccccaggaac taataccagt aatcaggtgg ctgttctgta tcaagacgtt 1860
aactgtacag aagtcccggt ggcgatccat gccgatcagc tgacacccac ctggcgcgta 1920
tatagtaccg gcagtaatgt gttccagaca cgggctgggt gtctgatcgg agccgagcat 1980
gtgaacaatt cttacgaatg cgatatcccc attggagcag gcatctgcgc ttcttatcag 2040
acccagacca attctcccag agccgctgct tctgtggcct ctcagtccat tatcgcctat 2100
acaatgtctc tgggagctga gaatagtgtt gcttatagca ataacagcat tgctatccct 2160
acgaatttta caattagcgt tactaccgaa attcttcccg tctccatgac aaagacctct 2220
gtggattgta ctatgtatat ctgcggcgat agtactgagt gttctaacct gctgttgcag 2280
tatggctctt tttgtaccca gctgaataga gcattgactg ggattgcagt ggagcaggac 2340
aaaaacactc aggaagtatt cgctcaagtg aaacaaatct acaagacacc tcctattaaa 2400
gacttcgggg gttttaactt tagccaaatt ctccctgacc ccagtaaacc ttccaaacgc 2460
tcatttatag aagatcttct ttttaacaaa gtgaccctgg ctgacgctgg attcataaaa 2520
cagtacggag actgcctggg tgatatcgcc gctcgcgatc tcatttgtgc acagaagttt 2580
aatggcctga ctgtgctccc accactcctg acggatgaga tgatcgctca atacacctcc 2640
gcactgctgg caggaaccat taccagcggc tggacatttg gagccggagc ggcattgcaa 2700
attccattcg ctatgcagat ggcctaccga ttcaacggta tcggtgtaac acaaaacgtg 2760
ctctacgaga atcagaaatt gatagcaaac caatttaatt ccgccattgg caagattcaa 2820
gattccctgt cctcaacggc tagcgccctt gggaagctgc aggatgtggt caaccagaac 2880
gcgcaggctt tgaatacgct ggtaaagcaa ctgtcaagta acttcggcgc tatctctagt 2940
gtgctgaacg atattttgtc caggctggac aaagtggaag ctgaggtaca gatcgatcga 3000
ttgattaccg gacgcctgca gtccctgcag acttacgtca cccagcagtt gataagagct 3060
gccgagatta gagcttccgc caacctggcc gccactaaga tgtcagagtg cgtgctcgga 3120
cagagtaagc gcgtggattt ttgcgggaaa ggctaccacc tgatgagctt ccctcagtca 3180
gccccacatg gagtggtgtt cctgcacgtg acttatgtgc ccgcagagaa gaactttact 3240
actgctcctg caatctgtca cgatggaaag gcgcactttc cgagagaagg cgtgttcgtc 3300
tcaaacggga cccactggtt cgtcacgcag cgcaatttct atgagcccca gatcattacc 3360
actgacaaca catttgtgag cggaaattgc gacgtggtga tcgggatcgt taataataca 3420
gtgtatgatc ccttgcagcc agaactggac agttttaaag aggaactgga taaatacttc 3480
aagaatcaca ccagcccaga tgtcgatctg ggggacatct ccggaattaa cgccagcgtg 3540
gttaacattc aaaaagagat tgatagactg aatgaagtgg ccaaaaatct caatgaaagc 3600
ctgatcgacc tgcaggaact ggggaaatat gagcagtaca tcaaatggcc atatattcca 3660
gaagcaccga gagacggcca ggcttacgtg agaaaggacg gagagtgggt cctcctgagt 3720
acatttcttc atcatcacca tcatcat 3747
<210> 26
<211> 3747
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 26
atggagactg atactttgct gctgtgggtg ctgttgctct gggtgccagg cagcaccggt 60
gttaacctta ctacccgcac tcagctgccc ccagcctaca ctaactcttt cacccgagga 120
gtgtattacc ccgacaaagt ctttcggagt tctgtcctgc actctaccca ggatttgttc 180
ctccccttct ttagcaatgt cacctggttt catgccatcc acgtgtccgg gactaacgga 240
accaagaggt tcgataaccc tgtccttcct ttcaacgatg gtgtgtattt tgcctccacg 300
gagaagtcta acatcatcag aggctggatc ttcggaacaa cccttgattc caaaacccaa 360
tcactgctca ttgtgaataa cgctactaac gttgtgatta aagtatgtga gttccaattc 420
tgcaacgacc cctttctcgg cgtgtattac cacaagaaca ataaatcatg gatggaaagt 480
gaattccgcg tctactcttc tgcaaacaat tgcaccttcg aatacgtcag ccagccattt 540
ttgatggacc tggaggggaa gcagggaaat tttaagaatc tgagagagtt tgtgttcaaa 600
aatattgacg ggtacttcaa gatctattct aagcataccc caataaatct ggtccgcgat 660
ctcccacagg gtttttctgc actcgaacca ctcgttgatc tgcctatcgg tatcaacatc 720
acaaggttcc agaccctgct tgctctgcac aggtcctatt tgactccggg agatagttct 780
tctggctgga cagccggagc tgctgcatac tacgtcgggt atttgcagcc ccgcactttt 840
ctcttgaaat acaacgagaa tggcaccatc acagacgcag tagactgcgc actcgaccca 900
ctcagcgaaa caaagtgtac tctgaagtct tttacagtgg aaaaaggaat ttatcagacg 960
agcaatttca gagtccagcc aacagagtct atcgtaagat tccctaatat cacaaatttg 1020
tgtcccttcg gagaggtctt caacgcaacc cgatttgcca gcgtgtatgc ctggaacaga 1080
aagaggatct ctaattgcgt agcggactac tctgtcctct acaatagcgc gagcttttca 1140
acatttaaat gttatggagt cagccctact aagctcaatg atctctgctt tacaaatgtt 1200
tacgctgact cctttgtaat aagaggagac gaggtgaggc agatagcccc tgggcaaact 1260
ggaaaaatcg ctgactacaa ctacaaactg cctgacgatt ttactggttg cgtgatagcc 1320
tggaattcca ataacctgga ctctaaagtt ggtggaaact ataattacct gtacaggctg 1380
tttcgcaaat caaatctgaa gcccttcgaa agggacatat ccaccgaaat ctaccaggct 1440
ggatcaacac cgtgcaacgg agtggagggc tttaattgtt attttcctct gcagtcttat 1500
gggttccaac ccactaacgg tgtgggatat cagccctata gagtcgtggt cctgagcttt 1560
gaactgttgc acgcacctgc aactgtgtgt ggacctaaga aaagtaccaa cttggtgaaa 1620
aataaatgtg tgaacttcaa ctttaacgga ctgaccggga ctggcgtgct gacggaaagc 1680
aacaaaaaat ttctgccatt tcaacagttc ggccgggata ttgcggatac aactgatgca 1740
gttagggacc cccagacgct cgaaatactc gacattaccc cctgcagctt tggaggagtg 1800
tcagtcatca cccctggtac taacacttct aaccaggtcg ctgtgttgta tcaggatgtt 1860
aactgcaccg aggttcccgt ggcgatccac gctgatcagc tgacgccgac ttggcgcgtg 1920
tatagtactg gctcaaacgt ctttcagact cgggcagggt gtcttattgg tgccgaacat 1980
gtgaacaatt cttatgaatg cgacattcca ataggggctg gcatttgcgc atcctaccaa 2040
acacaaacga actcacctag agctgccgct tctgtggcca gtcagtctat aatcgcttac 2100
acaatgtccc tcggcgccga gaatagcgtc gcctacagca acaattccat cgcaattccc 2160
