CN111607000A - 一种重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白及其制备方法和应用 - Google Patents
一种重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白及其制备方法和应用 Download PDFInfo
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Abstract
本发明公开了一种重组的非洲猪瘟p30亚单位可溶性融合蛋白及其制备方法和应用，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白包括OPTI‑p30蛋白和一种伴侣蛋白，其中所述OPTI‑p30蛋白的核苷酸序列为经优化后的序列OPTI‑p30如SEQ ID NO.1所示。本发明能够在大肠杆菌中直接可溶性表达P30，克服了现有技术中的诸多问题，且制备方法简单、成本低。
Description
技术领域
本发明属于兽用生物制品技术领域。涉及一种重组的非洲猪瘟p30亚单位可溶性融合蛋白，及在制备诊断、预防和治疗非洲猪瘟疫苗中的应用。
背景技术
非洲猪瘟(African swine fever，ASF)是由非洲猪瘟病毒(African swine fevervirus，ASFV)引起的猪的急性、热性、高度接触性传染病。猪是ASFV自然感染的唯一哺乳动物宿主，包括家猪和野猪，尤其是家猪，易感性极高。猪感染非洲猪瘟病毒后临床上以皮肤充血，内脏器官出血，高热为特征，发病率及死亡率高达100％。世界动物卫生组织将其列为A类疫病，我国也将其列为一类动物传染病。
非洲猪瘟自1927年在非洲大陆发现以来，对非洲和欧洲的养猪业造成极大打击。2018年8月份以来，在我国多个省份爆发，给我国养猪业带来了严重的经济损失。尽管已经有许多非洲猪瘟的研究性报告，人民对非洲猪瘟也有一定的了解，但目前世界上尚未有有效预防非洲猪瘟的疫苗和治疗该病的药物发现，迫切需要新型疫苗的研发和生产来预防非洲猪瘟。
ASFV病毒是具有囊膜的虫媒DNA病毒。病毒颗粒呈二十面体对称结构，平均直径200nm，表面有含糖脂类的囊膜覆盖。其病毒基因组为双股线性DNA，大小为170-190kb，整个基因组大约有150个ORF，编码150-200中蛋白质。p30蛋白由CP204L基因编码的，分子量大小约30kD-32kD的蛋白，因此，有时也称为p32蛋白。研究发现，p30蛋白与病毒进入宿主细胞的过程有关，针对p30的抗体能够抑制病毒的内化(Gomez-Puertas et al.,1996)。并且p30蛋白是ASFV的主要结构蛋白，具有很好的抗原性，能诱导机体产生中和抗体p30蛋白能够在感染的动物体内诱导产生中和抗体(Gemoz-Puertas et al.,1996)，通常作为诊断抗原(Oviedo et al.,1997)。因此，p30是一个很好的保护性抗原。目前，P30已经能成功的表达在原核系统，但表达产量低，且以包涵体形式存在(储德文，2013)，无法满足基因工程亚单位疫苗的开发。而利用本发明提供的方法可大量可溶性表达P30蛋白。在当前没有可能大规模制备灭活疫苗或弱毒疫苗的情况下，确定一种制备该病毒的免疫原性蛋白的方法，以便研究一种能够预防该疾病的亚单位疫苗或具有能够诊断该疾病的试剂具有重大的意义。
发明内容
在现有技术的基础上，本发明为了克服现有技术中存在的诸多缺陷(如表达产量低，包涵体表达，纯化困难成本相对较高等)，提供了一种制作成本低廉，可在大肠杆菌中直接可溶性表达p30的核苷酸序列及蛋白制备方法。
根据本发明的一个方面，本发明提供了一种可在大肠杆菌中表达p30蛋白的优化后的OPTI-p30的核苷酸序列。为了能够在大肠杆菌中高效表达P30蛋白，本发明根据GenBank:FR682468.1上已经存在公开的序列(密码子优化前的编码P30蛋白的核苷酸序列如SEQ ID NO.2所示)，我们对p30基因进行分析，首先对p30基因编码的密码子的在原核表达中的密码子偏好性进行了优化，其次对p30基因的GC含量和mRNA在大肠杆菌的稳定性进行优化。所述最终优化后OPTI-p30的核苷酸序列如SEQ ID NO.1所示。因此，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白包括OPTI-p30蛋白和一种伴侣蛋白，其中所述OPTI-p30蛋白的核苷酸序列为p30蛋白经优化后的序列OPTI-p30如SEQ ID NO.1所示。
在本发明的优选技术方案中，所述伴侣蛋白为TF蛋白。
在本发明的优选技术方案中，优化后的OPTI-p30直接在原核表达系统中也不能可溶性表达p30，为了能够得到可溶性表达P30蛋白，本发明在OPTI-p30核苷酸序列3’或5’端加入伴侣蛋白的基因编码序列，其中伴侣蛋白优选为TF，得到OPTI-TF-P30。优选地，5’端加入TF基因编码OPTI-TF-p30的基因编码序列如SEQ ID NO.3所示，其氨基酸序列如SEQ IDNO.4所示。
为了方便使用亲和层析的方法纯化融合蛋白，根据本发明的技术方案，优选地，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白在如SEQ ID NO.4所示的氨基酸序列的氨基末端或羧基末端连接有poly-His、FLAG、c-myc、HA和poly-Arg中的一种标签。
根据本发明的另一个方面，本发明提供了一种重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白的制备方法，所述制备方法包括以下步骤：1)将含有OPTI-p30蛋白和一种伴侣蛋白的基因克隆到原核表达载体中，得到含有非洲猪瘟病毒OPTI-p30蛋白和一种伴侣蛋白编码基因的重组质粒；其中，所述OPTI-p30蛋白的核苷酸序列为经优化后的序列OPTI-p30如SEQ ID NO.1所示；2)再将步骤1)所述重组质粒转化至大肠杆菌菌株的细胞中，得到重组表达的菌株；3)通过培养、筛选步骤2)中所述重组表达的菌株，得到高度表达的菌株；4)发酵培养步骤3)中所述高度表达的菌株，纯化后得到重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白。
在本发明的优选技术方案中，步骤1)中，所述伴侣蛋白为TF蛋白。
在本发明的优选技术方案中，含有非洲猪瘟病毒OPTI-p30蛋白和TF蛋白的核苷酸序列OPTI-TF-p30是OPTI-p30蛋白的5’端插入TF蛋白的核苷酸序列，所述OPTI-TF-p30蛋白的核苷酸序列如SEQ ID NO.3所示。
