Gorziglia M, Cashdollar L W, Hudson G R, Esparza J
J Gen Virol. 1983 Dec;64 ( Pt 12):2585-95. doi: 10.1099/0022-1317-64-12-2585.
The genes of a field isolate of human rotavirus were cloned into pBR322. The strategy used involved polyadenylation of denatured double-stranded (ds) genomic RNA, followed by cDNA synthesis using reverse transcriptase. Oligo(dC) tails were added to the 3' end of the single-stranded cDNA and then separated by alkaline agarose electrophoresis. Sized cDNA of both polarities were allowed to hybridize and inserted into the PstI site of pBR322. Transformations done with sized cDNA always resulted in the selection of hybrid plasmids carrying inserts with a size representative of the original cDNA. Four individual clones were selected for preliminary characterization. One clone has an insert of 1360 base pairs (bp) and corresponds to gene 6. The second clone has an insert of 1140 bp and corresponds to one of the genes in the triplet 7-8-9. The other two genes, with inserts of 780 and 660 bp, were identified as corresponding to dsRNA segments 10 and 11.
一株人轮状病毒野毒株的基因被克隆到pBR322中。所采用的策略包括对变性双链（ds）基因组RNA进行聚腺苷酸化，然后使用逆转录酶合成cDNA。寡聚（dC）尾被添加到单链cDNA的3'末端，然后通过碱性琼脂糖电泳进行分离。将两种极性的大小合适的cDNA进行杂交，并插入到pBR322的PstI位点。用大小合适的cDNA进行转化总是能筛选出携带具有原始cDNA大小代表性插入片段的杂交质粒。选择了四个单独的克隆进行初步鉴定。一个克隆有一个1360个碱基对（bp）的插入片段，对应于基因6。第二个克隆有一个1140 bp的插入片段，对应于三联体7 - 8 - 9中的一个基因。另外两个基因，插入片段分别为780和660 bp，被鉴定为对应于dsRNA片段10和11。