accaacttta caatcagcgt tacaactgag attctgccag tttcaatgac caagaccagc 2220
gttgactgca ccatgtatat ctgtggcgat agcaccgaat gcagcaacct tctgctccag 2280
tatgggtctt tctgcacaca gctgaacagg gcacttacag gcatcgcagt cgagcaggat 2340
aagaacacac aggaagtgtt tgcccaagtc aagcagatct acaagacccc tcctattaag 2400
gactttgggg gctttaactt ttcccagatt ttgccggacc caagtaagcc atctaaacgc 2460
agtttcatcg aggacttgct gtttaataaa gtaactctgg cagacgccgg atttatcaag 2520
cagtacggcg actgtctggg tgatatagct gcccgcgacc ttatttgcgc acagaagttc 2580
aacggattga ccgtgcttcc tccattgttg actgatgaga tgatcgccca gtatacctcc 2640
gcccttctgg ctggcaccat tacctctggt tggacatttg gagctggggc ggccctccaa 2700
ataccctttg caatgcagat ggcttacaga tttaacggta tcggggttac tcaaaatgta 2760
ctttacgaga accagaaact cattgccaac cagtttaata gcgcgattgg gaagatacag 2820
gattccctgt cctctacagc ctccgctttg ggtaagttgc aggatgtggt caaccagaac 2880
gcgcaagccc tcaacacgtt ggtaaaacag ctgagtagta attttggcgc tatctccagc 2940
gttcttaatg acatcctgtc ccggcttgat aaggtggagg ccgaagtcca gatcgaccgc 3000
ctgatcacag gaagattgca atcacttcaa acatatgtga cacagcagct gatccgagcg 3060
gctgagataa gggcctcagc caatctcgcc gcaactaaga tgtccgagtg cgtactggga 3120
cagagcaaaa gagtcgactt ttgtggaaag gggtaccatt tgatgtcttt tcctcagagc 3180
gctccccacg gcgtcgtgtt tttgcatgtc acatacgtcc ccgctgagaa gaattttact 3240
acagccccag ctatttgcca cgatggaaag gcacatttcc ctagggaggg tgtattcgtc 3300
agtaatggaa cccactggtt tgtcacccag agaaacttct acgagccaca gataatcacc 3360
actgacaata catttgtctc tggtaattgt gatgttgtga taggaatcgt aaataatact 3420
gtgtacgacc ctctccagcc tgagctggac agttttaaag aagagctgga taaatacttc 3480
aaaaaccata ccagccctga cgtagacctg ggggatatca gtggcattaa cgcctcagtc 3540
gtgaacatcc agaaggagat tgaccgcctc aacgaggtcg ctaagaacct gaatgaatcc 3600
ctgatcgacc tccaggaact gggcaaatat gagcaataca tcaagtggcc ttatattccc 3660
gaggctccta gggatggtca agcatacgtc cgcaaagacg gcgagtgggt tttgctgagc 3720
acgttcctcc accaccatca tcatcac 3747
<210> 27
<211> 3756
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 27
atggatgcca tgaagagagg attgtgctgt gttctgcttc tttgtggcgc cgtgttcgtg 60
tcacctagcg ttaatcttac cacccgaacc cagttgcctc ctgcttatac aaactctttc 120
acccgggggg tctattaccc agataaggtg ttcaggtcca gcgtgctgca cagcacgcag 180
gatctttttc tgcccttttt cagtaatgtg acctggtttc acgctatcca cgttagcggg 240
accaacggaa ccaagcgctt tgacaaccct gtgctgcctt tcaatgatgg agtgtacttc 300
gcctccactg agaaatccaa tataatccgc gggtggattt tcggcacgac actcgactcc 360
aagacgcaat ctcttctgat cgtgaataac gcaaccaatg ttgtcatcaa ggtatgtgag 420
tttcagtttt gtaacgatcc tttcctgggc gtgtactatc acaaaaacaa taagtcttgg 480
atggagagcg agttccgagt atattcatct gccaataatt gtaccttcga gtatgtctct 540
caaccttttc tgatggatct ggaggggaag cagggaaact tcaaaaactt gagggagttc 600
gtgttcaaaa atattgatgg ttactttaaa atctactcca agcacactcc aatcaatctg 660
gtgagagacc ttccacaggg tttctctgca ctggagccgt tggtggacct gccgattgga 720
ataaatatca cccggttcca aacgctgctg gcactgcatc ggtcctacct gacacctgga 780
gactctagct ccggctggac agcaggagct gctgcttatt atgtgggcta tctgcagcct 840
aggacattct tgctgaaata taacgaaaac ggaaccatca ctgatgcagt ggactgcgcg 900
ctcgaccctc tttccgagac taaatgcacc ttgaagtctt tcaccgttga gaagggcatc 960
taccagacta gtaacttccg ggtccagcca accgagagta tagtaagatt cccaaacatc 1020
accaatctct gtccatttgg ggaagtgttt aacgcaactc gatttgcttc cgtgtacgcc 1080
tggaatcgaa agcggatctc taattgtgtt gcagattata gcgtgttgta caattccgca 1140
agtttttcta ctttcaaatg ctatggtgta tctcccacaa agttgaatga tctctgcttc 1200
actaacgtgt acgcagattc attcgttata agaggagacg aagtgaggca gatagctccc 1260
ggccagacag ggaaaattgc tgactacaat tacaagcttc ccgatgattt cactgggtgc 1320
gtcatagcct ggaactcaaa taaccttgac agtaaagtag ggggaaatta caactatctc 1380
tatcggctct tccggaagtc taatcttaag cccttcgagc gagatatttc cacagaaatc 1440
taccaggctg gctctactcc ctgtaacgga gttgaagggt ttaattgtta cttccctttg 1500
caatcctatg gcttccagcc taccaacggg gtcgggtatc agccctatag agtggtcgtg 1560
ttgtcttttg aacttctcca cgcccctgct actgtgtgcg gtccaaagaa gagcactaat 1620
ctggtgaaaa acaagtgtgt gaacttcaac ttcaatggac tcacaggcac tggcgtactg 1680
accgaaagta ataagaagtt tctgcctttc caacaatttg gaagagacat tgccgatact 1740
actgacgcag tgcgagaccc gcagacgttg gaaatcttgg acatcacccc gtgctccttc 1800
gggggagtga gcgtaattac acctggaacg aacacttcca atcaggttgc agttctgtat 1860
caggacgtga attgtactga ggtgcccgtt gccatccacg cagaccagct tacaccaacc 1920
tggagggtct actctactgg gagtaacgtg tttcagacta gagcaggctg ccttatcgga 1980
gccgaacacg tgaataactc ttatgagtgt gacatcccca tcggagccgg catttgtgcc 2040
tcataccaga cccagactaa ttccccaaga gcggctgcat ctgtggcctc ccagagcata 2100
attgcgtaca ctatgtccct tggagctgaa aactccgttg catactccaa caactccatc 2160
gccatcccca ccaatttcac aatcagcgtg acgaccgaaa tcctgcctgt cagcatgact 2220
aagacctctg tggactgtac aatgtatatc tgcggagatt caaccgaatg ttctaacctc 2280
cttcttcagt atggttcctt ttgtacgcag ttgaatagag ccctcactgg tatagcagtc 2340
gaacaggata agaacaccca ggaggtgttt gctcaagtca aacagatcta caagacaccc 2400
cctatcaaag actttggggg cttcaatttc agccagattc tgccagatcc cagcaagcca 2460
tccaaacgga gtttcataga agacctcctg tttaacaagg tcacgctggc cgacgcggga 2520
tttatcaaac aatacggcga ttgcctcggt gatatcgcag ctcgcgacct catttgtgct 2580