在本发明的优选技术方案中，含有非洲猪瘟病毒OPTI-p30蛋白和TF蛋白的的氨基酸序列如SEQ ID NO.4所示。
在本发明的优选技术方案中，含有非洲猪瘟病毒OPTI-p30蛋白和TF蛋白在如SEQID NO.4所示的氨基酸序列的氨基末端或羧基末端连接有poly-His、FLAG、c-myc、HA和poly-Arg中的一种标签。
在本发明的优选技术方案中，为了能够在大肠杆菌中高效可溶性表达P30蛋白，本发明将OPTI-TF-p30-6His核苷酸序列导入原核表达载体。所述表达载体可以是任何一种原核表达载体，所述的表达载体具体但不限于pET30a表达载体、pBAD载体、pcold载体、pQE载体、pKK载体等。更优选地，所述载体为pET30a载体。
在本发明的优选技术方案中，步骤2)中，所述大肠杆菌菌株选自BL21(DE3)、BL21star(DE3)、arcticexpress中的一种菌株。更优选地，步骤2)中所述大肠杆菌菌株使用BL21(DE3)。
在本发明的技术方案中，优先地，所述的步骤3)中使用IPTG的浓度为0.1mmol/L。
在本发明的技术方案中，优先地，所述的步骤4)中使用裂解液：20mM Tris(pH8.7)，500mM NaCl，0.2％TritonX-114，5％甘油，1mMβ-ME。
根据本发明的再一个方面，本发明提供了一种所述非洲猪瘟病毒p72亚单位可溶性融合蛋白在制备诊断、预防和治疗非洲猪瘟的疫苗中的应用。
与现有技术相比，本发明公开的表达序列、表达载体以及相应的纯化制备方法克服了现有技术中的缺陷，解决了大肠杆菌中不能直接大量可溶性表达P30蛋白且产量低的问题。本发明能够在大肠杆菌中直接可溶性表达P30，克服了现有技术中的诸多问题，且制备方法简单、成本低。
附图说明
图1 P30蛋白编码的核苷酸优化前后序列比对结果。
图2 pET30-OPTI-TF-p30-6His构建示意图。
图3 pET30-OPTI-TF-p30-6His酶切鉴定结果。1-4：pET30a-OPTI-TF-P30质粒StyI酶切，条带大小分别为5199bp,2118bp，质粒酶切正确。M:DL10000Marker
图4 SDS-PAGE检测TF-P30蛋白诱导表达的结果。M是蛋白Marker 26619；1是诱导前上清；2是诱导前沉淀；3是诱导后上清；4是诱导后沉淀。
图5 SDS-PAGE检测TF-P30蛋白纯化后的结果。
图6 SDS-PAGE检测第十次取样4℃和-20℃保存的TF-P30蛋白。1是蛋白Marker26619，2，3:第十次样品4℃和-20℃蛋白样品，上样量3ug
具体实施方式
以下将结合附图和实施例对本发明做进一步说明，本发明的实施例仅用于说明本发明的技术方案，并非限定本发明。
使用试剂均为国产市售产品。
实施例1 P30蛋白表达及制备
1.1非洲猪瘟P30蛋白的选择
非洲猪瘟结构蛋白P30是由CP204L基因编码的一段多肽，P30蛋白合成的动力学表明该蛋白在感染早期被翻译，已有研究表明，p30蛋白能够在感染的动物体内诱导产生中和抗体(Gemoz-Puertas,P.,1996；Oviedo,J.M.,1997)。因此，利用p30蛋白作为抗原具有很好的防控非洲猪瘟的感染或作为诊断试剂检测是否猪感染了非洲猪瘟。尽管P30蛋白报道在多个系统中存在表达，但是，目前还没有报道在原核表达系统中能可溶性表达及纯化得到该蛋白，这也是本发明所要解决的一个重要的技术问题。
为了使所述亚单位TF-P30蛋白便于纯化，可在如SEQ ID NO.4所示的氨基酸序列的氨基末端或羧基末端连接如表一所示的标签，本实施例中具体以在羧基端添加Poly-His为例，将其连接于如SEQ ID NO.4所示的氨基酸序列的羧基末端处。
表一 标签及其氨基酸序列
1.2非洲猪瘟P30蛋白密码子优化
本实验室以2018年在中国报道流行的非洲猪瘟毒株亚型，参考Georgia 2007/1全基因序列(GenBank：FR682468.1)为模板，对编码非洲猪瘟P30蛋白的CP204L的核苷酸序列进行密码子优化，得到OPTI-p30序列，如SEQ ID NO.1所示，该序列合成工作委托南京金斯瑞生物科技有限公司完成。如图1所示，优化前后有27.3％的核苷酸序列不同。
1.3表达载体构建及验证
1.3.1 PCR扩增目的片段
1.3.1.1 PCR反应
(1)引物设计及合成
上游引物:5’-tccgaattcGAGAACCTGTACTTCCAGGGTGACTTCATCCTGAAC-3’
下游引物:5’-agactgcagTTAGTGGTGGTGGTGGTGGTGGAT-3’
(2)加样体系50μL，如下表所示：
1.3.1.2 PCR扩增程序：
1.3.1.3 PCR产物进行胶回收：按照试剂盒说明书操作(可使用天根生化科技有限公司货号为DP214-02的胶回收试剂盒)。
1.3.2 PCR产物及载体双酶切反应
(1)标记好需要用到的200μL PCR管，在PCR管中按照下表进行加样、混匀：50μL反应体系
(2)将步骤(1)中的PCR管置于相应酶最适温度恒温水浴锅中，水浴1-2h。
双酶切产物胶回收：取出上述双酶切体系，进行琼脂糖凝胶电泳以回收其中的DNA片段，方法同1.3.1.3中PCR产物胶回收。
1.3.3连接反应
(1)准备洁净的200μL PCR管若干，做好标记，置于EP管架上待用。
(2)在200μL PCR管按照下表进行加样、混匀。
(3)按照步骤(2)中表格完成加样后，将每个10μl反应体系置于PCR仪16℃反应2h；
(4)取出步骤(3)中的EP管，置于4℃保存。
1.3.4转化反应
将10μL连接反应液快速加入100μL感受态细胞中，并吹打混匀，冰浴30min；取出样品管，置于42℃水浴100s，然后立即冰浴2min；取出样品管，在超净工作台中，向样品管中加入600μL液体LB培养基，然后将样品管置于37℃恒温摇床，220rpm/min，培养1h；涂板：取出上步骤中样品管，室温离心8,000rpm/min，2min，去掉600μL上清液体，剩余上清液重悬管底部的菌体，将重悬的菌液放入相应的转化平板中心，用涂菌棒将转化平板中心的菌液均匀铺开；将上步骤平板正置于生化恒温培养箱中，37℃培养1h后，将转化平板倒置进行培养15h；观察转化结果。转化后阳性质粒命名为pET30a-OPTI-TF-P30。质粒构建示意图如图2所示。
1.3.5质粒抽提与双酶切鉴定
1.3.5.1质粒抽提：按试剂盒照说明书进行操作(可选用生工生物工程(上海)股份有限公司，货号为SK8192的质粒小提试剂盒)。
