DDRT-PCR法研究小鼠耐受全氟异丁烯中毒的相关基因

Identification of the related gene of mouse tolerant to exposure to perfluoroisobutylene by DDRT-PCR

目的 急性低剂量全氟异丁烯 (PFIB)染毒后的活存小鼠可对致死量PFIB产生耐受性 ,本文旨在寻找与耐受有关的基因对揭示PFIB中毒耐受机制。方法 采用静式全暴露法 3次染毒PFIB ,抽提耐受 1个致死量PFIB活存动物的肺组织tRNA ,采用DDRT- PCR方法 ,使用 3个锚定引物和 4个随机引物分别对耐受小鼠和对照小鼠肺组织总tRNA进行逆转录和PCR扩增反应 ,通过测序胶电泳和银染分析其基因表达差异 ,并对差异DNA片段进行克隆、测序。结果 耐受PFIB的小鼠与对照鼠相比 ,在基因表达上发生了明显变化。对耐受小鼠特有的 3个差异DNA片段进行克隆、测序 ,通过BLAST与Genbank中的基因比对显示 ,克隆的EST(expressedsequencetags)均与小鼠特定的基因高度同源。结论 特异基因的表达是小鼠耐受PFIB的基础 ,耐受动物特有的 3种差异表达EST对应的基因对于揭示PFIB耐受机理可能具有重要意义。

Objective\ The mice surviving acute exposure to low do se of perfluorois obutylene (PFIB) could tolerant to lethal dose of PFIB. Discovering the gene rel ated with the tolerance would be significant to find preventive agents for PFIB. Methods By static exposure to PFIB for 3 times,the survival ani mals could tolera te to lethal dose of PFIB.Three anchoring primers and 4 arbitary primers were ap plied in differential display reverse transcription PCR(DDRT-PCR) of lung tissu e s and the products were analyzed by sequencing PAGE and silver staining.The diff erential DNA fragments were cloned and sequenced.Results There were obvious diff erences of gene expresion between control and tolerant animals.Three expressed s equence tags(EST) have been cloned and sequenced,which were homologous to some m ouse gene in Genbank. Conclusion Specific gene up-regulation m inght be the cause of tolerance to PFIB, and they are very important to clarity the tolerant mecha nisms to PFIB.