cagaagttta atggcctgac tgtcttgcca cctctgctta cagacgagat gatagcccag 2640
tacacgtccg cgttgcttgc tggaaccatc acatctggtt ggacctttgg ggctggcgca 2700
gctctgcaaa ttcctttcgc catgcagatg gcatatcgct ttaatgggat cggcgtgacc 2760
caaaacgtgc tgtacgagaa tcagaaactc attgcaaacc aatttaactc tgcaattggt 2820
aagatccagg atagtctgag ctctaccgct tcagcactcg gcaagcttca ggacgtggtt 2880
aaccagaacg cccaggccct gaacactctg gttaagcagc ttagctccaa ttttggcgcc 2940
atatccagcg tgctgaacga tattctgagt agactggaca aggttgaggc cgaagtacaa 3000
atagatcgcc tgattactgg tcgcctgcag agcctgcaga catacgtcac tcagcagctg 3060
attcgcgctg cagagatacg cgccagtgcc aatcttgcag ccaccaaaat gagtgagtgc 3120
gtactcgggc agtccaaaag ggtggatttt tgtggcaaag gatatcattt gatgtccttc 3180
ccccaatcag ctccacacgg ggtggtgttc ctgcacgtta cctatgtacc ggctgaaaaa 3240
aatttcacta cggctcctgc aatctgccat gacggaaaag cacatttccc ccgggaggga 3300
gtgttcgttt ctaatgggac ccactggttt gtgacacagc gcaacttcta cgaaccacaa 3360
ataattacta ccgataacac ctttgtttca ggtaattgtg acgttgtcat tggaattgtc 3420
aacaacactg tgtatgaccc cctgcaacca gagctggact cctttaagga agaattggac 3480
aagtatttca aaaaccatac aagcccggac gttgatcttg gggatataag cggcataaat 3540
gcctctgtcg taaatatcca gaaagaaatc gataggttga atgaggttgc caaaaacctc 3600
aatgagagtt tgattgacct ccaagaactc ggcaagtacg agcagtacat aaagtggccg 3660
tatattcccg aggcgcctag agatggccag gcatatgtgc ggaaggacgg cgagtgggtt 3720
ttgttgagca cttttcttca ccaccaccac catcat 3756
<210> 28
<211> 3735
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 28
atgaaaacta tcattgccct tagctacatc ttttgtttgg tgctcggtgt gaacttgact 60
acacggacac agctgccccc tgcatatacc aactcattca caagaggcgt gtattacccc 120
gacaaggtct ttcgaagcag tgttcttcac tctacgcagg acctgtttct gcctttcttt 180
tccaacgtga cttggtttca cgcaatccac gtctctggaa ccaatggtac aaagcgcttc 240
gacaaccccg tgctgccctt caacgatggc gtttacttcg ccagcacgga gaaaagcaat 300
atcatacgag gatggatctt tggaactaca ctcgatagca agactcaatc tctgcttata 360
gtcaataacg ctaccaacgt cgtgatcaag gtgtgtgaat tccaattttg caatgatcct 420
tttctcggag tatactatca caaaaataac aaaagctgga tggagagcga gtttagggtg 480
tattcctcag ctaataattg cactttcgag tacgtgtctc agccatttct gatggacctg 540
gaaggtaaac agggtaactt taaaaacctg cgagaatttg tgtttaagaa tattgatgga 600
tacttcaaaa tttatagtaa acacactccc atcaatctcg tgagagacct tccacagggc 660
tttagcgccc tggagcctct ggtggatctg cctattggga ttaacatcac ccggtttcag 720
acccttctgg ctctgcatcg gagctacctg acccctggag actcctcatc cggctggaca 780
gctggcgccg ccgcgtacta cgtcggatat ctgcagccta gaaccttctt gttgaaatat 840
aacgagaatg ggacaattac cgacgccgtg gattgcgccc tcgatcccct ctctgaaacc 900
aagtgcaccc tcaaatcatt cactgtggag aagggcatat atcagacatc aaacttccgg 960
gtgcaaccga ccgagtccat agttcggttc cccaacatta ctaatctttg tcctttcgga 1020
gaagtgttca atgctacccg gtttgcatcc gtgtacgcat ggaacaggaa gcggatctca 1080
aactgcgtcg ccgactactc cgttctttac aattcagcaa gtttctctac attcaaatgt 1140
tacggcgtat cccctacaaa gctgaacgat ctgtgcttca ccaacgttta cgcggatagt 1200
ttcgtcatac ggggtgacga agtgcggcag attgcacctg gccaaactgg gaaaatagcc 1260
gactataatt acaaactgcc cgacgacttc actggatgtg tgatcgcttg gaattcaaac 1320
aacttggact ccaaagtagg ggggaactac aattacctct acaggctttt tcgaaagtct 1380
aatctcaagc cattcgagag agacattagc accgagattt atcaggccgg ctctacgcct 1440
tgcaatggcg tggagggatt taactgctat tttcctctcc agagctacgg cttccaacca 1500
accaatggag tcggatacca gccctacaga gtggtcgtgc tttcatttga acttcttcat 1560
gccccagcaa cagtgtgtgg tcctaaaaag tctaccaacc tggtgaagaa caaatgtgtg 1620
aatttcaact tcaacggtct caccggcacc ggggtgctga cagagagtaa taagaaattt 1680
ctcccatttc agcagttcgg ccgcgatatt gcggatacca cagatgctgt acgcgaccct 1740
cagacactgg agatacttga catcacccct tgcagctttg gaggcgtgtc agtcattaca 1800
cccggaacaa acaccagcaa tcaggtagca gtgctctatc aagacgtaaa ttgtacagaa 1860
gtgcctgtcg ccatccatgc tgatcagctg acccctacgt ggcgcgtgta ttccactggg 1920
agcaacgtgt ttcaaacacg ggcagggtgc ctgattgggg ccgagcacgt gaacaactct 1980
tacgaatgcg acatccctat cggagccgga atctgcgcct cttatcagac gcagactaac 2040
agccctaggg ccgcggcatc cgttgcgagc caatccatca tagcatatac aatgtcattg 2100
ggcgctgaga attctgttgc ttacagcaac aacagcatcg ccatcccaac gaatttcacg 2160
atcagtgtga ccaccgagat cctgcccgtt agcatgacca aaacctcagt cgattgtaca 2220
atgtacattt gtggggactc aacggaatgt agcaatctcc tgctccagta tggcagcttt 2280
tgtacacagt tgaatcgagc cctcaccggc atcgccgtcg aacaggataa gaacacgcag 2340
gaagtgttcg cccaggttaa acagatttac aaaacaccgc cgattaagga cttcggaggt 2400
ttcaatttta gccagatcct tcctgatccc agtaaaccct ctaagcggag ctttattgag 2460
gacctcctgt tcaataaggt gacactggct gacgccggat tcatcaagca atatggcgat 2520
tgcctcgggg acatagccgc cagagacctg atctgcgcgc agaaattcaa tggcctgacg 2580
gtcttgccac cactgctcac cgacgaaatg atcgcacaat atacctccgc ccttctggct 2640
ggaacaataa cttcagggtg gaccttcggg gcgggggccg ccttgcagat tccctttgcc 2700
atgcagatgg cgtatagatt taacggtatt ggggtcaccc agaatgtgct ttatgaaaat 2760
caaaagctca ttgcaaacca gtttaacagt gccatcggca aaatccaaga tagcctgagc 2820
agcacggcat cagcacttgg taagctgcag gacgtggtca accagaacgc ccaagcactg 2880
aatactctgg taaagcaact gagcagcaat tttggtgcca ttagcagtgt cctcaacgat 2940
