Pearson M N, Rohrmann G F
Department of Agricultural Chemistry, Oregon State University, Corvallis 97331-7301.
J Virol. 1995 Jan;69(1):213-21. doi: 10.1128/JVI.69.1.213-221.1995.
Homologous regions (hrs) were identified in the Lymantria dispar nuclear polyhedrosis virus (LdMNPV) genome. A 1.58-kb region surrounding hr4 was sequenced and found to have two distinct domains. Domain I (about 600 bp) is composed of seven repeats of about 80 bp including a series of palindromes containing MluI sites and overlapping XhoI and SacI sites. Domain II (about 700 bp) is composed of eight partially repeated sequences of 60 to 100 bp containing a 15- to 25-bp sequence that is 80 to 100% A+T in addition to a 6- to 10-bp palindrome containing an NruI site. Hybridization of a domain I sequence to cosmids containing the LdMNPV genome indicated its presence at eight positions (hr1 to -8) on the genome. In contrast, hybridization of domain II indicated that it was present only at the hr4 locus. A DpnI-based transient-replication assay was used to determine if subclones of hr4 transfected into LdMNPV-infected L. dispar cells functioned as replication origins. Subclones of hr4 containing either domain I or domain II replicated at very low or moderate levels, respectively. However, when domain I and domain II were linked on the same plasmid, high levels of replication were observed. A 1.4-kb region containing hr1 was also sequenced. It lies immediately upstream of the polyhedrin gene and contains six domain I-type repeats. Four-hundred-base-pair regions of domain I repeats from hr1 and hr4 showed 89% sequence identity. Plasmids containing the hr1 domain I replicated at low levels. However, hybrid plasmids in which the AT-rich hr4 domain II was inserted adjacent to hr1 domain I replicated to high levels, indicating that the AT-rich domain II greatly enhances replication. The orientation and position of domains I and II relative to each other did not have major effects on the levels of replication.
在舞毒蛾核型多角体病毒（LdMNPV）基因组中鉴定出同源区域（hrs）。对围绕hr4的一个1.58 kb区域进行了测序，发现其有两个不同的结构域。结构域I（约600 bp）由7个约80 bp的重复序列组成，包括一系列含有MluI位点以及重叠的XhoI和SacI位点的回文序列。结构域II（约700 bp）由8个60至100 bp的部分重复序列组成，其中包含一个15至25 bp的序列，该序列的A + T含量为80%至100%，此外还有一个含有NruI位点的6至10 bp回文序列。结构域I序列与含有LdMNPV基因组的黏粒杂交表明，其在基因组的8个位置（hr1至-8）存在。相比之下，结构域II的杂交表明它仅存在于hr4位点。基于DpnI的瞬时复制试验用于确定转染到受LdMNPV感染的舞毒蛾细胞中的hr4亚克隆是否作为复制起点发挥作用。含有结构域I或结构域II的hr4亚克隆分别以非常低或中等水平进行复制。然而，当结构域I和结构域II连接在同一质粒上时，观察到高水平的复制。还对包含hr1的一个1.4 kb区域进行了测序。它位于多角体蛋白基因的紧邻上游，包含6个结构域I型重复序列。来自hr1和hr4的结构域I重复序列的400 bp区域显示出89%的序列同一性。含有hr1结构域I的质粒以低水平复制。然而，其中富含AT的hr4结构域II插入到hr1结构域I相邻位置的杂交质粒则以高水平复制，这表明富含AT的结构域II极大地增强了复制。结构域I和结构域II相对于彼此的方向和位置对复制水平没有重大影响。
