PRV闽A株BamHI片段克隆及其第7片段的鉴定

Identification of Bam HI-7 Fragment and Cloning of Bam HI Restriction Enzyme Fragments of Pseudorabies VirusMin-A Strain DNA

用鸟枪法将ＰＲＶ闽Ａ株的ＢａｍＨＩ酶切片断克隆到ｐＢＲ３２２质粒中，再经抗性筛选、酶切鉴定和菌落原位杂交，证实已克隆了ＰＲＶ闽Ａ株１４个ＢａｍＨＩ酶切片段中的１２个，从而构建了其基因文库。通过Ｓｏｕｔｈｅｒｎ转印杂交和酶切图谱分析鉴定了重组质粒ｐＰＲ１２８，其插入片段包含了ＰＲＶ闽Ａ株糖蛋白ｇｐ５０基因在内的ＢａｍＨＩ－７片段，核酸长约６．８ｋｂ。

Pseudoraties virtis(PRV)Min-A strain isolated first in China was prop-agated in MDBK cells and its DNA was purified.The Bam HI restriction enzyme frag-ments of PRV Min-A strain DNA digested completely by Bam HI were cloned into theplasmid pBR322.The Bam HI restriction enzyme fragments except Bam HI-1,2 werecloned by resistance screening, restriction enzyme analysis and in situ hybridization.The genomic library of PRV Min-A strain DNAs was constructed.The recombinantplasmid pPR128 inserted with Bam HI-7 fragment that was approximately 6.8 kb andcontained complete gp50 gene was identified by restriction enzyme analysis and Southernblot hvbridization.