atcttgagca ggctggacaa agtggaagcc gaagttcaaa ttgaccggct gatcactgga 3000
cggctccaga gtctgcagac ttacgtcacc cagcagctga tccgcgccgc cgagatcagg 3060
gcttcagcaa acctggctgc taccaagatg tccgagtgcg ttctgggcca gagtaaacgg 3120
gttgacttct gcggaaaggg ataccacctc atgagctttc ctcaaagcgc tccgcacggg 3180
gtggtcttcc ttcatgtaac ctacgtgcct gcagagaaaa attttactac cgctcccgca 3240
atctgccacg acggcaaagc ccatttccca agggaagggg tcttcgtgtc aaatgggaca 3300
cactggtttg taacacaacg aaacttctac gagcctcaaa tcatcaccac tgataataca 3360
tttgtatctg gaaactgcga tgtcgtgatc gggatcgtca ataacacggt ttacgaccct 3420
ctccagccgg aactggactc tttcaaggaa gagctggaca agtatttcaa aaatcacacg 3480
agccctgatg tggatctggg tgacatttca gggatcaacg cctcagtggt gaatattcaa 3540
aaagagattg acaggctcaa cgaagtggca aagaatctca acgagtcatt gatagacctt 3600
caagagctgg gaaagtatga gcagtacatt aaatggccct atataccaga agctcccaga 3660
gacgggcaag cgtatgttcg caaggacggg gagtgggtgc ttctgagcac ctttctgcac 3720
caccaccacc accat 3735
<210> 29
<211> 3747
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 29
atggagctgg gcctctcatg ggtgtttctc gttgcaatcc ttgagggagt tcagtgtgaa 60
gtgaacttga caacaaggac gcaactgccc cccgcatata caaactcctt cactcgcgga 120
gtctactatc ctgataaagt cttcaggagt agtgtgttgc atagcactca agacctcttc 180
ctgccctttt tctcaaacgt gacgtggttt catgcaatcc atgtgtctgg cactaacggc 240
acgaagcgct tcgataatcc tgtgctgccg ttcaatgacg gggtctattt tgcatcaact 300
gaaaaaagta atattatccg gggctggata tttggcacta ccctcgatag caagacccag 360
agcctcctga tcgtgaacaa cgccacaaac gtggtcatta aagtgtgtga gttccagttc 420
tgtaacgatc cctttttggg ggtgtattac cataagaaca ataagtcctg gatggagagc 480
gagtttcggg tttattctag cgccaataat tgcacctttg agtatgtctc ccaaccattc 540
ctgatggatc tggaagggaa acagggtaac ttcaagaatc tcagagagtt tgttttcaaa 600
aacatagatg ggtattttaa gatttattcc aagcatacgc cgattaacct cgttagagat 660
ctgcctcagg gcttttctgc cttggaaccg ctggtggatc tccctattgg tatcaatata 720
actaggttcc agactctgct ggctctgcat cggtcatacc tgacccctgg agatagttct 780
tccggatgga cagcaggagc cgccgcatac tacgtcggct acctgcagcc aagaacattc 840
ctcctgaagt acaacgagaa cgggaccatt acagatgctg ttgactgcgc cctcgatccc 900
ctgagtgaga ccaagtgcac tcttaagtcc ttcaccgtcg aaaagggaat ctatcagacc 960
agcaatttcc gggtgcaacc aactgagtcc atcgttcgct ttcccaatat cacaaacttg 1020
tgtccattcg gggaggtatt taatgcaaca cgctttgcct ctgtttacgc gtggaatcgc 1080
aaaaggatct caaattgcgt cgccgactac agcgtgctgt ataactcagc atcattttca 1140
accttcaagt gttacggtgt ctcccctaca aagcttaatg acctttgctt taccaatgtg 1200
tatgccgatt ccttcgtaat taggggcgat gaagttcggc agattgcacc tggacagacg 1260
ggtaaaattg cagactataa ctacaagctc cccgatgatt tcacaggatg cgtgatagca 1320
tggaactcaa acaacctgga cagcaaggta ggcggcaatt acaactattt gtacagactg 1380
ttcaggaagt ccaatctgaa gccatttgag cgagacattt ccaccgagat ctaccaggca 1440
ggctctaccc cttgtaacgg ggtcgagggg ttcaactgct acttcccact tcagtcctac 1500
ggctttcagc ccactaatgg cgttggctac cagccatacc gagtggtggt gctctccttt 1560
gaattgctcc acgcacccgc taccgtgtgt ggtccaaaaa aaagcacaaa cctcgtgaaa 1620
aacaaatgtg tgaatttcaa cttcaatggg ctcacaggaa ctggagtgct gacagaatct 1680
aacaagaagt tcttgccctt ccagcagttc ggtagagaca tcgcggacac gaccgatgca 1740
gtgagagatc ctcagacatt ggaaatcctc gatattacac cgtgcagttt cggcggagta 1800
agtgtgataa caccaggcac caacacaagc aatcaggtcg ccgtgctgta tcaagatgtg 1860
aattgtaccg aggtccccgt ggcaattcac gcagaccaac tcacccccac ctggagagta 1920
tactccacag ggtccaatgt ctttcagact cgggccggat gtctcatcgg agctgaacac 1980
gtgaataata gttatgaatg tgacatccca atcggcgccg gaatttgtgc atcttatcaa 2040
actcagacaa actctccacg agccgctgca agtgttgcat cacagtccat aatcgcatac 2100
actatgagct tgggcgcgga aaactccgtc gcctattcca ataactcaat tgctattccc 2160
acaaatttca cgattagcgt gaccacagaa atcctgcctg tgagtatgac taagacttcc 2220
gtggactgca ccatgtacat ttgcggagac agcactgaat gtagcaacct gctcctgcaa 2280
tacggtagtt tttgcacaca actgaatagg gcactgacgg gcatcgcggt agaacaggac 2340
aagaatactc aggaggtgtt tgcacaagtg aagcagatat ataaaactcc cccaattaag 2400
gacttcggcg gtttcaactt ctcacagatc ttgcccgatc cttcaaagcc atccaaaagg 2460
agcttcatcg aggaccttct tttcaacaag gtgacactgg ccgatgccgg gttcattaag 2520
cagtacgggg attgtcttgg cgacatcgct gccagagacc tgatctgcgc ccagaagttc 2580
aatggcctca cagtgctccc tccattgctt actgacgaaa tgatcgctca atacacatct 2640
gccctgctgg ccggcactat tacatctgga tggacatttg gtgccggcgc cgcactgcaa 2700
attccattcg ctatgcagat ggcatatcgc tttaacggca tcggagtgac ccagaacgtg 2760
ctgtacgaga atcagaaatt gattgctaac cagttcaaca gtgccattgg caagattcag 2820
gattcactga gcagcacagc cagtgcactt ggtaagctcc aggatgtcgt caatcagaat 2880
gctcaagccc ttaacacctt ggtgaaacag ctttcctcaa attttggtgc gatctctagc 2940
gtactgaatg acatactttc tcgcctcgac aaggttgagg ccgaggttca gatcgatcgg 3000
ctcatcaccg gaagactgca gagcctccag acctatgtga ctcagcagct gataagagca 3060
gctgagattc gcgcttctgc taatttggcc gcgacaaaga tgtcagagtg tgtgctgggc 3120
cagtctaaaa gagttgattt ttgcggcaaa ggataccacc tgatgtcctt cccccaatca 3180
gctccgcacg gggtggtctt tctgcacgtt acttatgtcc cagcagagaa gaacttcacg 3240
acagcacccg caatctgtca tgacggaaag gcccactttc cgagagaagg agtctttgtg 3300
