CN115197918B - 一种病毒保护剂及其应用 - Google Patents
一种病毒保护剂及其应用 Download PDFInfo
- Publication number
- CN115197918B CN115197918B CN202210869669.9A CN202210869669A CN115197918B CN 115197918 B CN115197918 B CN 115197918B CN 202210869669 A CN202210869669 A CN 202210869669A CN 115197918 B CN115197918 B CN 115197918B
- Authority
- CN
- China
- Prior art keywords
- protective agent
- virus
- samples
- suspension
- rotavirus
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N7/00—Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/5306—Improving reaction conditions, e.g. reduction of non-specific binding, promotion of specific binding
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56983—Viruses
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2720/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsRNA viruses
- C12N2720/00011—Details
- C12N2720/12011—Reoviridae
- C12N2720/12311—Rotavirus, e.g. rotavirus A
- C12N2720/12351—Methods of production or purification of viral material
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/005—Assays involving biological materials from specific organisms or of a specific nature from viruses
- G01N2333/08—RNA viruses
- G01N2333/14—Reoviridae, e.g. rotavirus, bluetongue virus, Colorado tick fever virus
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Molecular Biology (AREA)
- Zoology (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Hematology (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Virology (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Urology & Nephrology (AREA)
- General Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Cell Biology (AREA)
- Biophysics (AREA)
- Food Science & Technology (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
本发明属于生物检测技术领域,具体涉及一种病毒保护剂及其应用,具体为在流行病学检测中的应用。该病毒保护剂,包括以下质量百分比的原料:蔗糖1~10%、氯化锌0.01~0.1%、吐温20 0.1~0.3%、D‑山梨醇质量百分比为10%的HBSS溶液8~15%,余量为水。该病毒保护剂能够在常温下保存样品达半年之久,且不影响高通量检测结果,在流行病学监测中具有良好的应用前景。
Description
技术领域
本发明属于生物检测技术领域,具体涉及一种病毒保护剂及其应用,具体为在流行病学检测中的应用。
背景技术
RNA病毒是一种不具细胞结构,只含一种核酸,必须寄生于活细胞内的非细胞生物,容易发生变异,往往有多种不同的基因型。以轮状病毒为例,病毒基因组由11段分节段的dsRNA组成,分别编码6种结构蛋白(VP)和6种非结构蛋白(NSP),有三层蛋白衣壳,最外层由VP4和VP7蛋白组成,是RV主要的中和抗原,根据VP4和VP7核苷酸序列的差异,RV的分型是根据P基因型(VP4,蛋白酶敏感)和G基因型(VP7,糖蛋白)来进行分型。目前已经发现的有36种G基因型,51种P基因型。
目前,世界范围内流行的G-P基因型组合的流行株主要是G1P[8]、G2P[4]、G3P[8]、G4P[8]、G9P[8]和G12P[8]。在我国,因腹泻住院的5岁以下的婴幼儿轮状病毒检测阳性率约为50%,灭活轮状病毒疫苗是一种安全有效地预防儿童因感染轮状病毒所引起疾病的疫苗,疫苗在接种后应该对流行地区的大多数流行株都具有保护作用,因此,对不同地区的RV进行持续的监测十分必要,尤其是引入疫苗之后,也应随时检测流行病学,检测其是否因为疫苗发生血清型的改变与迁移,是否会产生新的血清型。
在持续的监测过程中,经常会涉及到样品的保存,而现有的病毒保护剂保存时间过久时,样品容易发生降解,一般只能保存几个小时或者几天,远远难以满足流行病监测过程中样本的保存时间,因此,开发一种病毒保护时间长的保护剂对于病毒性流行病的监测具有重要意义。
