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1.
分别利用5’RACE和3’RACE确定了CMVSDRNA2的5’和3’末端序列,在此基础上,利用RTPCR得到了RNA2的5’端一半的cDNA克隆pC25和3’端一半的cDNA克隆pC23,并通过拼接构建了RNA2全长cDNA克隆pC2F。通过对pC25和pC23进行序列测定,得到了RNA2的全序列。序列分析结果表明CMVSDRNA2由3048nt组成,其中存在2个部分重叠的阅读框ORF1(79~2652nt)和ORF2(2414~2746nt),分别编码858aa的2a蛋白和111aa的2b蛋白,并在2a蛋白的序列中发现了动植物病毒复制酶所特有的两个保守序列。该株系RNA2核苷酸序列与分属CMVI亚组的Fny株系和II亚组的Q株系RNA2的核苷酸序列同源性分别为917%和756%;2a蛋白的氨基酸序列同源性分别为938%和677%,2b蛋白的氨基酸序列同源性分别为830%和513%。同源性比较的结果表明SD株系属于CMVI亚组。 相似文献
2.
从猪水泡病病毒(SVDV)细胞培养物的PEG浓缩毒中提取病毒RNA,经RT-PCR和套式PCR扩增病毒主要保护性抗原蛋白基因,将扩增产物1.6kb插入pUC18载体中,经亚克隆后用双脱氧链终止法测定其序列,与已发表的SVDV分离物该区序列作比较,核苷酸同源性为96%-97%,氨基酸同源性为98%,参与构成SVDV中和性抗原位点的几个氨基酸残基均很保守;与已发表的柯萨奇B5病毒的对应序列比较,两者核苷酸序列同源性为77%,而推导的氨基酸顺序同源性竞高达92%。本文结果有助于SVDV的分子流行病学研究,并为其和柯萨奇B5病毒的相互关系提供参考数据,为SVDV新型疫苗研究提供了基础材料 相似文献
3.
从水稻矮缩病毒(Ricedwarfvirus,RDV)中国福建分离物中克隆分离了最外层外壳蛋白基因(S2)全长cDNA,并对其进行了序列分析,结果表明RDVS2cDNA全长3512bp,仅含一个3348bp的阅读框架,编码一个含有1116个氨基酸的蛋白(P2)。与基因库中已知基因序列比较,发现它与日本RDVH株系相应片段的核苷酸和氨基酸同源率分别为946%和954%,与轮状病毒VP2氨基酸序列有一定的同源性。S2核苷酸序列二级结构预测结果表明,5’端50个核苷酸的二级结构为一个发夹结构和一个茎环结构。P2有4个富含亮氨酸的区域,位于N端亲水区域的10个氨基酸(AA69~78)残基形成一个α螺旋,这些特点均与轮状病毒VP2的结构特征相似。SDSPAGE和Western印迹分析表明在大肠杆菌中分段高效表达了S2编码蛋白的N端和C端。 相似文献
4.
以甜菜坏死黄脉病毒(BNYVV)内蒙分离物的总RNA为模板,通过反转录-PCR扩增获得BNYVVRNA3全长cDNA。将其克隆到pGEM-7Zf(+)上,得到重组质粒pGBY56。序列分析结果表明,内蒙分离物RNA3基因组全长为1775nt,其中包含3个开放阅读框架,分别编码25kD蛋白、4.6kD蛋白和一种由59个氨基酸组成的N蛋白。与法国F2分离物、德国G1分离物和日本S分离物相比,其核苷酸序列的同源性分别为96.4%、96.8%和97.3%。将25kD蛋白编码基因克隆到pJW2上,构建了该基因的原核表达载体。SDS-PAGE和Westernbloting分析结果表明,25kD蛋白基因在E.coliBL21(DE3)中经温度(42℃)诱导后,可特异地表达25kD蛋白 相似文献
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6.
黄瓜花叶病毒亚组Ⅰ和Ⅱ分离物外壳蛋白基因的序列分析与比较 总被引:13,自引:0,他引:13
对我国分离的经生物学和血清学鉴定为黄瓜花叶病毒(CMV)亚组Ⅰ和Ⅱ的各一分离物(GB、XB)的外壳蛋白(CP)基因进行了序列分析和比较。以提纯病毒RNA为模板,进行逆转录及PCR扩增,并通过常规基因克隆方法得到插入CP基因片段的重组克隆。对插入GB和XB两个分离物CP基因片段的重组克隆进行全序列测定,结果表明重组克隆序列长分别为777bp和792bp,均只含一个开放读框(ORF),长度为657nt,可编码218个氨基酸;两个分离物的CP基因核苷酸序列同源率为775%,氨基酸序列同源率为826%。与我国已报道的7个CMV分离物的CP基因序列比较,分离物GB的同源率为913%~973%,分离物XB的同源率为766%~784%。与国际上已报道部分CMV株系的CP基因序列相比较,分离物GB与亚组Ⅰ株系有更密切的亲缘关系,而分离物XB则与亚组Ⅱ株系的亲缘关系更密切。 相似文献
7.
