Bai J, Howes K, Payne L N, Skinner M A
Department of Molecular Biology, Institute for Animal Health, Compton, Nr Newbury, Berks, UK.
J Gen Virol. 1995 Jan;76 ( Pt 1):181-7. doi: 10.1099/0022-1317-76-1-181.
A genomic DNA library was constructed, in a bacteriophage lambda vector, from line 0 chick embryo fibroblasts (CEFs) infected with HPRS-103, an exogenous avian leukosis virus (ALV; envelope subgroup J) recently isolated from meat-type chickens. The library was screened at high stringency using a full length RAV-1 (subgroup A) proviral probe. From 10(6) plaques, two clones which hybridized strongly to the RAV-1 probe were isolated; one contained a full-length copy of the proviral genome of HPRS-103 and the other contained a copy lacking the 5'-long terminal repeat (LTR) and part of gag. The relative strength of hybridization of RAV-1 and HPRS-103 clones, to RAV-1 probes representing different parts of the proviral genome, indicated that the gag and pol genes of HPRS-103 share a high level of identity with those of RAV-1 but that the env gene and the LTRs are considerably less well conserved. Infectious virus was recovered from CEFs transfected with the full-length clone, as detected by ELISA. The recovered virus appeared to be identical to HPRS-103 by electron microscopy and by Southern blotting of proviral DNA. The recovered virus was shown to be of the same subgroup as HPRS-103 by serum neutralization and receptor interference assays. Sequence analysis of the env gene of HPRS-103 shows that it differs considerably from the env genes of other ALV subgroups, particularly in the host range determinants, consistent with the finding that HPRS-103 represents a new subgroup (designated J).
利用噬菌体λ载体,构建了来自感染HPRS - 103的0代鸡胚成纤维细胞(CEF)的基因组DNA文库。HPRS - 103是一种最近从肉用型鸡中分离出的外源性禽白血病病毒(ALV;囊膜J亚群)。使用全长RAV - 1(A亚群)前病毒探针在高严谨度下筛选该文库。从10^6个噬菌斑中,分离出两个与RAV - 1探针强烈杂交的克隆;一个包含HPRS - 103前病毒基因组的全长拷贝,另一个包含一个缺少5'-长末端重复序列(LTR)和部分gag基因的拷贝。RAV - 1和HPRS - 103克隆与代表前病毒基因组不同部分的RAV - 1探针的杂交相对强度表明,HPRS - 103的gag和pol基因与RAV - 1的gag和pol基因具有高度同源性,但env基因和LTRs的保守性要差得多。通过ELISA检测,从转染了全长克隆的CEF中回收了感染性病毒。通过电子显微镜和前病毒DNA的Southern印迹分析,回收的病毒似乎与HPRS - 103相同。通过血清中和试验和受体干扰试验表明,回收的病毒与HPRS - 103属于同一亚群。HPRS - 103的env基因序列分析表明,它与其他ALV亚群的env基因有很大差异,特别是在宿主范围决定簇方面,这与HPRS - 103代表一个新亚群(命名为J)的发现一致。