1.3.5.2双酶切鉴定
(1)标记好需要用到的200μL PCR管，按照下表进行加样：10μL反应体系
(2)将步骤(1)中的200μL PCR管10μL反应体系置于37℃恒温水浴锅中，水浴1h。
(3)将步骤(2)中的酶切体系样品进行琼脂糖凝胶电泳，检查插入片段大小是否正确；实验结果见图3所示，1-4：pET30a-OPTI-TF-P30质粒StyI酶切，条带大小分别为5199bp,2118bp，质粒酶切正确。
(4)选择插入片段正确的克隆送测序公司测序。
1.4 P30蛋白表达
1.4.1转化大肠杆菌BL21(DE3)
吸取1μL质粒加入100μL BL21(DE3)感受态细胞中，冰浴30min；42℃热激90s；冰浴2min；在超净台内加入500μL无抗性的LB培养液；37℃220rpm摇1h；吸取100μL菌液涂卡那抗性LB平板，37℃过夜培养，挑取单克隆并加入甘油于-80℃保存备用。
1.4.2小量诱导表达
甘油管保藏管菌株活化：取E.coli BL21(DE3)pET30a-OPTI-TF-P30菌株甘油保藏管解冻，用接种环挑取甘油管中菌悬液于卡那抗性平板(50μg/mL)上划线，37℃培养过夜。挑菌活化：于培养好的平板上挑取单克隆至3mL卡那抗性LB培养基中，37℃220r/min摇床培养3.5～5h，至OD600达到0.5～0.8。发酵接种：将活化好的菌悬液取以1：100的比例接种至15mL卡那抗性LB培养基中，37℃，220r/min培养。降温诱导：当OD600达到0.6-0.8时，取出5mL菌液，12000rpm离心5min，将菌体置于-20℃保存，即为“诱导前”。剩余菌液置于冰水浴10min，加入0.4ml 1M IPTG，终浓度0.2mM IPTG。将摇床温度降至20℃，诱导15h。菌体收集：发酵结束后，测量OD600，收集与“诱导前”等量菌体，12000r/min离心5min，收集菌体置于-20℃保存，即为“诱导后”。
诱导表达SDS-PAGE检测结果如图4所示，箭头指向为TF-p30蛋白。从图中可以看出，我们制备的TF-P30蛋白为95％为可溶性表达。
另外，参照1.3的步骤，我们也构建了pET28a-OPTI-p30，pQE30Xa--OPTI-p30，pColdIII--OPTI-p30，pBAD30--OPTI-p30等并参照1.4的步骤将该质粒进行了转化和小量诱导表达，发现这些和文献报道基本一致，基本上是不可溶表达，即在表达形成了包涵体。另pET30a-OPTI-MBP-p30、pET30a-OPTI-SUMO-p30，pET30a-OPTI-GST-p30等含有不同伴侣蛋白的质粒，并将该质粒进行了转化和小量诱导表达，发现使用这些质粒制备的蛋白也有不同程度的表达，—在后续的实验和生产中用pET30a--OPTI-TF-p30的阳性菌株作为实施例。
1.4.3大量诱导表达
甘油管保藏管菌株活化：取pET30a--OPTI-TF-p30菌株甘油保藏管解冻，用接种环挑取甘油管中菌悬液于卡那抗性平板上划线，37℃培养过夜。挑菌活化：于培养好的平板上挑取单克隆至3mL卡那抗性LB培养基中，37℃220r/min摇床培养5～6h，至OD600达到0.5～0.8。种子液培养：将活化好的菌悬液取450μL接种至450mL卡那抗性LB培养基中，37℃220r/min培养9～10h。发酵培养基配制：按照发酵培养基配方配制发酵培养基组分1于15L发酵罐中，安装组联发酵罐，发酵培养基组分2及补料培养基配制于蓝口瓶中，121℃高压蒸汽灭菌20min。发酵参数设置：Agit 400r/min；Temperature 37℃；pH 7.00；DO 40；Air 100％；Gasflow 2.0。发酵接种：于接种口将450mL发酵培养基组分2，3mL发酵培养基组分3，200μL消泡剂，9mL卡那抗生素(50mg/mL)补加到发酵罐中；将培养好的450mL种子液接种至9L发酵培养基中进行发酵罐放大培养，培养5～6h至OD600值到12～14。降温诱导：设置温度参数，将发酵罐温度降至20℃，取样，加入2.7mL IPTG(1M)至IPTG终浓度为0.3mmol/L，20℃诱导培养8h。发酵补料：待发酵培养至OD600达到17～19时，按5％的速度连续补加补料培养基(先将补料培养基组分1和2混匀)。菌体收集：发酵结束后，收集发酵液在8000r/min离心10min，收集菌体置于-20℃保存。
其中，上述过程中所用培养基如下：
发酵培养基组分1：酵母粉10g/L，胰蛋白胨20g/L，KH2PO4 1.14g/L，K2HPO4 0.9g/L，(NH4)2SO4 3.0g/L，MgSO4·7H2O 0.3g/L，NaCl 5g/L，pH 7.0；发酵培养基组分2：甘油30g/L；发酵培养基组分3：VB1 2mg/L；补料培养基组分1：酵母粉16.67g/L；胰蛋白胨33.33g/L；补料培养基组分2：甘油100g/L。
1.5 p30蛋白纯化
菌体破碎：向菌体中加入裂解液(10ml/g湿重)，搅拌均匀；将菌体样品倒入到细胞均质仪样品槽中，出样口用烧杯准备收集样品；以样品的90％流出出样口为一个循环，一个循环完成后，将出样口烧杯收集的样品倒回样品槽，共进行4个循环。离心：将上步骤中破碎完全的样品分装到250mL Beckman离心管中，12,000rpm，4℃离心30min，上清作为上样样品。镍柱平衡：用超纯水平衡2～3柱体积(CV)，排出20％的乙醇保存液；然后用裂解液平衡2～3柱体积(CV)。上样：取菌体破碎后上清作为样品与镍柱填料混合，于滚瓶机上混匀1h后进行流穿，并收集流穿液。去除内毒素：用50倍柱体积(CV)的清洗缓冲液组分1洗柱，用于去除内毒素。去除TritonX-114：用30倍柱体积(CV)的洗脱缓冲液组分1(elution buffer 1)(不含Triton X-114)洗柱，减少Triton X-114的残留。洗脱：20mM咪唑洗脱缓冲液洗杂，至考马斯亮蓝G250检测无蓝色；500mM咪唑洗脱缓冲液洗脱目的蛋白，至考马斯亮蓝G250检测无蓝色，收集浓缩目的蛋白。
结果：纯化结果如图5所示，1是Marker，2是纯化后的TF-p30蛋白。从图中可以看出，纯化后的TF-p30蛋白SDS-PAGE纯度均能达到80％以上；经过计算，在没有优化发酵条件的情况下，表达量能达到400mg/L以上。
其中蛋白纯化所需溶液如下：
