Complete nucleotide sequence and genomic organization of Periplaneta fuliginosa densonucleosis virus

We have cloned the replicative form of thePeriplaneta fuliginosa densonucleosis virus (PfDNV) genome and determined its complete sequence. The sequence has 5 454 nucleotides (nt), the genome consists of an internal unique sequence flanked by inverted terminal repeats (201 nt). The first 122 nt at the 5′ end and the terminal 122 nt at the 3′ end of both plus and minus strands can fold into a typical hairpin structure. The genome contains seven major open reading frames (ORFs). The plus strand has 4 ORFs occupying the 5′ half of the plus strand, whereas the others span the 5′ half of the minus strand. Two potential promoters were found at map units (m.u.) 3 and 97. Computer analysis of sequence homologies with other parvoviruses suggests that the plus strand ofPf DNV encodes very likely the nonstructural proteins and the minus strand probably encodes the structural proteins.     

Molecular cloning and nucleotide sequence of the attenuated hog cholera virus Lapinized Chinese strain

The genomic sequence of the attenuated hog cholera virus Lapinized Chinese strain (HCLV) was determined from overlapping cDNA clones. The viral RNA of HCLV stain comprised 12 310 nucleotide (nt) including 374 nt and 239 nt at the 5′ and 3′-noncoding region, respectively. The complete genome sequence contained one large open reading frame which encoded an amino acid sequence of 3 898 residues with a calculated molecular weight of 437×10³. Although there were mostly only small differences between the sequence of the HCLV strain and the published sequences of strains ALD, GPE⁻, Alfort and Brescia, there was one notable insertion of 12 nucleotides, TTTTCTTTTTTC in the 3′ non-coding region of HCLV strain. Supported by the National Pandeng Project, Genbank accession number AF091507 Wang Jiafu: born in 1972, Ph. D.     

Genome sequence variation analysis of two SARS coronavirus isolates after passage in Vero cell culture

SARS coronavirus is an RNA virus whose replication is error-prone, which provides possibility for escape of host defenses, and even leads to evolution of new viral strains during the passage or the transmission. Lots of variations have been detected among different SARS-CoV strains. And a study on these variations is helpful for development of efficient vaccine. Moreover, the test of nucleic acid characterization and genetic stability of SARS-CoV is important in the research of inactivated vaccine. The whole genome sequences of two SARS coronavirus strains after passage in Vero cell culture were determined and were compared with those of early passages, respectively. Results showed that both SAPS coronavirus strains have high genetic stability, although nearly 10 generations were passed. Four nucleotide variations were observed between the second passage and the llth passage of Sinol strain for identification of SARS inactivated vaccine. Moreover, only one nucleotide was different between the third passage and the 10th passage of Sino3 strain for SARS inactivated vaccine. Therefore, this study suggested it was possible to develop inactivated vaccine against SARS-CoV in the future.     

A complete sequence and comparative analysis of a SARS-associated virus （Isolate BJO1）

The genome sequence of the Severe Acute Respiratory Syndrome (SARS)-assoclated virus provides essential information for the identification of pathogen(s), exploration of etiology and evolution, interpretation of transmission and pathogenesis, development of diagnostics, prevention by future vaccination, and treatment by developing new drugs.We report the complete genome sequence and comparative analysis of an isolate (B J01) of the coronavirus that has been recognized as a pathogen for SARS. The genome is 29725 nt in size and has 11 ORFs (Open Reading Frames). It is composed of a stable region encoding an RNA-dependent RNA polymerase (composed of 20RFs) and a variable region representing 4 CDSs (coding sequences) for viral structural genes (the S, E, M, N proteins) and 5 PUPs (putative uncharacterized proteins). Its gene order is identical to that of other known coronaviruses. The sequence alignment with all known RNA viruses places this virus as a member in the family of Coronaviridae. Thirty putative substitutions have been identified by comparative analysis of the 5 SARS-associated virus genome sequences in GenBank. Fifteen of them lead to possible amino acid changes (non-synonymous mutations) in the proteins. Three amino acid changes, with predicted alteration of physical and chemical features, have been detected in the S protein that is postulated to be involved in the immunoreactions between the virus and its host.Two amino acid changes have been detected in the M protein,which could be related to viral envelope formation. Phylogenetic analysis suggests the possibility of non-human origin of the SARS-associated viruses but provides no evidence that they are man-made. Further efforts should focus on identifying the etiology of the SARS-associated virus and ruling out conclusively the existence of other possible SARS-related pathogen(s).     