发明内容
针对现有技术所存在的技术问题,本发明的目的之一在于提供一种病毒保护剂,该病毒保护剂能够在常温下保存样品达半年之久,且不影响高通量检测结果。
本发明采用如下技术方案来实现上述技术目的:
本发明提供一种病毒保护剂,包括以下质量百分比的原料:蔗糖1~10%、氯化锌0.01~0.1%、吐温20 0.1~0.3%、D-山梨醇质量百分比为10%的HBSS溶液8~15%,余量为水。
可选的,包括以下重量百分比的原料:蔗糖2~5%、氯化锌0.02~0.05%、吐温200.2~0.3%、D-山梨醇质量百分比为10%的HBSS溶液15%,余量为水。
本发明的目的之二在于保护上述病毒保护剂在流行病学检测中的应用。
本发明的目的之三在于保护上述病毒保护剂在轮状病毒腹泻样本处理中的应用。
其中,所述病毒保护剂添加在粪便悬浮液离心后得到的上清中,病毒保护剂的添加量为1-2mL。
本发明的目的之四在于保护一种轮状病毒腹泻样本病毒血清型鉴定方法,包括以下步骤:步骤一,制备粪便悬浮液,离心,收集上清;步骤二,向上清中加入病毒保护剂;步骤三,采用高通量测序的方法进行测序并分析病毒血清型。
可选的,还包括采用ELISA试剂盒初步鉴定的步骤,所述ELISA试剂盒鉴定时采用的是步骤一中得到的上清,和/或采用一代测序进行鉴定的步骤,所述一代测序采用的是步骤一中得到的上清或者步骤二中得到的添加了病毒保护剂的上清。
在流行病学的检测时,可先采用ELISA试剂盒进行第一步的初筛鉴定,之后再采用一代测序技术进行初步分型检测,对于未分型的样品则进一步采用高通测序技术集合生物信息学方法分析,既避免了一代测序通量低,样品要求高,检出效率低的问题,又避免了完全采用高通量测序费用高的问题。
该方法的准确性已经得到验证确定,将其应用于流行病学监测中,有利于在流行病学监测中发现罕见血清型、罕见血清型组合或新的血清型论病毒,具有良好的应用前景。
可选的,所述粪便悬浮液为采用TNC缓冲液得到的悬浮液。1×TNC(Tris-NaCl-CaCl2)缓冲液配制如下:10mL 1M CaCl2溶液,2.24g Tris(0.02M)和16.38g NaCl(0.28M),使用DEPC水定容至2L,高温灭菌后备用。
可选的,当步骤一中的粪便来自于尿不湿或尿布时,向其上加入TNC缓冲液,静置后震荡,得到混悬液,使用细胞筛网过滤后即得到粪便悬浮液。其中细胞筛网为100um,BeyoGoldTM细胞过滤器100μm孔径,采用独立纸塑包装,无菌。
附图说明
图1、图2为本发明实施例1部分样品VP7蛋白基因电泳结果。
具体实施方式
下面结合具体实施例对本发明作进一步的详细说明,以使本领域的技术人员更加清楚地理解本发明。
以下各实施例,仅用于说明本发明,但不止用来限制本发明的范围。基于本发明中的具体实施例,本领域普通技术人员在没有做出创造性劳动的情况下,所获得的其他所有实施例,都属于本发明的保护范围。
在本发明实施例中,若无特殊说明,所有原料组分均为本领域技术人员熟知的市售产品;在本发明实施例中,若未具体指明,所用的技术手段均为本领域技术人员所熟知的常规手段。
实施例1
本实施例提供一种轮状病毒临床样本的基因检测及分型方法,包括以下步骤:
(1)参与Ⅲ期临床试验的205份腹泻病例的粪样或尿不湿样品的收集及处理
粪样的处理:取适量粪便样品置于离心管中,加入1×TNC缓冲液,制备得到10%(质量/体积)的粪便悬浮液,漩涡震荡,5000rpm离心5min,收集上清至冻存管中;
尿不湿样品的处理:将盛放有尿不湿/尿不湿样品的粪便收集盒从冷冻冰箱中取出,向各个粪便盒中加入1×TNC缓冲液5~10mL,旋紧粪便盒盖后静止放置10min,将粪便盒放置在漩涡震荡仪上震荡1min,使粪便盒内呈混悬液状态(使用无菌镊子或者无菌注射器等对溶胀后的尿不湿样品进行挤压),使用一次性细胞筛网过滤,将粪便盒内的混悬液通过筛网进入到离心管中,5000rpm离心5min,取上清。
(2)通过ProSpecTTM轮状病毒ELISA检测试剂盒检测,收集并整理呈轮状病毒阳性的样品。
(3)样品RNA提取及一代测序
使用病毒RNA提取试剂盒QIAamp Viral RNAMini Kit(购于美国QIAGEN公司,货号:52906)抽提各临床样品的病毒总RNA;使用一步法逆转录PCR试剂盒PrimeScript IIHigh Fidelity One Step RT-PCR Kit(购于Takara公司,货号:R026A)进行一步法RT-PCR;
其中,RT-PCR扩增体系所用引物如下表1所示,RT-PCR反应体系如下表2所示,RT-PCR扩增程序如下表3所示:
表1轮状病毒VP4、VP6、VP7蛋白基因扩增引物
表2 RT-PCR反应体系
名称 | 体积(μl) |
2×One Step High Fidelity Buffer | 25 |
PrimeScript II RT Enzyme Mix | 1 |
PrimeSTAR GXL for 1step RT-PCR | 4 |
上游引物(20μM) | 1 |
下游引物(20μM) | 1 |
样品RNA | 4 |
RNase Free dH2O | 14 |
总计 | 50 |
表3 RT-PCR扩增程序
PCR反应结束后,将产物进行1%琼脂糖凝胶电泳,鉴定PCR扩增条带,以VP7部分条带为例,如图1所示。PCR产物条带大小鉴定正确后,将扩增产物送取生工生物工程(上海)股份有限公司进行测序,测序结果使用Sequencher5.4.5软件拼接双向测序序列,处理将各片段拼接得到的基因组序列并整理,分型结构如下表4所示:
表4 PCR分型结果汇总
VP7 | VP6 | VP4 | |
完成分型数量 | 127/205(62.0%) | 89/205(43.4%) | 83/205(40.5%) |
其中G型未成功分型样品75例,其血清型分型结果如下表5所示:
表5 75例G型未成功分型的样品的血清型分型结果
(4)样品RNA提取及高通量测序
选取其中的20份样品的RNA质检,结果如表6所示。
合成cDNA并制备文库,采用Illumina Hiseq 2500测序仪进行测序,采用PE150模式,高通量测序得到的原始图像数据经碱基识别转化为原始测序序列,称之为Raw Data,结果以FASTQ格式存储;高通量测序得到的Raw Data中会有少量reads包含测序接头、低质量碱基等,使用Fastp对Raw Data进行质控,并使用SortMeRNA和默认的rRNA数据库去除样品中的rRNA序列,将数据质控后得到的Clean Data比对到参考基因组(A组轮状病毒)上,得到拼接后的序列,将高通量测序得到的各样品拼接序列使用NCBI BLAST进行序列比对。统计20份样品分型结果,见下表7所示,同时比较一代测序及高通量测序的分型结果及检出率。
表6A0001-A0020 20份样品RNA浓度
表7 20份样品一代测序及高通量测序的分析结果及检出率
通过表7可以观察到:一代测序和高通量测序均得出分型结果的样品,一代测序及高通量测序结果呈现出一致性;根据检出率及一致性比对,一代测序VP7、VP6、VP4的检出率分别为20%、10%、25%,高通量测序的检出率依次为85%、75%、85%。
实施例2病毒保护剂的选择确定
在实施例1中表6、表7所述的20份经过1×TNC缓冲液处理后得到上清样本分别取1mL并加入1mL不同的病毒保护剂,将其放置不同时间后采用高通量测序的方法进行检测,不同配方的检测结果如下表8所示:
表8不同病毒保护剂的检测结果分析
研究表明,当采用氯化锌0.01-0.1%w/v、蔗糖1-10%w/v、吐温20 0.1-0.3%w/v及含有D-山梨醇的HBSS溶液8-15%w/v配制的溶液作为保护剂使用时,不影响高通量测序的检出率。
实施例3
将实施例1中75例G型未成功分型的样本分别加入实施例2中配方4制备得到的保护剂,提取RNA后直接进行高通量测序,测序结果如表9所示:
表9 75例G型未成功分型的样本采用高通量测序后的分型结果
/>
/>
如表9所示,除少数样品由于病毒含量太低,无法组装序列以外,其余均成功进行序列组装并得出分型结果,说明该方法具有很好的敏感性,可以大大提高轮状病毒临床样本的检测效率。且该方法的检测结果中存在G3P、G4P等罕见轮状病毒G-P血清型组合,因此,使用一代测序和高通量测序技术相结合的分型检测方法对于不同血清型的轮状病毒能够进行扩增及全基因组分型,分型效率高,可避免漏检。该方法快速、高效,有望广泛应用于轮状病毒的流行病学监测,也可用于疫苗临床试验期间的样品分型检测,因疫苗引入后的流行病学迁移与否,新的轮状病毒血清型、血清型组合的监测等。
实施例4
将实施例1中的205例样本分别加入实施例2中配方4制备得到的保护剂,提取RNA后直接进行高通量测序,统计分析分型样本的检出率,结果如表10所示:
表10 RT-PCR与高通量测序检出率的比较
检出率(%) | VP7 | VP6 | VP4 |
PCR方法 | 62.0% | 43.4% | 40.5% |
高通量测序 | 98.0% | 74.6% | 94.6% |
通过分析可知,采用高通量测序可将病毒样本VP7、VP6、VP4的分型了由62.0%、43.4%、40.5%提高至98.0%、74.6%、94.6%,临床样本可以不通过一代测序进行初步分型,直接进行高通量测序分型,分型效果显著。
实施例5病毒保护剂的稳定性研究
将实施例1中选用的A0001-A0020共计20份样品分别分装至两个保存管中,每管加入1mL样品,其中一批加入病毒保护剂另一批不加入病毒保护剂,均常温放置半年,其中病毒保护剂的配方为:0.02%氯化锌、15%D-山梨醇-HBSS、5%蔗糖、0.2%吐温20,滤膜过滤后使用。
将两批样品进行RNA提取,使用QIAamp Viral RNA Mini Kit试剂盒进行RNA提取。
将提取后的RNA样品进行高通量测序,比较检出率结果如表11所示:
表11添加保护剂和不添加保护剂的样品放置半年后的检出率比较
检出率(%) | VP7 | VP6 | VP4 |
未加入保护剂 | 65% | 65% | 50% |
加入保护剂 | 95% | 85% | 90% |
由此可知,本发明所述的病毒保护剂可以延长样品的放置时间达半年之久,高通量测序可以应对任何处理条件下的轮状病毒腹泻样品的分型监测。
在此有必要指出的是,以上实施例仅限于对本发明的技术方案做进一步的阐述和说明,并不是对本发明的技术方案的进一步的限制,本发明的方法仅为较佳的实施方案,并非用于限定本发明的保护范围。凡在本发明的精神和原则之内,所作的任何修改、等同替换、改进等,均应包含在本发明的保护范围之内。
Claims (9)
1.一种病毒保护剂,其特征在于,该病毒保护剂用于轮状病毒腹泻样品的保护,其由以下质量百分比的原料制备而成:
蔗糖 1~10%、氯化锌 0.01~0.1%、吐温20 0.1~0.3%、D-山梨醇质量百分比为10%的HBSS溶液 8~15%,余量为水。
2.根据权利要求1所述的病毒保护剂,其特征在于,由以下重量百分比的原料制备而成:
蔗糖 2~5%、氯化锌 0.02~0.05%、吐温20 0.2~0.3%、D-山梨醇质量百分比为10%的HBSS溶液 15%,余量为水。
3.权利要求1~2任一项所述的病毒保护剂在轮状病毒腹泻样本处理中的应用。
4.根据权利要求3所述的病毒保护剂在轮状病毒腹泻样本处理中的应用,其特征在于,所述病毒保护剂添加在粪便悬浮液离心后得到的上清中。
5.根据权利要求4所述的病毒保护剂在轮状病毒腹泻样本处理中的应用,其特征在于,病毒保护剂的添加量为1-2mL。
6.一种轮状病毒腹泻样本病毒血清型鉴定方法,其特征在于,所述方法用于不以疾病的诊断和治疗为目的的轮状病毒流行病学血清型的监测,包括以下步骤:
步骤一,制备粪便悬浮液,离心,收集上清;
步骤二,向上清中加入权利要求1或2所述的病毒保护剂;
步骤三,采用高通量测序的方法进行测序并分析病毒血清型。
7.根据权利要求6所述的轮状病毒腹泻样本病毒血清型鉴定方法,其特征在于,还包括采用ELISA试剂盒初步鉴定的步骤,所述ELISA试剂盒鉴定时采用的是步骤一中得到的上清,
和/或
采用一代测序进行鉴定的步骤,所述一代测序采用的是步骤一中得到的上清或者步骤二中得到的添加了病毒保护剂的上清。