对同一地区两例输血后丙型肝炎进行PCR分型,结果为Ⅱ型。分别将其NS5区基因部分片段克隆到pUC18和M13mp18中,序列分析结果表明在NS5区基因相同区段,二者的核苷酸同源性为89.04%,氨基酸同源性为90.75%。而且在分离株V中第70~72位发现阅读框内终止密码子TGA。与Ⅱ型代表株HCVBK序列相比,相同区段的核苷酸和氨基酸同源性分别为:91.5%,91.92%和91.91%,91.91%。对比分析表明,在同一地区基因型相同的不同分离株间NS5区基因仍存在较大变异,分离株V可能为缺陷性病毒 相似文献
8.
采用HGVNS5特异的2对引物,对两个香港株和一个广东株HGVRNA进行逆转录套式PCR扩增,PCR产物克隆入pUC19,重组质粒转化DH5α和JM109菌株。PCR和酶切法鉴定阳性克隆,双脱氧链末端终止法测定核苷酸序列并进行同源性分析。结果发现核苷酸变异呈散在分布,三株间核苷酸和氨基酸序列同源性分别为93.3%~94%及97%~99.2%,与已报道的中国株(CN)相比,则同源性分别为90%~91.2%和94%~96.3%,与美国株(PNF2161及R10291)相比,为87.1%~89.5%和95.2%~97%,而与西非株(GBVC)相比,则达91.4%~93.8%和97%~97.9%。提示HGVNS5区核苷酸和氨基酸序列相对保守,不同HGV株存在一定的地区差异。 相似文献
9.
马铃薯卷叶病毒基因间隔区的克隆及序列分析 总被引:6,自引:0,他引:6
根据已报道的马铃薯卷叶病毒基因组序列.设计合成一对特异性引物,以马铃薯卷叶病毒中国分离株(PLRV-Ch)的RNA为模板,反转录合成cDNA第一条链,经PCR扩增后克隆于pUC19质粒中,进一步用PCR鉴定、限制酶切分析和序列分析,结果表明:PLRV-Ch基因间隔区由197个核苷酸组成,与国外报道的荷兰PLRV-N加拿大PLRV-C,澳大利亚PLRV-A,苏格兰PLRV-S各株系核苷酸序列具有很高的同源性,同源率依次为99%、98%、93%、98%。 相似文献
10.
以甜菜坏死黄脉病毒(BNYVV)内蒙分离物(NM)RNA为模板,通过反转录和PCR扩增得到了BNYVV RNA4基因组的cDNA克隆pGBF6。序列分析结果表明,pGBF6含有全长RNA4 cDNA插入片段,大小为1465个核苷酸,含有一个849个核苷酸的开放阅读框架,编码产生由282个氨基酸组成的分子量为31kDa的蛋白。与法国F2分离物RNA4相比,其核苷酸序列和由此推导的氨基酸序列同源性分别 相似文献
11.
Nucleotide Sequence Analysis of DNA 总被引:7,自引:0,他引:7
RAY WU 《Nature: New biology》1972,236(68):198-200
There are three major obstacles to the analysis of the nucleotide sequence in a DNA molecule starting from a known location in the DNA molecule. First, it is difficult to obtain large quantities of homogeneous DNA. Second, even the smallest DNA molecules contain several thousand nucleotides which make sequence analysis prohibitive. Third, there are no highly base-specific DNAases available for degrading DNA for sequence analysis. We have overcome some of these obstacles; first, by incorporating highly labelled deoxynucleotides into DNA in vitro, small amounts of material can be used for sequence analysis. Second, the nucleotide sequence of DNA molecules can now be determined from the 5′-terminal. Thus, two dodecanucleotide sequences corresponding to the two cohesive ends of λ DNA have been determined1 and a nona-decanucleotide sequence corresponding to one cohesive end of phage 186 DNA has been completed2. So far, our approach is limited to starting the analysis from the 5′-ends of a DNA molecule. A more general approach is being developed for starting the analysis from other selected parts of a DNA molecule with the use of specifically designed primers. 相似文献
12.