菌液裂解液：组分为50mM NaH2PO4，500mM NaCl，1mMβ-ME，0.5％NP-40，0.05％Tween-20，0.2％TritonX-114，最后调节pH至8.0。清洗缓冲液1：50mM NaH2PO4，1M NaCl，0.05％Tween-20，0.5％NP-40，1mMβ-ME，0.2％TritonX-114，最后调节pH至7.0。洗脱缓冲液组分1：50mM NaH2PO4，0.5％NP-40，500mM NaCl，最后调节pH至8.0。洗脱缓冲液组分2：50mMNaH2PO4，0.5％NP-40，500mM NaCl，500mM咪唑，最后调节pH至8.0。
1.6 TF-P30蛋白稳定性
将实施例5纯化后的蛋白用PBS稀释到600ug/ml，共10ml，分成2份，每份5ml；一份置于-80℃冰箱，一份置于4℃冰箱中，每周取样一次，每次0.5ml，连续取样10次；每次取样后用BCA检测蛋白浓度，结果如下表所示：
从蛋白浓度的变化来看，两组实验过程中蛋白基本保持稳定。为了进一步验证处理后的蛋白的含量是否变化，我们用第十次的样品进行SDS-PAGE检测。具体结果如图6所示：1是蛋白Marker 26619，2，3：第十次样品4℃和-20℃蛋白样品，上样量3μg；从图中可以看出，处理后的样品(第十次取样)仍然具有良好的稳定性。
1.7 TF-P30亚单位疫苗的制备
1.7.1疫苗制备
水相准备：根据疫苗中TF-P30蛋白含量，使用PBS(或生理盐水)将TF-P30蛋白稀释适当浓度，即为水相；
油相准备：根据制备疫苗总量，按照抗原相和佐剂重量比为1:1，体积比为46:54，量取适量ISA 201VG佐剂；
乳化：将水相和油相都预热到33℃，将水相缓缓加到油相中，200-500rpm搅拌20-30min，于20℃静置1h置于4℃过夜；
分装、贮存：根据需要进行分装，检合格后于4℃保存备用。
1.7.2疫苗质检
物理性状采用眼观的方法观察外观(是否是乳白色乳剂)；
采用清洁吸管吸取少量疫苗滴于冷水中，观察(除第1滴外)，疫苗应为云雾状扩散，判定为水包油包水剂型；
将疫苗10ml加入离心管中，以3000r/min离心15min，管底析出的水相应≤0.5mL，判为稳定；
使用粘度仪对疫苗进行粘度检测，应在20-50cp，判为合格。
1.7.3 TF-P30亚单位疫苗对小鼠安全性实验
取健康16-20g SPF雌性小鼠(购于浙江中医药大学)20只，分别随机分4个组，每组5只，按照如下方法进行安全性实验。
单剂量一次免疫组：每组5只按肌肉注射接种100μL(25μg/只)，连续观察2周。
单剂量二次免疫组：每组5只按肌肉注射接种100μL(25μg/只)，连续观察2周。2周后以相同方法剂量再接种一次，继续观察2周。
超剂量一次免疫组：每组5只按肌肉注射接种100μL(200μg/只)，连续观察2周。
对照组：每组5只按肌肉注射接种100μL(PBS配制的疫苗)，连续观察2周。
实验期间，每天观察动物的精神，采食、活动、饮水、注射部位炎症变化和排泄情况等临床变化，记录动物的异常情况。
经过连续观察，对注射TF-P30蛋白的小鼠的临床症状进行比较，单剂量，二次免疫剂量，超剂量免疫组和对照组，均饮食正常，精神无不良变化，排泄正常，注射部位未发现发炎现象，无死亡小鼠出现，接种动物没有遇到任何不良反应。说明，本发明制备的疫苗蛋白即使是以高剂量注射免疫(200μg)，也无明显副作用，是一种安全的免疫蛋白。
1.7.4 ELISA检测
(1)包被：用包被液(50mM碳酸盐缓冲液，pH 9.5)将纯化的TF-P30蛋白稀释至0.5μg/ml，每种抗原均包被8孔(4孔加小鼠血清样品，4孔加封闭液作为对照)，每种抗原均加入100μl/孔，封口膜封好后4℃冰箱放置过夜；
(2)洗涤：从冰箱取出酶标板后，用PBST洗板5次；
(3)封闭：每孔加入200μl封闭液(5％脱脂奶)，封口膜封好后37℃孵育2h；
(4)血清稀释：将用TF-P30蛋白免疫后的小鼠的阳性血清(二免14天后的阳性血清)用封闭液稀释500倍(如499μl稀释液中加入1μl血清，混匀即可)；
(5)洗涤：同(2)；
(6)加样：加入稀释血清，同时用封闭液做阴性对照，37℃孵育1h；
(7)洗涤：同(2)；
(8)加二抗：每孔加入稀释(稀释比为1:5000)的HRP标记的兔抗鼠IgG二抗100μl，37℃孵育0.5h；
(9)洗涤：同(2)；
(10)显色：避光条件下每孔加入100μl的TMB显色液，37℃孵育10min；
(11)终止：每孔加入50μl终止液(2M H2SO4)，终止反应；
(12)检测：于450nm波长测定样品OD值，分析数据；
(13)结果如下表所示：包被TF-P30蛋白的能与血清特异性结合，OD450均值为1.133；包被TF-P30蛋白与封闭液均没有特异性结合，OD450均值为0.049。这说明，TF-P30蛋白可以作为Elisa试剂盒的抗原，在摸索合适的包被浓度和血清稀释比后，可以开发一种检测非洲猪瘟感染和免疫的诊断试剂盒。
本发明通过上面的实施例进行举例说明，但是，应当理解，本发明并不限于这里所描述的特殊实例和实施方案。在这里包含这些特殊实例和实施方案的目的在于帮助本领域中的技术人员实践本发明。任何本领域中的技术人员很容易在不脱离本发明精神和范围的情况下进行进一步的改进和完善，因此本发明只受到本发明权利要求的内容和范围的限制，其意图涵盖所有包括在由附录权利要求所限定的本发明精神和范围内的备选方案和等同方案。
<110>浙江海隆生物科技有限公司
<120>一种p30蛋白的制备方法和应用
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ATGAATCACAAAGTGATGCAAGTTTCAGTTGAAACCACTCAAGGCCTTGGCCGCCGTGTAACGATTACTATCGCTGCTGACAGCATCGAGACCGCTGTTAAAAGCGAGCTGGTCAACGTTGCGAAAAAAGTACGTATTGACGGCTTCCGCAAGGGCAAAGTGCCAATGAATATCGTTGCTCAGCGTTATGGCGCGTCTGTACGCCAGGACGTTCTGGGTGACCTGATGAGCCGTAACTTCATTGACGCCATCATTAAAGAAAAAATCAATCCGGCTGGCGCACCGACTTATGTTCCGGGCGAATACAAGCTGGGTGAAGACTTCACTTACTCTGTAGAGTTTGAAGTTTATCCGGAAGTTGAACTGCAAGGTCTGGAAGCGATCGAAGTTGAAAAACCGATCGTTGAAGTGACCGACGCTGACGTTGACGGCATGCTGGATACTCTGCGTAAACAGCAGGCGACCTGGAAAGAAAAAGACGGCGCTGTTGAAGCAGAAGACCGCGTGACCATCGACTTCACCGGTTCTGTAGACGGCGAAGAGTTCGAAGGCGGTAAAGCGTCTGATTTCGTACTGGCGATGGGCCAGGGTCGTATGATCCCGGGCTTTGAAGACGGTATCAAAGGCCACAAAGCTGGCGAAGAGTTCACCATCGACGTGACCTTCCCGGAAGAATACCACGCAGAAAACCTGAAAGGTAAAGCAGCGAAATTCGCTATCAACCTGAAGAAAGTTGAAGAGCGTGAACTGCCGGAACTGACCGCAGAGTTCATCAAACGTTTCGGCGTTGAAGATGGTTCCGTAGAAGGTCTGCGCGCTGAAGTGCGTAAAAACATGGAGCGCGAGCTGAAGAGCGCCATCCGTAACCGCGTTAAGTCTCAGGCGATCGAAGGTCTGGTAAAAGCTAACGACATCGACGTACCGGCTGCGCTGATCGACAGCGAAATCGACGTTCTGCGTCGCCAGGCTGCACAGCGTTTCGGTGGCAACGAAAAACAAGCTCTGGAACTGCCGCGCGAACTGTTCGAAGAACAGGCTAAACGCCGCGTAGTTGTTGGCCTGCTGCTGGGCGAAGTTATCCGCACCAACGAGCTGAAAGCTGACGAAGAGCGCGTGAAAGGCCTGATCGAAGAGATGGCTTCTGCGTACGAAGATCCGAAAGAAGTTATCGAGTTCTACAGCAAAAACAAAGAACTGATGGACAACATGCGCAATGTTGCTCTGGAAGAACAGGCTGTTGAAGCTGTACTGGCGAAAGCGAAAGTGACTGAAAAAGAAACCACTTTCAACGAGCTGATGAACCAGCAGGCGTCCGCGGGTCTGGAAGTTCTGTTCCAGGGGCCCTCCGCGGGTCTGGTGCCACGCGGTAGTGGTGGTATCGAAGGTAGGCATATGGAGCTCGGTACCCTCGAGGGATCCGAATTCGAGAACCTGTACTTCCAGGGTGACTTCATCCTGAACATTAGCATGAAGATGGAAGTGATCTTTAAAACCGACCTGCGTAGCAGCAGCCAAGTGGTTTTCCACGCGGGTAGCCTGTACAACTGGTTTAGCGTGGAAATCATTAACAGCGGCCGTATCGTTACCACCGCGATTAAGACCCTGCTGAGCACCGTGAAGTATGACATCGTTAAAAGCGCGCGTATTTACGCGGGTCAGGGCTATACCGAGCACCAGGCGCAAGAGGAATGGAACATGATCCTGCACGTTCTGTTCGAGGAAGAGACCGAAAGCAGCGCGAGCAGCGAGAACATTCACGAAAAGAACGATAACGAGACCAACGAATGCACCAGCAGCTTCGAGACCCTGTTTGAGCAAGAACCGAGCAGCGAAGTGCCGAAGGACAGCAAACTGTACATGCTGGCGCAGAAAACCGTTCAACACATCGAGCAGTATGGCAAGGCGCCGGATTTCAACAAAGTGATTCGTGCGCACAACTTTATCCAGACCATTTACGGCACCCCGCTGAAGGAAGAGGAAAAAGAAGTGGTTCGTCTGATGGTTATCAAGCTGCTGAAGAAAATTAGCTTCTACCTGACCTATATCTAA
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<211>688
<212>PRT
<213>TF-P30的蛋白质氨基酸序列
<400>4
MNHKVMQVSVETTQGLGRRVTITIAADSIETAVKSELVNVAKKVRIDGFRKGKVPMNIVAQRYGASVRQDVLGDLMSRNFIDAIIKEKINPAGAPTYVPGEYKLGEDFTYSVEFEVYPEVELQGLEAIEVEKPIVEVTDADVDGMLDTLRKQQATWKEKDGAVEAEDRVTIDFTGSVDGEEFEGGKASDFVLAMGQGRMIPGFEDGIKGHKAGEEFTIDVTFPEEYHAENLKGKAAKFAINLKKVEERELPELTAEFIKRFGVEDGSVEGLRAEVRKNMERELKSAIRNRVKSQAIEGLVKANDIDVPAALIDSEIDVLRRQAAQRFGGNEKQALELPRELFEEQAKRRVVVGLLLGEVIRTNELKADEERVKGLIEEMASAYEDPKEVIEFYSKNKELMDNMRNVALEEQAVEAVLAKAKVTEKETTFNELMNQQASAGLEVLFQGPSAGLVPRGSGGIEGRHMELGTLEGSEFENLYFQGDFILNISMKMEVIFKTDLRSSSQVVFHAGSLYNWFSVEIINSGRIVTTAIKTLLSTVKYDIVKSARIYAGQGYTEHQAQEEWNMILHVLFEEETESSASSENIHEKNDNETNECTSSFETLFEQEPSSEVPKDSKLYMLAQKTVQHIEQYGKAPDFNKVIRAHNFIQTIYGTPLKEEEKEVVRLMVIKLLKKISFYLTYI
序列表
<110> 浙江海隆生物科技有限公司
<120> 一种重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白及其制备方法和应用
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<170> SIPOSequenceListing 1.0
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<211> 606
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<213> 密码子优化前的编码P30蛋白的核苷酸序列(DNA)
<400> 4
<210> 4
<211> 2046
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<213> 编码TF-P30蛋白的核苷酸序列(DNA)
<400> 4
<210> 4
<211> 688
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<213> TF-P30的蛋白质氨基酸序列(PRT)