8.根据权利要求6或7所述的轮状病毒腹泻样本病毒血清型鉴定方法,其特征在于,所述粪便悬浮液为采用TNC缓冲液得到的悬浮液。
9.根据权利要求8所述的轮状病毒腹泻样本病毒血清型鉴定方法,其特征在于,当步骤一中的粪便来自于尿不湿或尿布时,向其加入TNC缓冲液,静置后震荡,得到混悬液,使用细胞筛网过滤后即得粪便悬浮液。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202210869669.9A CN115197918B (zh) | 2022-07-22 | 2022-07-22 | 一种病毒保护剂及其应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202210869669.9A CN115197918B (zh) | 2022-07-22 | 2022-07-22 | 一种病毒保护剂及其应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN115197918A CN115197918A (zh) | 2022-10-18 |
CN115197918B true CN115197918B (zh) | 2023-07-18 |
Family
ID=83584027
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202210869669.9A Active CN115197918B (zh) | 2022-07-22 | 2022-07-22 | 一种病毒保护剂及其应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN115197918B (zh) |
Family Cites Families (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
AU2015291181A1 (en) * | 2014-07-18 | 2017-02-02 | Msd Wellcome Trust Hilleman Laboratories Pvt. Ltd. | A thermostable freeze dried rotavirus vaccine formulation and process to prepare thereof |
CN109055321A (zh) * | 2018-08-27 | 2018-12-21 | 武汉博沃生物科技有限公司 | 一种轮状病毒的培养方法 |
-
2022
- 2022-07-22 CN CN202210869669.9A patent/CN115197918B/zh active Active
Also Published As
Publication number | Publication date |
---|---|
CN115197918A (zh) | 2022-10-18 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Zhang et al. | Virome comparisons in wild-diseased and healthy captive giant pandas | |
Oka et al. | Cell culture isolation and sequence analysis of genetically diverse US porcine epidemic diarrhea virus strains including a novel strain with a large deletion in the spike gene | |
Rahman et al. | Characterization of a novel P [25], G11 human group a rotavirus | |
Gao et al. | Clades of Adeno-associated viruses are widely disseminated in human tissues | |
Randhawa et al. | DNA sequencing of viral capsid protein VP-1 region in patients with BK virus interstitial nephritis | |
Vogtlin et al. | Quantification of feline herpesvirus 1 DNA in ocular fluid samples of clinically diseased cats by real-time TaqMan PCR | |
Fujii et al. | Spread and predominance in Japan of novel G1P [8] double-reassortant rotavirus strains possessing a DS-1-like genotype constellation typical of G2P [4] strains | |
Koopmans et al. | Optimization of extraction and PCR amplification of RNA extracts from paraffin-embedded tissue in different fixatives | |
Decaro et al. | Molecular characterisation of the virulent canine coronavirus CB/05 strain | |
Amer et al. | Isolation and molecular characterization of type I and type II feline coronavirus in Malaysia | |
