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逆转录病毒载体介导表达伪狂犬病病毒PK基因MDBK细胞系的建立
引用本文:李祥敏,钱平,金梅林,陈焕春.逆转录病毒载体介导表达伪狂犬病病毒PK基因MDBK细胞系的建立[J].中国生物化学与分子生物学报,2003,19(3):396-400.
作者姓名:李祥敏  钱平  金梅林  陈焕春
作者单位:华中农业大学畜牧兽医学院动物病毒室,武汉,430070
基金项目:国家“8 63”项目 (No.2 0 0 1AA2 13 0 5 1),湖北省自然科学基金资助项目( 2D0 1ABD110 )资助~~
摘    要:细胞凋亡 (apoptosis)是机体在生理条件下受刺激后 ,经过多种途径的信号传导导致细胞产生一系列形态和生物化学方面的改变而引起的细胞自我消亡的过程 ,是程序性细胞死亡 (programmedcelldeath ,PCD)的主要机制 ,它对于机体的发育 ,内外环境的平衡等都具有重要的作用1,2 ] .研究表明 ,许多病毒具有诱导或抑制细胞凋亡的功能 ,但其发生的机理仍然不清 .随着分子生物学研究的深入 ,人们发现病毒的某些基因与细胞凋亡相关 ,其编码的基因产物或因病毒感染诱导细胞表达的蛋白抑制或诱导细胞发生凋亡 .最近研究表明 ,伪狂犬病病毒 (psedorabies…

收稿时间:2003-06-20
修稿时间:2002年9月3日

Construction of MDBK Cells Expressing Pseudorabies Virus PK Gene by Retroviral Expression Vector
LI Xiang min,QIAN Ping,JIN Mei lin,CHEN Huan chun.Construction of MDBK Cells Expressing Pseudorabies Virus PK Gene by Retroviral Expression Vector[J].Chinese Journal of Biochemistry and Molecular Biology,2003,19(3):396-400.
Authors:LI Xiang min  QIAN Ping  JIN Mei lin  CHEN Huan chun
Institution:(Laboratory of Animal Virology, College of Animal Husbandry and Veterinary Science, Huazhong Agricultural University, Wuhan 430070, China
Abstract:Based on the nucleotide sequence of pseudorabies virus (PrV) Ka strain, a pair of primers was designed. PrV Ea strain PK gene was cloned and inserted into retroviral expression vector pLXSN (neo r). The recombinant plasmid pLXSNPK was co transfected with packaging cell PA317 using lipofectin method. The transfectants were selected by G418 (300 mg/L) for 2 weeks. Viral supernatants gaining stable cell clones were collected, the total RNA was extracted and the PK gene was amplified by RT PCR. It was shown that the PK gene had been inserted into the retroviral genome. The positive viral supernatant was collected to infect the interesting target cell MDBK. The infected MDBK cells were selected by G418 (800 mg/L) and analyzed by indirect immunofluorescence and SDS PAGE and Western blotting. It was confirmed that a MDBKPK cells expressing PK protein was obtained. It is useful to study the biological function of PrV PK gene in the process of PrV inducing culture cells apoptosis.
Keywords:pseudorabies virus  PK  gene  retroviral expression vector  MDBKPK cells
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