Guenter Stoesser Wendy Baker Alexandra van den Broek Evelyn Camon Maria Garcia-Pastor Carola Kanz Tamara Kulikova Rasko Leinonen Quan Lin Vincent Lombard Rodrigo Lopez Nicole Redaschi Peter Stoehr Mary Ann Tuli Katerina Tzouvara Robert Vaughan 《Nucleic acids research》2002,30(1):21-26
The EMBL Nucleotide Sequence Database (aka EMBL-Bank; http://www.ebi.ac.uk/embl/) incorporates, organises and distributes nucleotide sequences from all available public sources. EMBL-Bank is located and maintained at the European Bioinformatics Institute (EBI) near Cambridge, UK. In an international collaboration with DDBJ (Japan) and GenBank (USA), data are exchanged amongst the collaborating databases on a daily basis. Major contributors to the EMBL database are individual scientists and genome project groups. Webin is the preferred web-based submission system for individual submitters, whilst automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO). Database releases are produced quarterly. Network services allow free access to the most up-to-date data collection via FTP, email and World Wide Web interfaces. EBI’s Sequence Retrieval System (SRS), a network browser for databanks in molecular biology, integrates and links the main nucleotide and protein databases plus many other specialized databases. For sequence similarity searching, a variety of tools (e.g. Blitz, Fasta, BLAST) are available which allow external users to compare their own sequences against the latest data in the EMBL Nucleotide Sequence Database and SWISS-PROT. All resources can be accessed via the EBI home page at http://www.ebi.ac.uk. 相似文献
13.
Nucleotide Sequence of Canine Smad3 总被引:1,自引:0,他引:1
Kazutoshi Sugiyama Ryo Ooishi Yoshii Nishino Masayuki Funaba Masaru Murakami 《Biochemical genetics》2010,48(3-4):202-207
The whole genome sequence of a Boxer dog suggested that the amino acid sequence of the carboxyl terminus of a putative Smad3 is SSVF-COOH, not SSVS-COOH as in all Smad3 sequences identified in many species. Because phosphorylation of the last two serines at the carboxyl terminus is generally indispensable for Smad3-mediated signaling, the role of Smad3 may be unique in dogs. The present study determines the nucleotide sequence of the coding region of canine Smad3 and deduces the carboxyl terminal amino acids of Smad3 in several breeds. Except for the Boxer, the deduced amino acid sequence was SSVS-COOH in all dogs examined. In addition, the nucleotide at position 1204 in the Boxer was different from that of the other dogs. Furthermore, there was a SNP at nt 240. The present study indicates that the carboxyl terminal amino acid of canine Smad3 is not unique, although it is unknown in the Boxer breed. 相似文献
14.
The EMBL Nucleotide Sequence Database (http://www.ebi.ac.uk/embl.html) constitutes Europe's primary nucleotide sequence resource. Main sources for DNA and RNA sequences are direct submissions from individual researchers, genome sequencing projects and patent applications. While automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO), the preferred submission tool for individual submitters is Webin (WWW). Through all stages, dataflow is monitored by EBI biologists communicating with the sequencing groups. In collaboration with DDBJ and GenBank the database is produced, maintained and distributed at the European Bioinformatics Institute (EBI). Database releases are produced quarterly and are distributed on CD-ROM. Network services allow access to the most up-to-date data collection via Internet and World Wide Web interface. EBI's Sequence Retrieval System (SRS) is a Network Browser for Databanks in Molecular Biology, integrating and linking the main nucleotide and protein databases, plus many specialised databases. For sequence similarity searching a variety of tools (e.g. Blitz, Fasta, Blast etc) are available for external users to compare their own sequences against the most currently available data in the EMBL Nucleotide Sequence Database and SWISS-PROT. 相似文献
15.
The EMBL Nucleotide Sequence Database is a comprehensive database of DNA and RNA sequences directly submitted from researchers and genome sequencing groups and collected from the scientific literature and patent applications. In collaboration with DDBJ and GenBank the database is produced, maintained and distributed at the European Bioinformatics Institute (EBI) and constitutes Europe's primary nucleotide sequence resource. Database releases are produced quarterly and are distributed on CD-ROM. EBI's network services allow access to the most up-to-date data collection via Internet and World Wide Web interface, providing database searching and sequence similarity facilities plus access to a large number of additional databases. 相似文献
16.
