Gill-associated nidovirus of Penaeus monodon prawns transcribes 3′-coterminal subgenomic mRNAs that do not possess 5′-leader sequences
Gill-associated nidovirus of Penaeus monodon prawns transcribes 3′-coterminal subgenomic mRNAs that do not possess 5′-leader sequences
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DOI：
10.1099/0022-1317-83-4-927
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发表时间：
2002-04-01
影响因子：
3.8
通讯作者：
Walker, PJ
中科院分区：
文献类型：
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作者：
Cowley, JA;Dimmock, CM;Walker, PJ
对斑节对虾鳃相关病毒（GAV）ssRNA基因组中约20 kb的5 '端部分进行序列分析，发现其含有与巢状病毒目冠状病毒和动脉炎病毒相同的ORF 1a-1b复制酶基因。对ORF 1a-1b下游至3 '-poly（A）尾的GAV基因组的剩余6.2 kb相似序列进行序列分析，鉴定出两个高度保守的基因间序列，其中29/32个核苷酸是保守的。使用探针对四个推定的GAV ORF和总RNA或poly（A）选择的RNA进行北方杂交，鉴定了两个3 '-共末端亚基因组（sg）mRNA，其类似于6 kb和类似于5.5 kb。引物延伸和5 '-RACE分析表明，sgmRNA起始于两个保守基因间序列的中心区域中相同的5'-AC位置。这两种方法都没有提供任何证据表明GAV sgmRNA与基因组5 '-前导RNA序列融合，就像脊椎动物冠状病毒和动脉炎病毒的情况一样。还通过RNA酶/DNA酶消化来自GAV感染的对虾组织的总RNA鉴定了大小相当于26.2kb基因组RNA的细胞内双链（ds）RNA和两种sgRNA。仅鉴定出在保守基因间序列中的相同位置处起始的两个sgmRNA以及不存在与这些sgmRNA融合的5 '-基因组前导序列证实了GAV具有很少的基因，并表明其利用可能类似于脊椎动物圆环病毒但不同于冠状病毒和动脉炎病毒的转录机制。
Sequence analysis of the similar to 20 kb 5'-terminal portion of the ssRNA genome of gill-associated virus (GAV) of Penaeus monodon prawns has previously established that it contains an ORF1a-1b replicase gene equivalent to those of the coronavirus and arterivirus members of the order Nidovirales. Sequence analysis of the remaining similar to 6.2 kb of the GAV genome downstream of ORF1a-1b to a 3'-poly(A) tail has identified two highly conserved intergenic sequences in which 29/32 nucleotides are conserved. Northern hybridization using probes to the four putative GAV ORFs and either total or poly(A)-selected RNA identified two 3'-coterminal subgenomic (sg) mRNAs of similar to 6 kb and similar to 5.5 kb. Primer extension and 5'-RACE analyses showed that the sgmRNAs initiate at the same 5'-AC positions in the central region of the two conserved intergenic sequences. Neither method provided any evidence that the GAV sgmRNAs are fused to genomic 5'-leader RNA sequences as is the case with vertebrate coronaviruses and arteriviruses. Intracellular double-stranded (ds)RNAs equivalent in size to the 26,2 kb genomic RNA and two sgRNAs were also identified by RNase/DNase digestion of total RNA from GAV-infected prawn tissue. The identification of only two sgmRNAs that initiate at the same position in conserved intergenic sequences and the absence of 5'-genomic leader sequences fused to these sgmRNAs confirms that GAV has few genes and suggests that it utilizes a transcription mechanism possibly similar to the vertebrate toroviruses but distinct from coronaviruses and arteriviruses.