tccaatggga cccactggtt cgtgacacaa agaaattttt acgaacccca aatcattacg 3360
actgataaca ccttcgtgag tggcaactgc gatgttgtta taggcatcgt taataacacg 3420
gtttacgatc ccttgcagcc ggaactggat tctttcaaag aagagctgga caagtacttc 3480
aaaaaccaca catctccaga cgtggatctg ggcgatataa gtggtataaa cgcaagtgta 3540
gtaaacatac agaaagaaat cgatagactg aacgaggttg caaaaaacct taacgaaagt 3600
ctcatcgatc ttcaggagct cggcaagtac gagcaatata ttaagtggcc atacattccc 3660
gaagcaccac gcgatggaca ggcttacgtg cgcaaggacg gtgaatgggt cttgctttct 3720
acgttcttgc accatcacca ccaccat 3747
<210> 30
<211> 3759
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 30
atgccgatgg gcagtctgca accattggct actctctatc tccttggcat gctcgtagcc 60
tcatgcctgg gcgtgaatct gacaacgcgg actcagctcc ctccagctta tactaactcc 120
tttacgagag gtgtgtacta cccagacaag gttttccgga gctcagtgct gcatagcacg 180
caggacttgt tcctgccttt cttctcaaac gtgacttggt tccacgctat ccatgtgagc 240
ggcaccaacg gtactaagag atttgataat ccggttctcc cattcaacga tggcgtgtac 300
ttcgcctcaa cggagaagag taacattatt cggggctgga tttttggaac cactctcgac 360
tcaaagactc agtctctgct gatagtgaac aatgctacca atgttgtcat caaagtgtgt 420
gagttccagt tctgcaatga tccattcctg ggcgtttatt atcacaaaaa taataagtca 480
tggatggagt ctgaattccg cgtgtacagt agcgccaaca actgtacttt tgaatatgtg 540
agtcaacctt ttctgatgga cctggagggg aagcagggta atttcaagaa cctgagagaa 600
tttgtgttta agaatataga tgggtatttt aaaatttact ccaagcacac gcccatcaac 660
cttgtcagag atctcccaca agggttctcc gcgctggaac ctctggttga cctgcccatc 720
ggaatcaaca tcaccaggtt ccaaacgctg ttggccttgc atcggtctta cctgaccccc 780
ggtgacagta gttctggttg gactgcagga gcggccgctt actacgtggg gtacctgcaa 840
ccccgcacct ttctgttgaa gtacaatgag aatgggacca taactgacgc tgtggattgt 900
gctcttgacc ccctgagcga gaccaaatgc acccttaaat cattcactgt ggaaaaggga 960
atttaccaga catctaactt cagagttcag cctacagagt ctatcgtccg gttccccaac 1020
ataaccaacc tctgcccgtt tggcgaggtg tttaacgcta cgagatttgc gtcagtgtac 1080
gcttggaata gaaaacgaat ctctaattgc gtggctgact actccgttct gtacaatagc 1140
gcaagtttta gcaccttcaa gtgctacgga gtttcaccaa caaagcttaa cgacctgtgt 1200
tttaccaacg tgtacgccga cagttttgtt attagaggtg atgaagtaag acagatcgcc 1260
ccgggccaga ctggcaaaat tgcggattat aactacaagc ttccagatga tttcacagga 1320
tgcgtgatcg cctggaattc aaacaacctc gacagcaagg tgggcgggaa ctacaactac 1380
ttgtaccgac tgtttcgcaa gtctaacctg aaaccgttcg aacgggatat ttccacagaa 1440
atctaccagg ccgggtctac tccatgcaac ggcgtcgaag gatttaactg ttatttccca 1500
ctgcagagct atggattcca gccaacgaat ggcgtgggtt atcaacccta ccgcgttgtg 1560
gttctgtcat ttgagctgct tcatgctcca gccaccgtct gcggtcctaa aaaatccaca 1620
aatcttgtga agaacaaatg tgttaatttc aacttcaatg ggcttacagg cacaggggtt 1680
ctcacagagt ctaacaaaaa gttcctgccc tttcagcagt ttggtcgcga tatcgcagac 1740
accactgatg ctgtgcgcga cccacagacg ctggaaatcc ttgatatcac tccctgttca 1800
ttcggcggag tcagtgtgat cacgccagga accaatacct ctaaccaggt ggcggtactt 1860
taccaggacg ttaactgcac tgaggttccc gtcgctattc acgccgacca gcttactccg 1920
acatggagag tttactccac tggttccaac gttttccaga caagggccgg ttgtcttatc 1980
ggagccgaac acgtgaataa cagctatgaa tgcgatatcc ctattggagc cggcatctgt 2040
gctagctatc aaacccagac taactcacca cgagccgctg catcagtcgc cagtcagtcc 2100
atcattgctt atacgatgtc cctcggggcc gagaacagcg tcgcatatag taacaatagt 2160
atcgctatcc ctacaaattt cacaatttcc gtgaccactg aaatactccc agtgtctatg 2220
acgaagacaa gtgtggactg tacaatgtat atctgcggag atagtaccga atgtagtaat 2280
ttgctgctcc aatacggctc attttgtacg cagctgaaca gggcacttac tggaatcgca 2340
gtggagcagg acaaaaatac ccaggaagtc ttcgcgcagg tcaaacagat ctacaagacc 2400
ccgccaataa aggattttgg aggatttaac ttctcccaga tcctgcccga tccctccaag 2460
ccgagtaaga ggtcttttat cgaagatctt ttgtttaaca aagtcacctt ggctgacgcc 2520
ggctttatta agcagtacgg agactgtctg ggcgacattg ccgcaagaga tttgatctgt 2580
gctcagaagt ttaatggtct gacagtgttg cccccactgc tcacagatga gatgatcgcc 2640
cagtatacta gtgctctgct cgccgggacg atcacctctg gatggacatt tggagccggg 2700
gctgcgttgc aaatcccgtt cgcaatgcag atggcttacc gatttaacgg cataggggtg 2760
acgcaaaatg tcttgtacga gaaccaaaag ctgattgcta accagtttaa cagcgctata 2820
ggaaaaattc aagactctct ctcctctacc gcgagcgcac tcggaaagct ccaggatgtg 2880
gtcaaccaga acgcccaagc cctcaatacg cttgtaaagc agctgtcctc caatttcggg 2940
gccatcagca gcgttttgaa cgatattctg agtcggctgg acaaggttga ggccgaggtg 3000
cagatcgata gactcatcac aggaaggctg cagtcattgc agacctacgt gacacagcag 3060
ctgattaggg ccgccgaaat cagggcttct gccaaccttg ccgctactaa gatgagtgag 3120
tgtgtgctcg gccagagcaa acgagtggac ttctgcggta agggctatca cctgatgtca 3180
tttccgcagt ccgcacctca tggtgtggtc ttcctgcatg tgacatacgt gccagcagaa 3240
aagaatttta ccaccgcgcc agccatttgc catgacggga aggctcattt ccctagagaa 3300
ggcgtcttcg tgagcaatgg gacacactgg ttcgttacac agagaaactt ttacgaaccc 3360
cagattatca cgacagacaa cacgttcgtt tccggaaact gtgatgtcgt catcggcata 3420
gtgaacaata ccgtctacga ccccctgcaa ccagagctgg attcattcaa agaagagctg 3480
gacaaatact tcaagaacca tacatctcct gacgttgatc tcggggacat aagtggcatc 3540
aacgctagcg tggtaaatat ccaaaaggaa attgatagac tgaatgaagt cgccaagaat 3600
ctcaacgaaa gcctgattga cttgcaggaa ctggggaaat acgagcaata cattaagtgg 3660
ccctacattc ctgaagctcc ccgggacgga caagcttatg tcagaaagga cggcgagtgg 3720