<400> 4
Met Asn His Lys Val Met Gln Val Ser Val Glu Thr Thr Gln Gly Leu
1 5 10 15
Gly Arg Arg Val Thr Ile Thr Ile Ala Ala Asp Ser Ile Glu Thr Ala
20 25 30
Val Lys Ser Glu Leu Val Asn Val Ala Lys Lys Val Arg Ile Asp Gly
35 40 45
Phe Arg Lys Gly Lys Val Pro Met Asn Ile Val Ala Gln Arg Tyr Gly
50 55 60
Ala Ser Val Arg Gln Asp Val Leu Gly Asp Leu Met Ser Arg Asn Phe
65 70 75 80
Ile Asp Ala Ile Ile Lys Glu Lys Ile Asn Pro Ala Gly Ala Pro Thr
85 90 95
Tyr Val Pro Gly Glu Tyr Lys Leu Gly Glu Asp Phe Thr Tyr Ser Val
100 105 110
Glu Phe Glu Val Tyr Pro Glu Val Glu Leu Gln Gly Leu Glu Ala Ile
115 120 125
Glu Val Glu Lys Pro Ile Val Glu Val Thr Asp Ala Asp Val Asp Gly
130 135 140
Met Leu Asp Thr Leu Arg Lys Gln Gln Ala Thr Trp Lys Glu Lys Asp
145 150 155 160
Gly Ala Val Glu Ala Glu Asp Arg Val Thr Ile Asp Phe Thr Gly Ser
165 170 175
Val Asp Gly Glu Glu Phe Glu Gly Gly Lys Ala Ser Asp Phe Val Leu
180 185 190
Ala Met Gly Gln Gly Arg Met Ile Pro Gly Phe Glu Asp Gly Ile Lys
195 200 205
Gly His Lys Ala Gly Glu Glu Phe Thr Ile Asp Val Thr Phe Pro Glu
210 215 220
Glu Tyr His Ala Glu Asn Leu Lys Gly Lys Ala Ala Lys Phe Ala Ile
225 230 235 240
Asn Leu Lys Lys Val Glu Glu Arg Glu Leu Pro Glu Leu Thr Ala Glu
245 250 255
Phe Ile Lys Arg Phe Gly Val Glu Asp Gly Ser Val Glu Gly Leu Arg
260 265 270
Ala Glu Val Arg Lys Asn Met Glu Arg Glu Leu Lys Ser Ala Ile Arg
275 280 285
Asn Arg Val Lys Ser Gln Ala Ile Glu Gly Leu Val Lys Ala Asn Asp
290 295 300
Ile Asp Val Pro Ala Ala Leu Ile Asp Ser Glu Ile Asp Val Leu Arg
305 310 315 320
Arg Gln Ala Ala Gln Arg Phe Gly Gly Asn Glu Lys Gln Ala Leu Glu
325 330 335
Leu Pro Arg Glu Leu Phe Glu Glu Gln Ala Lys Arg Arg Val Val Val
340 345 350
Gly Leu Leu Leu Gly Glu Val Ile Arg Thr Asn Glu Leu Lys Ala Asp
355 360 365
Glu Glu Arg Val Lys Gly Leu Ile Glu Glu Met Ala Ser Ala Tyr Glu
370 375 380
Asp Pro Lys Glu Val Ile Glu Phe Tyr Ser Lys Asn Lys Glu Leu Met
385 390 395 400
Asp Asn Met Arg Asn Val Ala Leu Glu Glu Gln Ala Val Glu Ala Val
405 410 415
Leu Ala Lys Ala Lys Val Thr Glu Lys Glu Thr Thr Phe Asn Glu Leu
420 425 430
Met Asn Gln Gln Ala Ser Ala Gly Leu Glu Val Leu Phe Gln Gly Pro
435 440 445
Ser Ala Gly Leu Val Pro Arg Gly Ser Gly Gly Ile Glu Gly Arg His
450 455 460
Met Glu Leu Gly Thr Leu Glu Gly Ser Glu Phe Glu Asn Leu Tyr Phe
465 470 475 480
Gln Gly Asp Phe Ile Leu Asn Ile Ser Met Lys Met Glu Val Ile Phe
485 490 495
Lys Thr Asp Leu Arg Ser Ser Ser Gln Val Val Phe His Ala Gly Ser
500 505 510
Leu Tyr Asn Trp Phe Ser Val Glu Ile Ile Asn Ser Gly Arg Ile Val
515 520 525
Thr Thr Ala Ile Lys Thr Leu Leu Ser Thr Val Lys Tyr Asp Ile Val
530 535 540
Lys Ser Ala Arg Ile Tyr Ala Gly Gln Gly Tyr Thr Glu His Gln Ala
545 550 555 560
Gln Glu Glu Trp Asn Met Ile Leu His Val Leu Phe Glu Glu Glu Thr
565 570 575
Glu Ser Ser Ala Ser Ser Glu Asn Ile His Glu Lys Asn Asp Asn Glu
