Phylogenetic analysis of a highly conserved region of the polymerase gene from 11 coronaviruses and development of a consensus polymerase chain reaction assay.
Phylogenetic analysis of a highly conserved region of the polymerase gene from 11 coronaviruses and development of a consensus polymerase chain reaction assay.
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DOI：
10.1016/s0168-1702(99)00017-9
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发表时间：
1999-04
期刊：
影响因子：
5
通讯作者：
Gangopadhyay NN
中科院分区：
文献类型：
--
作者：
Stephensen CB;Casebolt DB;Gangopadhyay NN
根据抗原性交叉反应和结构蛋白基因序列分析，目前冠状病毒属病毒分为三组。用聚合酶链式反应(PCR)方法扩增出8株冠状病毒聚合酶基因的开放阅读框(ORF)1b中高度保守的922个核苷酸片段，并进行了序列分析。将这些序列与已发表的另外三种冠状病毒的序列进行了比较。在这一比较中，发现当比较传统冠状病毒组中的病毒时，核苷酸替换频率(每100个核苷酸)变化在46.40到50.13之间，而在这些组中比较时，除了一个例外(人类冠状病毒(HCV)229E)，变化在2.54到15.89之间。(与同一群体的其他成员相比，229E的替换频率从35.37到35.72不等。)对这些Poll基因序列的系统发育分析得到了与前面描述的分组密切对应的分组，包括将两种禽类冠状病毒--鸡传染性支气管炎病毒(IBV)和火鸡冠状病毒(TCV)--归入同一组的最新数据[Guy，J.S.，Barnes，H.J.，Smith L.G.，Breslin，J.，1997。鸟类Dis.41：583-590]。在相同的反应条件下，从所有三组冠状病毒中鉴定出一对简并引物，扩增出251bp的区域。这种针对冠状病毒属的共识聚合酶链式反应分析可能有助于识别未知的冠状病毒。
Viruses in the genus Coronavirus are currently placed in three groups based on antigenic cross-reactivity and sequence analysis of structural protein genes. Consensus polymerase chain reaction (PCR) primers were used to obtain cDNA, then cloned and sequenced a highly conserved 922 nucleotide region in open reading frame (ORF) 1b of the polymerase (pol) gene from eight coronaviruses. These sequences were compared with published sequences for three additional coronaviruses. In this comparison, it was found that nucleotide substitution frequencies (per 100 nucleotides) varied from 46.40 to 50.13 when viruses were compared among the traditional coronavirus groups and, with one exception (the human coronavirus (HCV) 229E), varied from 2.54 to 15.89 when compared within these groups. (The substitution frequency for 229E, as compared to other members of the same group, varied from 35.37 to 35.72.) Phylogenetic analysis of these pol gene sequences resulted in groupings which correspond closely with the previously described groupings, including recent data which places the two avian coronaviruses—infectious bronchitis virus (IBV) of chickens and turkey coronavirus (TCV)—in the same group [Guy, J.S., Barnes, H.J., Smith L.G., Breslin, J., 1997. Avian Dis. 41:583–590]. A single pair of degenerate primers was identified which amplify a 251 bp region from coronaviruses of all three groups using the same reaction conditions. This consensus PCR assay for the genus Coronavirus may be useful in identifying as yet unknown coronaviruses.
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