gtgctcctca gcaccttcct gcatcaccat caccaccac 3759
<210> 31
<211> 3777
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 31
atggatagca agggcagcag ccagaaagga tctcgactgc tgcttctgct ggttgtttcc 60
aacctgctcc tttgccaggg tgtggtgagc gtaaacctga ctacccgaac acagctcccg 120
ccagcataca ccaattcctt cacacggggg gtgtattatc cagataaagt cttcagaagt 180
agtgtactgc attcaaccca agacctgttc ctgccatttt tttccaacgt gacctggttt 240
cacgccattc acgtttcagg aacaaacggg actaaaagat ttgataaccc cgtgctcccg 300
tttaacgatg gggtgtactt cgctagtaca gagaagtcca acatcatcag gggctggata 360
ttcggaacaa cgctggactc caagacccag agtctcctga ttgtgaataa tgctactaat 420
gtggtcatta aggtgtgcga gttccagttc tgcaacgacc cttttctggg ggtatactac 480
cataagaata ataaaagctg gatggagtct gaattcagag tatacagctc cgccaacaat 540
tgcacattcg aatacgtcag ccagccattc cttatggatc tggagggtaa gcaggggaat 600
ttcaagaacc tgcgagaatt tgtattcaag aatattgacg gctattttaa gatctactca 660
aagcacacgc ctatcaatct cgtgagggac cttccgcagg gattctctgc actggaaccg 720
ctcgtggatc tgccgattgg aattaacatt acgaggttcc agactctcct tgctttgcat 780
aggtcctatc tgacacccgg agatagttct agcgggtgga cagctggcgc agctgcttat 840
tatgtcggat atctgcaacc aaggaccttc ctcttgaagt ataacgagaa cggcaccatt 900
acagacgcag tcgactgcgc tcttgatcct ctcagcgaaa cgaagtgcac gctcaagtct 960
tttactgtgg agaagggcat ctatcagacc tccaacttta gggtacagcc cacagaatcc 1020
attgttaggt ttccaaacat cacaaatttg tgtccttttg gagaggtgtt taatgctaca 1080
agattcgcca gcgtctacgc gtggaacaga aagcgaatat caaactgtgt agccgactac 1140
tctgtgctct acaatagcgc cagcttcagc acctttaagt gctacggagt cagcccgaca 1200
aaactgaacg atctttgctt cacaaatgtg tatgctgact ccttcgtcat tcgcggcgat 1260
gaggtgagac agatcgcccc agggcagaca ggaaaaatcg ctgattataa ttacaaactt 1320
cccgacgact ttacagggtg cgttatcgca tggaactcca ataatcttga ttctaaagtg 1380
ggaggaaact acaactactt gtacaggctg tttcggaaga gcaatctgaa gcccttcgag 1440
agagacatta gtactgagat ctaccaggca ggttctacac cttgcaatgg cgtggagggg 1500
ttcaactgtt actttcccct tcagtcatac ggttttcagc caaccaatgg cgtcggctac 1560
cagccttaca gagttgtcgt actgtccttc gagctgctgc acgcacctgc tactgtgtgc 1620
ggtcctaaaa agtcaacaaa tctggtcaaa aataaatgtg tgaacttcaa ctttaacgga 1680
ctgactggaa ctggtgtcct taccgaaagc aacaagaaat ttcttccatt ccaacagttc 1740
gggcgggata tcgcagacac aactgatgcc gtccgcgacc cccagactct ggaaatactg 1800
gatatcacgc cttgctcatt cggcggcgtc tcagttatta ctcctgggac caatacaagc 1860
aaccaggtgg ctgtactgta tcaggatgtt aattgtacgg aggtgcccgt ggctattcac 1920
gctgatcaac tcactcctac ttggcgcgtg tacagcaccg gcagtaacgt atttcaaaca 1980
agagctgggt gcctgattgg ggctgaacac gtcaataatt catacgagtg tgatatccct 2040
attggcgccg ggatctgcgc gagttatcag acgcaaacta attcaccaag ggctgcagct 2100
agcgtggcct cacagagtat catagcttac actatgagcc tcggagccga gaacagtgtt 2160
gcttacagca ataacagtat cgccatccct acaaacttta ctatctctgt gacgactgaa 2220
atacttcccg tgtctatgac aaaaacctct gttgattgta caatgtatat ctgtggggat 2280
agcaccgagt gttctaacct cctgctgcag tacggtagtt tctgtacaca gcttaatcgc 2340
gctctgacag gaattgcggt cgagcaggat aagaacacgc aagaggtgtt cgcacaggtg 2400
aagcagatat ataagacccc tcccatcaaa gattttggag ggtttaattt tagtcagatt 2460
ctccccgacc cgtccaagcc ttcaaagcgg tcttttatcg aggacctgct gtttaacaag 2520
gtgaccttgg cagatgccgg tttcatcaaa caatatggag actgcttggg cgatattgct 2580
gcgcgggacc tgatttgcgc ccagaagttc aacgggttga ctgtcctgcc gcctctgctg 2640
accgatgaga tgattgctca atacaccagc gctcttctgg caggcaccat tacttccggc 2700
tggacatttg gagctggtgc tgcgctgcag atcccctttg ccatgcagat ggcttacaga 2760
ttcaacggga tcggagtgac ccaaaacgtt ctgtacgaga accagaaatt gattgctaac 2820
caattcaaca gcgccattgg taagatccag gattccctgt cctctaccgc cagtgcactt 2880
gggaaactcc aggatgtagt gaaccagaac gcccaggccc tcaacactct ggtgaaacag 2940
ctgagctcta acttcggcgc aatttccagc gtactgaacg acattctctc tagactggat 3000
aaagtcgagg ctgaagtcca gatcgacagg ctcattaccg gcagactgca gtccctgcag 3060
acatatgtaa cccagcagct gatcagggca gccgaaatcc gcgccagtgc taacctggcc 3120
gctacaaaga tgtccgagtg tgtgctggga cagtcaaagc gcgtggactt ctgcgggaaa 3180
gggtaccacc ttatgagttt tcctcagtct gctccccatg gggttgtgtt tctccatgta 3240
acatatgtcc ctgctgagaa aaattttaca acggctccag ccatttgtca cgatgggaaa 3300
gcccacttcc cccgagaagg cgtgtttgtt tcaaacggaa ctcattggtt tgtcacacag 3360
cgcaacttct atgagccaca gataatcacc acggacaaca catttgtgtc tgggaactgt 3420
gatgtggtca tcggaatcgt aaacaacaca gtctacgatc cccttcagcc tgagctggac 3480
tcctttaaag aggagctgga caagtacttc aagaaccaca cttctcccga cgtggacctg 3540
ggcgatatca gtggcatcaa cgccagtgtg gtcaatatcc agaaggaaat tgacaggctc 3600
aacgaagtag ccaagaatct gaacgaatca ctgattgatt tgcaggaact ggggaagtac 3660
gagcagtata tcaagtggcc atacatccct gaagccccta gggatggtca agcatatgtt 3720
agaaaggacg gggagtgggt tctgctgagc acctttctcc accatcatca tcatcac 3777
<210> 32
<211> 3765
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 32
atggcagctc cggcactgtc atggagattg cccctcttga ttttgttgtt gcctcttgct 60
acatcttggg cctccgcagt caatctcaca accaggactc aactgcctcc cgcctacacc 120
aattccttta ctagaggcgt ctactacccc gataaagtgt tcaggtccag cgtgctgcac 180
agcacccaag atctgtttct gccattcttt agtaatgtca cctggtttca tgccattcac 240