580 585 590
Thr Asn Glu Cys Thr Ser Ser Phe Glu Thr Leu Phe Glu Gln Glu Pro
595 600 605
Ser Ser Glu Val Pro Lys Asp Ser Lys Leu Tyr Met Leu Ala Gln Lys
610 615 620
Thr Val Gln His Ile Glu Gln Tyr Gly Lys Ala Pro Asp Phe Asn Lys
625 630 635 640
Val Ile Arg Ala His Asn Phe Ile Gln Thr Ile Tyr Gly Thr Pro Leu
645 650 655
Lys Glu Glu Glu Lys Glu Val Val Arg Leu Met Val Ile Lys Leu Leu
660 665 670
Lys Lys Ile Ser Phe Tyr Leu Thr Tyr Ile
675 680
Claims (13)
1.一种重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白，其特征在于，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白包括OPTI-p30蛋白和一种伴侣蛋白，其中所述OPTI-p30蛋白的核苷酸序列为p30蛋白经优化后的序列OPTI-p30如SEQ ID NO.1所示。
2.根据权利要求1所述的非洲猪瘟病毒p30亚单位可溶性融合蛋白，其特征在于，所述伴侣蛋白为TF蛋白。
3.根据权利要求2所述的非洲猪瘟病毒p30亚单位可溶性融合蛋白，其特征在于，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白的核苷酸序列OPTI-TF-p30是OPTI-p30蛋白的5’端插入TF蛋白的核苷酸序列，所述OPTI-TF-p30蛋白的核苷酸序列如SEQ ID NO.3所示。
4.根据权利要求3所述的非洲猪瘟病毒p30亚单位可溶性融合蛋白，其特征在于，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白的氨基酸序列如SEQ ID NO.4所示。
5.根据权利要求4所述的非洲猪瘟病毒p30亚单位可溶性融合蛋白，其特征在于，所述非洲猪瘟病毒p30亚单位可溶性融合蛋白在如SEQ ID NO.4所示的氨基酸序列的氨基末端或羧基末端连接有poly-His、FLAG、c-myc、HA和poly-Arg中的一种标签。
6.一种重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白的制备方法，其特征在于，所述制备方法包括以下步骤：
1)将含有OPTI-p30蛋白和一种伴侣蛋白的基因克隆到原核表达载体中，得到含有非洲猪瘟病毒OPTI-p30蛋白和一种伴侣蛋白编码基因的重组质粒；其中，所述OPTI-p30蛋白的核苷酸序列为经优化后的序列OPTI-p30如SEQ ID NO.1所示；
2)再将步骤1)所述重组质粒转化至大肠杆菌菌株的细胞中，得到重组表达的菌株；
3)通过培养、筛选步骤2)中所述重组表达的菌株，得到高度表达的菌株；
4)发酵培养步骤3)中所述高度表达的菌株，纯化后得到重组的非洲猪瘟病毒p30亚单位可溶性融合蛋白。
7.根据权利要求6所述的制备方法，其特征在于，步骤1)中，所述伴侣蛋白为TF蛋白。
8.根据权利要求7所述的制备方法，其特征在于，含有非洲猪瘟病毒OPTI-p30蛋白和TF蛋白的核苷酸序列OPTI-TF-p30是OPTI-p30蛋白的5’端插入TF蛋白的核苷酸序列，所述OPTI-TF-p30蛋白的核苷酸序列如SEQ ID NO.3所示。
9.根据权利要求8所述的制备方法，其特征在于，含有非洲猪瘟病毒OPTI-p30蛋白和TF蛋白的的氨基酸序列如SEQ ID NO.4所示。
10.根据权利要求9所述的制备方法，其特征在于，含有非洲猪瘟病毒OPTI-p30蛋白和TF蛋白在如SEQ ID NO.4所示的氨基酸序列的氨基末端或羧基末端连接有poly-His、FLAG、c-myc、HA和poly-Arg中的一种标签。
11.根据权利要求6所述的制备方法，其特征在于，步骤1)中，所述原核表达载体为pET28a、pET30a、pBAD、pcold、pQE、或pKK。
12.根据权利要求6所述的制备方法，其特征在于，步骤2)中，所述大肠杆菌菌株选自BL21(DE3)、BL21star(DE3)、arcticexpress中的一种菌株。
13.一种如权利要求1～5任一所述的非洲猪瘟病毒p30亚单位可溶性融合蛋白在制备诊断、预防和治疗非洲猪瘟的疫苗中的应用。
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113533722A (zh) * | 2021-07-21 | 2021-10-22 | 百沃特(天津)生物技术有限公司 | 一种用于非洲猪瘟病毒阻断elisa抗体检测的试剂盒及其检测方法、判定方法和用途 |
| CN115074372A (zh) * | 2021-03-10 | 2022-09-20 | 浙江海隆生物科技有限公司 | 一种重组的非洲猪瘟病毒j18l亚单位蛋白及其制备方法和应用 |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101831407A (zh) * | 2010-04-23 | 2010-09-15 | 天津出入境检验检疫局动植物与食品检测中心 | 抗非洲猪瘟病毒单克隆抗体的杂交瘤细胞系及其所分泌的单克隆抗体 |
| CN103992995A (zh) * | 2014-06-05 | 2014-08-20 | 山东大学 | 一种高表达水溶性肝素酶i融合蛋白及其编码基因 |
| RU2015102383A (ru) * | 2015-01-26 | 2016-08-10 | Государственное научное учреждение Всероссийский научно-исследовательский институт ветеринарной вирусологии и микробиологии Российской академии сельскохозяйственных наук, RU | ЭКСПРЕССИРУЮЩИЕ ПЛАЗМИДНЫЕ ВЕКТОРЫ pET32a(+)ASFV/p30e1 И pET32b(+)ASFV/p30e2 ДЛЯ СИНТЕЗА РЕКОМБИНАНТНЫХ БЕЛКОВ, СОСТОЯЩИХ ИЗ ФРАГМЕНТА p30 ВИРУСА АФРИКАНСКОЙ ЧУМЫ СВИНЕЙ, ТИОРЕДОКСИНА И ПОЛИГИСТИДИНОВЫХ УЧАСТКОВ |