A complete sequence and comparative analysis of a SARS-associated virus(Isolate BJ01)

Severe Acute Respiratory Syndrome (SARS) is a newly identified infectious disease[1—5]. The global outbreak of SARS has been threatening the health of people worldwide and has killed 353 people and infected more than 5462 in 27 countries, as reported by WHO on April 29, 2003 (http://www.who.int/csr/sarscountry/en). Although it has been recognized that a variant of virus from the family of coronavirus might be the candidate pathogen of SARS[1—5], its identity as the unique pathogen sti…     

IB-ELISA诊断试剂的研究

目的　为鸡群日常监测传染性支气管炎病毒 (IBV)抗体提供一个方便、特异、快速的检测方法。方法　利用 9日龄SPF鸡胚尿囊腔接种培养IBV ,以及鸡胚成纤维细胞传代培养IBV ,经纯化后 ,测定蛋白含量 ,作为包被抗原 ,利用过碘酸盐法标记兔抗鸡IgG作为酶结合物 ,用TMB作为显色剂 ,建立起酶联免疫吸附试验 (ELISA)技术 ,检测血清样品中传染性支气管炎抗体。结果　上述两种方法制备的抗原皆可用于此试验 ,与NDV、MDV、IBDV、AIV、APIV、REV、ILTV、IC、MG、MS等抗血清无交叉反应 ,与IDEXXIB ELISA比较 ,结果吻合较好。结论　本研究制备的IB ELISA诊断试剂 ,可用于日常监测传染性支气管炎病毒抗体     

SARS相关冠状病毒刺突糖蛋白的研究(综述)

概述了SARS—CoV的S蛋白的结构及功能相关研究。严重急性呼吸综合症相关的冠状病毒(SARS-CoV)引起2003年我国南方非典型肺炎爆发流行，波及多个国家和地区。目前全球许多学对SARS-CoV进行了广泛的研究，发现S蛋白是病毒表面的主要蛋白，它构成冠状病毒科特征性的冠状样结构，在严重急性呼吸综合症的发病机制起着关键性作用，可介导表达相关受体的宿主细胞感染。现已鉴定出SARS-CoV的S蛋白相关受体，同时它在抗病毒感染中是一个关键靶蛋白。     

SARS冠状病毒及相关病毒的分子系统学分析

根据SARS冠状病毒及其相关病毒的基因组核酸序列和3种不同蛋白质序列，应用最大简约法和最小进化法重建系统发育树；并对SARS冠状病毒的11个推测蛋白质(ORF)做BLAST分析。结果表明，SARS冠状病毒和鼠肝炎病毒——牛冠状病毒分支构成姊妹群。其单系群性质得到强有力的统计学支持。这暗示了SARS的爆发可能源自种间屏障的突破事件，该病毒天然宿主可能为猪、牛或鼠。SARS冠状病毒与已知的人冠状病毒分属冠状病毒科的不同分支，因此致病机制可能有很大不同。3个基因的系统树分支格局的一致性表明：SARS冠状病毒这3个主要基因与其他冠状病毒间不存在重组，但全部11个ORF的BLAST分析却认为其基因组上一些小的区段可能与其他病毒存在重组。     

Cloning and expression of the vp39 gene ofBombyx mori nuclear polyhedrosis virus inE. coli

The nuclear capsid protein gene (vp39) ofBombyx mori nuclear polyhedrosis virus (BmNPV) was amplified successfully by PCR technique and inserted into pGEM 3zf(+). The 5′ and 3′ terminal area of the amplified vp39 gene were sequenced with silver-staining dideoxy method. Bmvp39 gene was sub-cloned into the expression vector pRSET-A, and transformed intoE. coli BL21. This gene was highly expressed by IPTG induction. SDS-PAGE analysis showed that the expressed protein is about 38 kd, and the expressed amount reached maxium in 4 h with IPTG induction. Supported by the National Natural science Foundation of China and the Doctoral Foundation of Edn carto Committee Liu Deli: born in 1954, Doctoral Candidate from Huazhong Normal University To whom correspondence should be addressed: (027-7882712-2938)     