gtttctggaa cgaacggaac taagcgcttt gataatcctg tcctgccctt taacgatggt 300
gtgtactttg cctctactga gaaaagcaac attatccggg gatggatctt cggcactaca 360
ctggactcaa aaacacagag tttgctgatc gttaataacg cgaccaacgt ggtaattaag 420
gtgtgtgagt ttcagttttg taacgatccc ttcttgggag tgtactatca caagaataac 480
aagtcttgga tggaatccga attcagggtc tacagctccg ccaataattg cacatttgag 540
tatgtaagcc agccctttct gatggatctc gaaggcaaac agggcaactt caagaatctg 600
agggagttcg tgtttaaaaa cattgacgga tacttcaaaa tttatagcaa gcatactccc 660
atcaatcttg ttagagatct tcctcaaggg ttttccgctt tggagccatt ggtcgatttg 720
ccaatcggga tcaacatcac tagattccag acactgctcg ctctccatag atcctatctg 780
acaccaggtg actcttcttc cggctggaca gctggagctg cagcctacta tgttggctat 840
ctgcaacctc gaaccttcct tctgaaatac aatgaaaatg ggaccatcac agatgctgtt 900
gattgtgccc tcgaccctct gtccgagaca aaatgtaccc tcaaatcttt cactgtcgag 960
aaaggcatct accagacttc taattttaga gtgcagccaa ccgagtccat tgttcgcttc 1020
ccgaatataa caaatctctg cccatttggc gaagtcttta atgcaactcg gtttgcgtca 1080
gtttacgcgt ggaaccgcaa gagaattagc aattgcgttg ccgattactc tgtactgtat 1140
aacagcgcca gcttttccac tttcaagtgt tacggggtgt ctccaaccaa gctgaatgat 1200
ctctgtttta ccaacgtcta cgccgactct tttgtgatta ggggtgatga ggtccgccag 1260
atagctcctg ggcaaacagg aaaaatcgct gactacaact ataagcttcc tgacgacttt 1320
accgggtgcg ttatcgcctg gaacagcaac aacctggata gcaaggtggg gggcaattac 1380
aactacctgt acaggctgtt caggaagagc aacctcaagc cattcgagag agacattagc 1440
acggaaatct accaggctgg atctacgccc tgtaacggag tggagggttt caactgttat 1500
tttccactgc aaagctatgg cttccagcca accaatggag tcggttatca gccctaccgc 1560
gtcgtcgtcc tgtcattcga gctccttcat gctccggcca ccgtttgcgg ccctaagaag 1620
tcaaccaatc tcgtgaaaaa taaatgcgtt aactttaatt tcaatgggct cactggaact 1680
ggcgttttga ccgaatcaaa taagaagttc ctgccctttc agcagtttgg ccgagacatc 1740
gcagacacca ctgacgcagt gcgcgatcca cagacacttg agattctcga catcacacca 1800
tgttcattcg gtggcgtgtc cgtgatcaca ccaggaacca ataccagcaa ccaggtagca 1860
gtgttgtatc aggatgttaa ctgtacggag gtccccgtgg ccattcatgc tgaccagctt 1920
acccccacat ggcgggtgta cagtaccgga agcaatgtct tccagacgcg ggccggttgt 1980
cttattgggg cggaacatgt gaacaactct tacgagtgcg atatccctat aggcgctgga 2040
atttgcgcct cataccagac ccaaacaaac tcccctcgcg ccgctgccag tgttgcatct 2100
cagtccatca tcgcctatac tatgagcttg ggtgccgaaa acagtgtggc atattccaac 2160
aacagtatcg ctattcccac caacttcaca atatctgtga caaccgagat cctgcccgta 2220
agtatgacta agacttcagt ggactgcacc atgtacatat gcggtgactc tacagaatgc 2280
tctaacctgc ttcttcagta tgggagcttc tgcacacagc tcaacagggc tctgacagga 2340
atcgctgttg agcaagataa gaacactcag gaggtctttg cacaggtgaa gcagatatat 2400
aaaacacccc caatcaagga cttcggagga ttcaacttca gtcagatact gcccgaccca 2460
tctaagccat ccaagcgcag cttcatcgaa gacctccttt ttaataaagt cacactcgct 2520
gacgcggggt tcattaagca gtacggagat tgtcttggtg atatcgctgc tcgggacttg 2580
atttgcgctc agaaatttaa cggtcttacc gtgctgccac cactgctgac cgacgagatg 2640
atcgcccagt acaccagcgc acttcttgca ggaacaatca catcaggatg gaccttcggt 2700
gctggggctg cactgcagat accttttgcc atgcaaatgg catatcggtt caacggaatc 2760
ggagtcacgc aaaacgtgct gtacgaaaac cagaagctca ttgctaacca gttcaatagc 2820
gccatcggaa aaattcagga ttctctgtca agcaccgctt ccgcactggg taagctgcag 2880
gatgtagtca atcagaatgc ccaggctctt aacacccttg ttaaacagct ctcatccaac 2940
ttcggtgcta tttctagcgt gctgaacgat atcctgagcc ggctggacaa agtggaagcg 3000
gaagtgcaga tcgaccgact gatcacaggc agacttcaga gcctgcagac ttatgtcacc 3060
cagcagctga ttagggctgc tgaaatcaga gcaagcgcaa acctggctgc aaccaagatg 3120
tccgagtgcg tattggggca aagcaagaga gttgacttct gcgggaaggg ctaccacctg 3180
atgagtttcc cacagagtgc accccacggg gtggtattcc tccatgtgac ctatgtgcct 3240
gctgaaaaga actttactac cgccccagcc atttgtcacg atgggaaagc tcacttcccc 3300
cgcgaagggg tcttcgtctc caatggaact cactggtttg tcacacagcg caatttttac 3360
gagccgcaga tcattactac cgataacacg ttcgttagcg gaaactgcga cgtagtgatt 3420
ggcatagtga ataacactgt ctacgatcct ctgcaaccgg agctggactc cttcaaagaa 3480
gagctggata agtacttcaa gaatcatacc tcacccgatg tcgatctcgg ggatatctcc 3540
ggtatcaatg ccagcgtagt gaatattcaa aaagaaatag ataggctgaa cgaggttgct 3600
aagaacctga atgaaagcct catagacctg caagaactgg gcaaatatga acagtacatt 3660
aagtggccct acatccccga ggcaccaagg gatgggcagg cttatgtgcg gaaagatgga 3720
gagtgggtgc ttctctcaac tttcctccac caccatcatc accac 3765
<210> 33
<211> 3738
<212> DNA
<213> Artificial Sequence
<220>
<223> 编码重组Spike蛋白的核苷酸序列
<400> 33
atgagggctt ggatattctt tctgctgtgt ttggctggac gggcactcgc tgttaatctg 60
accacaagga cgcagctgcc acccgcttat accaattctt ttacccgggg ggtgtattat 120
cccgacaagg tttttagatc ttctgttctg cacagcaccc aggacctttt cctgccgttc 180
ttctcaaacg taacctggtt tcatgcaatt cacgtgtctg gaacaaatgg tactaaacgg 240
ttcgataacc ctgttcttcc ttttaacgac ggtgtgtatt ttgcaagtac cgagaagtct 300
aacataatcc gaggttggat ttttgggacg accctggata gcaagacaca gtcccttctc 360
atcgtcaata acgcaacgaa cgttgtaatt aaggtgtgtg aatttcagtt ttgcaatgac 420
cccttccttg gtgtgtatta tcataaaaac aacaaaagtt ggatggagag cgaatttagg 480
gtctatagtt cagctaataa ttgcacattt gaatacgtat ctcagccctt cttgatggat 540