| RU2647573C1 (ru) * | 2017-03-14 | 2018-03-16 | Федеральное государственное бюджетное научное учреждение "Федеральный исследовательский центр вирусологии и микробиологии" (ФГБНУ ФИЦВиМ) | ШТАММ БАКТЕРИЙ ESCHERICHIA COLI KRX pET32b/ASFV/p30-ПРОДУЦЕНТ ХИМЕРНОГО РЕКОМБИНАНТНОГО БЕЛКА p30 ВИРУСА АФРИКАНСКОЙ ЧУМЫ СВИНЕЙ |
| CN108441514A (zh) * | 2018-03-28 | 2018-08-24 | 扬州大学 | 一种表达非洲猪瘟病毒cp204l基因重组腺病毒载体、构建方法及重组腺病毒制备方法 |
| CN109254155A (zh) * | 2018-09-25 | 2019-01-22 | 扬州大学 | 一种检测非洲猪瘟病毒抗原胶体金免疫层析试纸及制备方法和应用 |
-
2019
- 2019-02-26 CN CN201910142630.5A patent/CN111607000B/zh active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101831407A (zh) * | 2010-04-23 | 2010-09-15 | 天津出入境检验检疫局动植物与食品检测中心 | 抗非洲猪瘟病毒单克隆抗体的杂交瘤细胞系及其所分泌的单克隆抗体 |
| CN103992995A (zh) * | 2014-06-05 | 2014-08-20 | 山东大学 | 一种高表达水溶性肝素酶i融合蛋白及其编码基因 |
| RU2015102383A (ru) * | 2015-01-26 | 2016-08-10 | Государственное научное учреждение Всероссийский научно-исследовательский институт ветеринарной вирусологии и микробиологии Российской академии сельскохозяйственных наук, RU | ЭКСПРЕССИРУЮЩИЕ ПЛАЗМИДНЫЕ ВЕКТОРЫ pET32a(+)ASFV/p30e1 И pET32b(+)ASFV/p30e2 ДЛЯ СИНТЕЗА РЕКОМБИНАНТНЫХ БЕЛКОВ, СОСТОЯЩИХ ИЗ ФРАГМЕНТА p30 ВИРУСА АФРИКАНСКОЙ ЧУМЫ СВИНЕЙ, ТИОРЕДОКСИНА И ПОЛИГИСТИДИНОВЫХ УЧАСТКОВ |
| RU2647573C1 (ru) * | 2017-03-14 | 2018-03-16 | Федеральное государственное бюджетное научное учреждение "Федеральный исследовательский центр вирусологии и микробиологии" (ФГБНУ ФИЦВиМ) | ШТАММ БАКТЕРИЙ ESCHERICHIA COLI KRX pET32b/ASFV/p30-ПРОДУЦЕНТ ХИМЕРНОГО РЕКОМБИНАНТНОГО БЕЛКА p30 ВИРУСА АФРИКАНСКОЙ ЧУМЫ СВИНЕЙ |
| CN108441514A (zh) * | 2018-03-28 | 2018-08-24 | 扬州大学 | 一种表达非洲猪瘟病毒cp204l基因重组腺病毒载体、构建方法及重组腺病毒制备方法 |
| CN109254155A (zh) * | 2018-09-25 | 2019-01-22 | 扬州大学 | 一种检测非洲猪瘟病毒抗原胶体金免疫层析试纸及制备方法和应用 |
Non-Patent Citations (4)
| Title |
|---|
| ARJUN THAPA等: "Purification of inclusion body−forming peptides and proteins in soluble form by fusion to Escherichia coli thermostable proteins", 《BIOTECHNIQUES》 * |
| NCBI: "ACCESSION NO: AB213654, Expression vector pColdTF DNA, complete sequence", 《GENBANK DATABASE》 * |
| NCBI: "ACCESSION NO: AHN94305，p30 [African swine fever virus]", 《GENBANK DATABASE》 * |
| S. D. KOLLNBERGER等: "Identification of the principal serological immunodeterminants of African swine fever virus by screening a virus cDNA library with antibody", 《JOURNAL OF GENERAL VIROLOGY》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115074372A (zh) * | 2021-03-10 | 2022-09-20 | 浙江海隆生物科技有限公司 | 一种重组的非洲猪瘟病毒j18l亚单位蛋白及其制备方法和应用 |
| CN113533722A (zh) * | 2021-07-21 | 2021-10-22 | 百沃特(天津)生物技术有限公司 | 一种用于非洲猪瘟病毒阻断elisa抗体检测的试剂盒及其检测方法、判定方法和用途 |
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