Cloning and expression of the vp39 gene of Bombyx mori nuclear polyhedrosis virus in E. coli

The nuclear capsid protein gene (vp39) ofBombyx mori nuclear polyhedrosis virus (BmNPV) was amplified successfully by PCR technique and inserted into pGEM 3zf(+). The 5′ and 3′ terminal area of the amplified vp39 gene were sequenced with silver-staining dideoxy method. Bmvp39 gene was sub-cloned into the expression vector pRSET-A, and transformed intoE. coli BL21. This gene was highly expressed by IPTG induction. SDS-PAGE analysis showed that the expressed protein is about 38 kd, and the expressed amount reached maxium in 4 h with IPTG induction. Supported by the National Natural science Foundation of China and the Doctoral Foundation of Edn carto Committee Liu Deli: born in 1954, Doctoral Candidate from Huazhong Normal University To whom correspondence should be addressed: (027-7882712-2938)     

Southern rice black-streaked dwarf virus: A new proposed <Emphasis Type="Italic">Fijivirus</Emphasis> species in the family <Emphasis Type="Italic">Reoviridae</Emphasis>

For the past several years, a novel dwarf disease has been observed on rice （Oryza sativa） in some regions of Guangdong Province and Hainan Province, southern China. Infected plants showed stunting, dark leaf and small enations on stem and leaf back. Typical Fijivirus viroplasma containing crystalline arrayed spherical virons approximately 70--75 nm in diameter and tubular structures were detected in ultrathin sections by an electron microscope in parenchyma phloem cells of the infected plants. The virus was transmitted to rice seedlings by white-backed planthoppers, Sogatella furcifera （Hemiptera： Delphacidae）, collected in the diseased fields. Analysis of dsRNA extracts from infected plants revealed ten linear segments, which were similar to the electrophoretic profile of Rice black-streaked dwarf virus （RBSDV）. RT-PCR with a single primer which matched to a linker sequence ligated to both 3＇ ends of the viral genomic dsRNAs resulted in amplification of genome segments 9 （S9） and 10 （S10） cDNA products. The complete nucleotide sequences of S9 and S10 were obtained from clones of the RT-PCR amplicon exhibited characteristic properties of Fijivirus including low GC content （34.5% and 35.6%）, genus conserved 5＇ and 3＇ termini sequences and similar genome organization. Blast searches indicated that the sequences of S9 and S10 shared 68.8%--74.9% and 67.1% --77.4% nucleotide identities with those of viruses in the Fijivirus group 2, respectively. These values were similar to those among other viruses in the Fijivirus group 2 and considerably lower than those among RBSDV isolates. Phylogenetic trees based on S9 and S10 nucleotide sequences and their putative amino acid sequences showed that this virus represented a separate branch among other Fijiviruses. The virus was also detected by a nested RT-PCR assay in corn （Zea mays）, barnyard grass （Echinochloa crusgalll）, Juncellus serotinus and flaccidgrass （Pennisetum flaccidum） in and/or adjacent to the infected rice fields. I     

SARS冠状病毒核衣壳蛋白cDNA的克隆和原核表达载体的构建

目的克隆SARS冠状病毒核衣壳(N)蛋白.方法用RT-PCR技术克隆SARS冠状病毒N蛋白全长cDNA,cDNA经序列分析鉴定后,克隆到pET30a表达载体.结果实现了SARS冠状病毒N蛋白基因的克隆.结论可以对重组成功的pET30a-N进行进一步的应用.     

