Self-excision of the BAC sequences from the recombinant Marek's disease virus genome increases replication and pathogenicity.
Self-excision of the BAC sequences from the recombinant Marek's disease virus genome increases replication and pathogenicity.
复制标题
DOI：
10.1186/1743-422x-5-19
复制
发表时间：
2008-01-30
期刊：
影响因子：
4.8
通讯作者：
Nair V
中科院分区：
文献类型：
--
作者：
Zhao Y;Petherbridge L;Smith LP;Baigent S;Nair V
疱疹病毒全长基因组克隆为细菌人工染色体（BAC），极大地促进了对几种疱疹病毒基因组的操作以鉴定致病决定簇。我们以前曾报道过高致癌性马立克氏病病毒（MDV）株RB-1B的BAC克隆（pRB-1B 5）的构建，这已被证明是一个有价值的资源，阐明几个致癌决定因素。尽管BAC复制子保留在基因组中，但重组病毒能够在易感鸡中诱导肿瘤。然而，尚不清楚BAC的存在是否会影响重组病毒的全部致癌潜力。为了使BAC衍生病毒与亲本RB-1B株的接近性最大化，我们通过恢复Us 2并通过在mini-F质粒的loxP位点内引入SV 40-cre盒来修饰现有的pRB-1B 5克隆，以允许在鸡细胞中自切除质粒序列。重组病毒在体外和体内的复制都有明显的提高。BAC序列的切除也将致病性增强到与亲本病毒相似的水平，因为重组病毒和亲本病毒感染组中马立克氏病的累积发病率没有显示出显著差异。因此，我们已经能够对这种高致癌性病毒的现有BAC克隆进行显著改进，这肯定会增加其作为鉴定这种主要禽类病原体的致癌决定因素的研究的有价值工具的有用性。
Cloning of full length genomes of herpesviruses as bacterial artificial chromosomes (BAC) has greatly facilitated the manipulation of the genomes of several herpesviruses to identify the pathogenic determinants. We have previously reported the construction of the BAC clone (pRB-1B5) of the highly oncogenic Marek's disease virus (MDV) strain RB-1B, which has proven to be a valuable resource for elucidating several oncogenic determinants. Despite the retention of the BAC replicon within the genome, the reconstituted virus was able to induce tumours in susceptible chickens. Nevertheless, it was unclear whether the presence of the BAC influenced the full oncogenic potential of the reconstituted virus. To maximize the closeness of BAC-derived virus to the parental RB-1B strain, we modified the existing pRB-1B5 clone by restoring the Us2 and by introducing SV40-cre cassette within the loxP sites of the mini-F plasmid, to allow self-excision of the plasmid sequences in chicken cells. The reconstituted virus from the modified clone showed significant improvement in replication in vitro and in vivo. Excision of the BAC sequences also enhanced the pathogenicity to levels similar to that of the parental virus, as the cumulative incidence of Marek's disease in groups infected with the recombinant and the parental viruses showed no significant differences. Thus, we have been able to make significant improvements to the existing BAC clone of this highly oncogenic virus which would certainly increase its usefulness as a valuable tool for studies on identifying the oncogenic determinants of this major avian pathogen.