Wang Yong Jie, Yao Qin, Chen Ke Ping, Wang Yong, Lu Jian, Han Xu
Institute of Life Sciences, Jiangsu University, Zhenjiang, 212013, P. R. China.
Virus Genes. 2007 Aug;35(1):103-8. doi: 10.1007/s11262-006-0034-3. Epub 2006 Oct 18.
The genome of Bombyx mori densovirus (China isolate), termed as BmDNV-3, is composed of two kinds of different single-stranded linear DNA molecules (VD1 and VD2). In this study, the viral DNA molecules were purified and cloned into pUC119 vector, and the complete nucleotide sequence was determined. Sequence analysis showed that VD1 genome consisted of 6,543 nts including inverted terminal repeats (ITRs) of 224 nts, and VD2 genome consisted of 6,022 nts including ITRs of 524 nts. Comparison of the complete genome sequence between BmDNV-3 and BmDNV-2 (Yamanashi isolate) showed an identity of 98.4% in VD1 and 97.7% in VD2, with a total number of 228 bp substitutions, 11 bp deletions and 3 bp insertions found in BmDNV-3. A single nucleotide "A" deletion at nt 1589 in BmDNV-3 caused a frame shift mutation and brought about a premature stop codon, thus dividing VD2 of BmDNV-3 into two ORFs (named VD2 ORF1a and VD2 ORF1b) within that region, while there was only one ORF (named VD2 ORF1) in the corresponding region of BmDNV-2 (Yamanashi isolate). Comparative polymorphisms of ORFs and ITR regions of the two viral genomes showed that highly variable regions were mainly located in VD1 ORF3, VD1 ORF4, VD2 ORF2, and ITRs of BmDNV-3. Northern blots analysis revealed that VD1 had 1.1 kb and 1.5 kb transcripts from the left half of its plus strand, and one transcript about 3.3 kb from the right half of its minus strand. Sequencing of 3' and 5' RACE products showed that the 1.1 kb transcript started at nt 290 and ended at nt 1437, the 1.5 kb transcript started at nt 1423 and ended at nt 2931, and the 3.3 kb transcript started at nt 6287 and ended at nt 2922. These results help us to further understand the variation between different DNV genera and its possible causes, providing clues for studying the evolutionary history of densoviruses.
家蚕浓核病毒（中国分离株），命名为BmDNV - 3，其基因组由两种不同的单链线性DNA分子（VD1和VD2）组成。在本研究中，对病毒DNA分子进行了纯化，并克隆到pUC119载体中，测定了其完整的核苷酸序列。序列分析表明，VD1基因组由6543个核苷酸组成，包括224个核苷酸的反向末端重复序列（ITRs），VD2基因组由6022个核苷酸组成，包括524个核苷酸的ITRs。BmDNV - 3与BmDNV - 2（山梨分离株）的完整基因组序列比较显示，VD1的同一性为98.4%，VD2的同一性为97.7%，在BmDNV - 3中总共发现了228个碱基对的替换、11个碱基的缺失和3个碱基的插入。BmDNV - 3中第1589位核苷酸处的单个核苷酸“A”缺失导致了移码突变，并产生了一个提前终止密码子，从而将BmDNV - 3的VD2在该区域内分为两个开放阅读框（命名为VD2 ORF1a和VD2 ORF1b），而在BmDNV - 2（山梨分离株）的相应区域只有一个开放阅读框（命名为VD2 ORF1）。两种病毒基因组的开放阅读框和ITR区域的比较多态性表明，高变区主要位于BmDNV - 3的VD1 ORF3、VD1 ORF4、VD2 ORF2和ITRs中。Northern印迹分析显示，VD1在其正链左半部分有1.1 kb和1.5 kb的转录本，在其负链右半部分有一个约3.3 kb的转录本。3'和5' RACE产物的测序表明，1.1 kb的转录本从第290位核苷酸开始，到第1437位核苷酸结束，1.5 kb的转录本从第1423位核苷酸开始，到第2931位核苷酸结束，3.3 kb的转录本从第6287位核苷酸开始，到第2922位核苷酸结束。这些结果有助于我们进一步了解不同浓核病毒属之间的差异及其可能的原因，为研究浓核病毒的进化历史提供线索。