Cloning and characterization of vernalization-related gene (ver203F) cDNA 3′ end

Based on the cDNA fragment sequence of vernalization-related geneverc203 cloned by differential screening in our lab, the 5′ primer has been designed. The cDNA 3′ end ofver203 gene (1 197 bp) has been cloned by the RACE method. And it is identified by Northern blotting that its expression is special in vernalization treatment. After comparing the sequence in the nucleotide sequence databases of Genbank, EMBL and DDBJ, the gene has homology withHordeum vulgare jesmonate-induced protein gene. It is suggested that this gene might be related to the signal transduction mediated by jamonate.     

Bioinformatic Analysis of Putative Gene Products Encoded in SARS—HCoV Genome

The cause of severe acute respiratory syndrome (SARS) has been identified as a new coronavirus named as SARS-HCoV. Using bioinformatic methods, we have performed a detailed domain search. In addition to the viral structure proteins, we have found that several putative polypeptides share sequence similarity to known domains or proteins. This study may provide a basis for future studies on the infection and replication process of this notorious virus.     

Study on synthesis and biological activities of novel water-soluble metalloporphyrins

The synthesis and characterization of a series of metal porphyrins, M^II(Por), {Por=dianionic of 5, 10, 15, 20-tetrakis [4-(4′-bromobutyloxy) phenyl]porphyrinato and 5, 10, 15, 20-tetrakis [4-(4′-butyloxypyridine bromide)phenyl]porphyrinato, M=Zn, Cu, Mn, Co, Ni, Ru-CO} were described. The complexes 3a-3e were prepared from the reactions of compound 2 with metal acetates in chloroform, and the treatment of 3a-3f with excess of pyridine gave corresponding complexes 4a-4f. These new compounds were identified by absorption spectroscopies,¹H-NMR and elemental analyses. The results of biological activity testing for 4a-4f revealed that 4a and 4c had stronger inhibiting action on the growth metabolism ofEsche richia coli. Foundation item: Supported by the National Natural Science Foundation of China (No: 29972035) and the Zi-Qiang Science Foundation of Wuhan University, 2000 Biography: Li Zao-ying (1949-), famale, Professor, research interests, study on synthesis and biological activities of metalloprphrins.     

Molecular cloning and nucleotide sequence of classical swine fever virus strain Shimen

According to the previously published CSFV sequences, 18 paris of partially overlapping primers which span the entire genome of CSFV strain Shimen were designed and synthesized. Each cDNA fragment of strain Shimen was amplified by RT-PCR method from the anticoagulant blood of strain Shimen infected pig. The PCR fragments were cloned into pGEM-T vector respectively and sequenced. The results show that we have obtained the nucleotide sequence of strain Shimen. The viral RNA consists of 12 297 nucleotides including noncoding regions of 373 and 227 bases at the 5′ and 3′ end, respectively, and a single large open reading frame spanning 11 697 nucleotides in the middle, which encodes an amino acid sequence of 3 989 residues with a calculated molecular weight of 437.6×10³. The precisely sequencing of 5′ and 3′ termini is undertaking. Supported by the National Pandeng Project Huang Qianhua: born in 1968. Graduate student     

引起人非典型性肺炎的冠状病毒基因组与其它冠状病毒基因组的初步比较

一种在世界范围内突然爆发的致命流行病——急性呼吸窘迫综合症(SARS)击倒了数干人，一种全新的冠状病毒被认为是其病原．5个SARS相关冠状病毒的全基因组序列已经完成．我们进行了SARS相关冠状病毒和其它冠状病毒的基因组进化分析和序列比较，结果显示：1．SARS相关冠状病毒不直接来自于任何已知的冠状病毒；2．E蛋白的基因可能是在近期从其它病毒横向转移到SARS病毒中来的；3．Sl和S2基因发生了较大范围的缺失或插入突变．这些基因横向转移和突变改变了SARS相关病毒的表面结构和抗原性，极有可能是导致其获得侵染人类细胞的主要原因．     

国内鸡传染性支气管炎病毒的分离与鉴定

对国内五个省患呼吸系统和肾脏系统疾病的肉鸡和蛋鸡19份样品进行了IBV分离和鉴定,结果表明RNZ与M41同型.HV、NB-90、TJ株与美国G株和澳大利亚T株血清型相同,证明国内存在不同的IBV血清型株.