ctggaaggta aacaaggcaa cttcaagaac cttagggagt ttgttttcaa gaatattgat 600
ggttatttta agatttactc taaacatacc cccatcaatt tggtgcggga tttgccacag 660
ggattcagtg ccttggaacc cctggtcgac cttccaatcg ggattaatat tacacgattc 720
caaaccctct tggcactgca ccggtcctac ctgacacccg gagactcatc ttcaggctgg 780
acggcaggag cagcggctta ttacgtcggt tatctccagc cgaggacctt tctccttaaa 840
tataatgaga acggtacaat cacagatgcc gttgactgtg ccctcgaccc cctgtcagag 900
accaagtgta ctttgaagag ttttactgtc gagaagggca tctatcagac ttctaatttc 960
cgagtgcagc caactgagtc catagtcaga ttcccaaaca tcactaatct ttgcccattc 1020
ggcgaagttt tcaacgcgac aagattcgct tctgtatacg cgtggaatag gaagcggatc 1080
agcaattgtg tagctgacta cagtgtgctg tataactcag cctcttttag tactttcaag 1140
tgctacggcg tgtccccaac taagctgaat gacctgtgct tcacaaacgt gtacgctgat 1200
agcttcgtca tccgcgggga tgaggtcaga cagatagctc ctggccaaac tgggaagatt 1260
gccgactaca actataagct gcccgatgac ttcaccgggt gcgtcatcgc ttggaactca 1320
aacaatttgg actcaaaagt aggtgggaat tacaactacc tttacaggct ctttcggaaa 1380
agtaacctca aacccttcga gagagacatc agtaccgaga tataccaggc tgggtctact 1440
ccatgcaatg gggttgaggg atttaactgt tattttccct tgcaaagtta tggcttccag 1500
cctaccaacg gggttgggta tcagccgtac agggtggttg tgctgagctt cgaactcctc 1560
catgcgccag ctacagtttg tggaccaaaa aagagtacaa accttgttaa aaacaagtgc 1620
gtgaatttca acttcaacgg acttactggc actggtgtac tgacagaatc caacaaaaag 1680
ttcctccctt tccagcaatt tggaagagac attgctgata ccactgatgc agtaagggac 1740
ccccaaacct tggagatcct ggacattaca ccttgctcat tcggcggcgt ttcagtcatc 1800
acacccggga caaacacaag caaccaggtg gctgtgctct atcaggacgt gaattgcaca 1860
gaggtccccg ttgccattca tgcagatcaa ctcaccccta catggagggt gtactctact 1920
ggatctaacg tgtttcagac acgcgccggg tgtcttatag gcgctgagca cgtgaataat 1980
agctatgagt gcgacatccc tatcggtgct ggcatctgtg catcctacca gactcagact 2040
aatagtccaa gggctgccgc aagtgtagct tcccagtcaa tcattgcata taccatgtcc 2100
cttggcgccg agaatagtgt cgcatactct aataacagca ttgcaatccc caccaatttt 2160
accatcagtg ttacaacaga gattctgcca gttagcatga caaagacatc cgtcgactgt 2220
accatgtaca tctgtggcga tagcaccgag tgttccaatc tcctcctgca gtacgggtcc 2280
ttttgtactc agctcaacag ggctcttact ggtatagcag tagagcagga taaaaacacc 2340
caggaggtct ttgcccaggt taaacagata tacaagaccc cgcctatcaa ggattttgga 2400
ggattcaact tttctcagat actgccagat ccaagtaagc catctaaacg gtctttcatt 2460
gaggacttgc tgttcaacaa ggttaccctt gctgacgctg gattcattaa acagtacgga 2520
gattgtctgg gtgacattgc cgcccgcgac ttgatctgtg cacagaaatt caatggactg 2580
actgtgctcc ctcccctcct gactgacgag atgattgctc agtatacgtc cgcactgttg 2640
gcaggaacta ttacatctgg atggactttt ggagccggag cggctctgca aatacccttt 2700
gcaatgcaga tggcctacag gttcaacggg atcggagtga cccagaatgt actttatgag 2760
aaccagaagc tgattgccaa tcaattcaac agcgccatcg gcaaaataca ggactctctg 2820
agctctactg cttctgcgct cggcaagttg caagacgtgg tgaaccagaa cgctcaggcg 2880
ctgaacacac tcgtcaagca gctgtcttcc aatttcgggg ctatctcctc cgtgctgaac 2940
gatatactct ccaggctgga caaggtggaa gccgaggtcc agatcgacag actcatcacg 3000
ggcagacttc agtctttgca gacctatgtt actcagcagc ttattagagc tgccgagata 3060
cgggcctcag cgaatctggc agccaccaaa atgtctgagt gcgtgctggg gcagtccaag 3120
cgagtggact tttgcggaaa ggggtaccat ctgatgagct tccctcagtc agctcctcac 3180
ggcgtggtat tcctgcacgt cacttacgtg cctgccgaaa agaacttcac taccgcccca 3240
gcaatctgcc acgacggcaa agcccacttt cctagagagg gggtttttgt gtctaacggc 3300
acccactggt tcgtcactca gagaaacttt tatgagcccc agataataac cacagacaac 3360
acattcgtgt ctggcaactg cgacgttgtg ataggaatcg tgaataatac cgtgtacgat 3420
cccctgcagc ctgaactcga ttctttcaaa gaggaactgg acaaatactt taagaatcat 3480
acttcccctg atgtcgacct gggtgacatt tcaggtatca acgccagcgt agtgaatatc 3540
caaaaggaga ttgataggct caatgaggtt gcaaaaaacc tcaacgagag tttgatcgac 3600
ttgcaggaac tgggcaaata tgaacagtac ataaaatggc cttatatccc agaagctcct 3660
cgcgatggcc aggcatatgt ccgaaaggac ggtgaatggg ttctgctgag cacctttctt 3720
caccaccatc atcatcat 3738

Claims (10)

1.以下多肽作为信号肽在制备重组新型冠状病毒Spike蛋白中的应用：
SEQ ID No. 10所示氨基酸序列组成的多肽。
2.一种制备重组新型冠状病毒Spike蛋白的方法，该方法包括：
构建含有编码重组新型冠状病毒Spike蛋白的多核苷酸的表达载体，转染哺乳动物细胞以分泌表达重组新型冠状病毒Spike蛋白；
其中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列中包括了编码信号肽的核苷酸序列，所述信号肽为：SEQ ID No. 10所示氨基酸序列组成的多肽。
3.根据权利要求2所述的方法，其中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列除包括编码信号肽的核苷酸序列外，还包括编码SEQ ID No. 1所示氨基酸序列的核苷酸序列。
4.根据权利要求2或3所述的方法，其中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列还包括：编码设计在重组新型冠状病毒Spike蛋白的C端的T4 fibritin三聚体基序的核苷酸序列，所述T4 fibritin三聚体基序包括SEQ ID No. 12所示氨基酸序列；
其中，所述T4 fibritin三聚体基序的C端带有或不带有poly His标签。
5.根据权利要求2所述的方法，其中，所述编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列包括：编码SEQ ID No. 22所示氨基酸序列的核苷酸序列。
6.根据权利要求2所述的方法，其中，编码重组新型冠状病毒Spike蛋白的多核苷酸的核苷酸序列如SEQ ID No. 32所示。
7.根据权利要求2所述的方法，其中，所述哺乳动物细胞为人胚肾293细胞。
8.根据权利要求2所述的方法，其还包括纯化所表达的重组新型冠状病毒Spike蛋白的过程。
9.一种用于制备重组新型冠状病毒Spike蛋白的表达载体，其为权利要求2~8任一项所述方法中所用到的含有编码重组新型冠状病毒Spike蛋白的多核苷酸的表达载体。
10.一种用于制备重组新型冠状病毒Spike蛋白的宿主细胞，其是由权利要求9所述的表达载体转染哺乳动物细胞得到的能分泌表达重组新型冠状病毒Spike蛋白的细胞。
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CN113929786A (zh) * 2021-04-13 2022-01-14 武汉大学 新型冠状病毒突变株s蛋白及其亚单位疫苗
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