Homogenization of Tandemly Repeated Nucleotide Sequences by Distance-Dependent Nucleotide Sequence Conversion 总被引:2,自引:1,他引:1
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Previous work revealed that recurrent mutations (= mutation occurring more than once) in the tandemly repeated arrays present in nontranscribed spacers (NTS) of ribosomal RNA genes (rDNA) are clustered, i.e., they most frequently occur in repeats with adjacent or alternate distribution. A possible explanation is that the likelihood of heteroduplex formation, a prerequisite of gene conversion, decreases with the distance between repeats. To test this possibility, evolution of an array of 11 initially homogeneous repeats was computer simulated using three models, two assuming that the likelihood of heteroduplex formation decreases with increasing distance between the repeats and one assuming that it is constant. Patterns of mutation distribution obtained in computer simulations were compared with the distribution of mutations found in the repeated arrays in the NTS of seven rDNA clones. The patterns of mutations generated by the models assuming that the likelihood of heteroduplex formation decreases as distance between the repeats increases agreed with the patterns observed in rDNA; the patterns generated by the model assuming that the likelihood is independent of distance between repeats disagreed with the patterns observed in the rDNA clones. The topology of the heteroduplex formed between DNA in adjacent repeats predicts that the most frequently occurring conversions in the NTS repeated arrays will be shorter than the length of the repeat. The topology of the heteroduplex also predicts that if the heteroduplex leads to crossing over a circular repeat is excised. It is speculated that the circle can transpose or can be amplified via rolling circle replication and subsequently transpose. It is also shown that homogenization of the NTS repeated arrays proceeds at different rates in different species. 相似文献
17.
Jack Lenz Robert Crowther Anthony Straceski William Haseltine 《Journal of virology》1982,42(2):519-529
The sequence of 2,191 nucleotides encoding the env gene of murine retrovirus Akv was determined by using a molecular clone of the Akv provirus. Deduction of the encoded amino acid sequence showed that a single open reading frame encodes a 638-amino acid precursor to gp70 and p15E. In addition, there is a typical leader sequence preceding the amino terminus of gp70. The locations of potential glycosylation sites and other structural features indicate that the entire gp70 molecule and most of p15E are located on the outer side of the membrane. Internal cleavage of the env precursor to generate gp70 and p15E occurs immediately adjacent to several basic amino acids at the carboxyl terminus of gp70. This cleavage generates a region of 42 uncharged, relatively hydrophobic amino acids at the amino terminus of p15E, which is located in a position analogous to the hydrophobic membrane fusion sequence of influenza virus hemagglutinin. The mature polypeptides are predicted to associate with the membrane via a region of 30 uncharged, mostly hydrophobic amino acids located near the carboxyl terminus of p15E. Distal to this membrane association region is a sequence of 35 amino acids at the carboxyl terminus of the env precursor, which is predicted to be located on the inner side of the membrane. By analogy to Moloney murine leukemia virus, a proteolytic cleavage in this region removes the terminal 19 amino acids, thus generating the carboxyl terminus of p15E. This leaves 15 amino acids at the carboxyl terminus of p15E on the inner side of the membrane in a position to interact with virion cores during budding. The precise location and order of the large RNase T(1)-resistant oligonucleotides in the env region were determined and compared with those from several leukemogenic viruses of AKR origin. This permitted a determination of how the differences in the leukemogenic viruses affect the primary structure of the env gene products. 相似文献
18.
本试验测定了已克隆的貂肠炎病毒(MEV)复制型(RF)DNA的核苷酸序列,确定MEV基因组全长约为5064个核苷酸(nucleotides,nt),推测了3'端和5'端结构,在5'端非编码区有3个51 nt的重复。MEV基因组序列与犬细小病毒(CPV)、猫细小病毒(FPV)有很高的同源性,结构基因区的同源性分别达99.1%和99.9%,但在5'端非编码区有较大差异。MEV基因组结构与CPV和FPV基本一致,有两个大的开放阅读框架,分别编码688和722个氨基酸。在map unit(m.u.)3.7和m.u.39处有两个启动子,在m.u.97处有poly A位点。NS2、VP1和VP2的mRNA都发生剪接。 相似文献
19.
如何向基因库(GenBank)发送核酸序列数据 总被引:2,自引:0,他引:2
基因库(GenBank)是美国国家生物技术信息中心(NationalCenterforBiotechnologyInformationNCBI)开发的一个核酸序列数据库,其网址为:htp://wwwncbinlmnihgov/。它是公开获取的... 相似文献
20.
The Nucleotide Sequence of Human Acylamino Acid-Releasing Enzyme 总被引:3,自引:0,他引:3
Mitta Masanori; Ohnogi Hiroshi; Mizutani Shigetoshi; Sakiyama Fumio; Kato Ikunoshin; Tsunasawa Susumu 《DNA research》1996,3(1):31-35
The nucleotide sequence of a cDNA coding for the human acylaminoacid-releasing enzyme (AARE, also known as acylpeptide hydrolase)[EC 3.4.19.1] subunit has been determined. The amino acid sequenceof human AARE subunit deduced from its cDNA nucleotide sequenceshowed a high degree of identity (91.5%) with both the correspondingproteins from the pig and the rat. The AARE cDNA shows 99.2%identity with a 3.3 kb cDNA transcribed from a locus (DNF15S2)on the short arm of human chromosome 3, whose deletion is associatedwith small cell lung cancer, taking into consideration thatthe sequence of the 3.3-kb cDNA previously reported was causedby misreading. 相似文献