Pongsuwanna et al. | Serological and genomic characterization of porcine rotaviruses in Thailand: detection of a G10 porcine rotavirus | |
Ataseven et al. | Detection and sequence analysis of equine gammaherpesviruses from horses with respiratory tract disease in Turkey | |
CN111321248B (zh) | 非洲猪瘟病毒mgf-505r基因荧光pcr检测试剂、试剂盒及其应用 | |
Li et al. | Isolation and identification of a recombinant porcine epidemic diarrhea virus with a novel insertion in S1 domain | |
Zobba et al. | Molecular survey of parvovirus, astrovirus, coronavirus, and calicivirus in symptomatic dogs | |
Rubio-Guerri et al. | Novel adenovirus detected in captive bottlenose dolphins (Tursiops truncatus) suffering from self-limiting gastroenteritis | |
Ma et al. | The Spodoptera frugiperda Sf9 cell line is a heterogeneous population of rhabdovirus-infected and virus-negative cells: Isolation and characterization of cell clones containing rhabdovirus X-gene variants and virus-negative cell clones | |
Opriessnig et al. | Porcine astrovirus type 5-associated enteritis in pigs | |
Hoa-Tran et al. | Detection of three independently-generated DS-1-like G9P [8] reassortant rotavirus A strains during the G9P [8] dominance in Vietnam, 2016–2018 | |
Tomaszewski et al. | Detection and heterogeneity of herpesviruses causing Pacheco's disease in parrots | |
Fu et al. | Molecular characterization of a novel wheat-infecting virus of the family Betaflexiviridae | |
Thilakarathne et al. | Development and application of a combined molecular and tissue culture-based approach to detect latent infectious laryngotracheitis virus (ILTV) in chickens | |
Parthiban et al. | Molecular detection of porcine parvovirus 1–associated reproductive failure in southern India | |
Xue et al. | Isolation of feline panleukopenia virus from Yanji of China and molecular epidemiology from 2021 to 2022 | |
Cong et al. | A multiplex xTAG assay for the simultaneous detection of five chicken immunosuppressive viruses |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
CB02 | Change of applicant information |
Address after: 100000 Floor 2, Building 2, Yard B, Shuangqiao Road, Chaoyang District, Beijing Applicant after: China Biotechnology Co.,Ltd. Applicant after: WUHAN INSTITUTE OF BIOLOGICAL PRODUCTS Co.,Ltd. Address before: 100000 2nd Floor, Building 2, Yard B2, Shuangqiao Road, Chaoyang District, Beijing Applicant before: China Biotechnology Co.,Ltd. Applicant before: WUHAN INSTITUTE OF BIOLOGICAL PRODUCTS Co.,Ltd. |
|
CB02 | Change of applicant information | ||
GR01 | Patent grant | ||
GR01 | Patent grant |