氡染毒小鼠骨髓细胞差异表达cDNA文库的构建和初步鉴定  

Construction and identification of subtracted cDNA library in bone marrow cells of radon-exposed mice

作　　者：李建祥[1] 傅春玲[2] 陈锐[3] 聂继华[1] 周建伟[4] 童建[1] 

机构地区：[1]苏州大学放射医学与公共卫生学院卫生毒理学教研室,215123 [2]苏州大学放射医学与公共卫生学院营养与食品教研室,215123 [3]苏州大学附属第二医院呼吸内科 [4]南京医科大学

出　　处：《中华放射医学与防护杂志》2008年第1期21-23,共3页Chinese Journal of Radiological Medicine and Protection

基　　金：国家自然科学基金资助项目（30371226）;教育部博士点专项基金资助项目（20050285009）;江苏省重点实验室开放课题（KJS0622）

摘　　要：目的构建氡染毒小鼠骨髓细胞差异表达基因的cDNA文库。方法采用HD-3型多功能氡室对雄性BALB／c小鼠动态吸入染毒，建立实验动物模型。按照实验动物吸入氡及其子体的累积剂量分为实验组（105WLM）和对照组（室内本底浓度，1WLM）。取骨髓细胞提取总RNA，采用SMART和抑制性消减杂交技术，构建正、反向消减cDNA文库，消减cDNA片段插入pMD18-T载体转化大肠杆菌，以含差异表达cDNA片段的菌液为模板进行巢式PCR扩增。结果获得了完整无降解的总RNA，得到高效率的消减cDNA文库；克隆了244个有插入片段和长度在100～1100bp之间的单克隆。结论成功构建了氡暴露小鼠骨髓细胞差异表达的正、反向cDNA消减文库。Objective To construct and identify subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation, Methods Adult male BALB/c mice, weighing 18-22 g, were placed in a multi- functional radon chamber. One group of mice was exposed to radon up to the accumulative dose of 105 work level month （WLM）, The control group of mice was housed in a room with an accumulative dose of 1 WLM. To construct a subtracted cDNA library enriched with differentially expressed genes, the SMART technique and the suppression subtractive hybridization were performed, The obtained forward and reverse cDNA fragments were directly inserted into pMD18-T vector and transformed into E, coli JM109. The inserting cDNA fragments were screened by the blue-and-white blot screening and nested PCR of bacterium liquid, Results The 244 of 285 white bacteria clones obtained randomly were positive clones contained 100-1100 bp inserted cDNA fragments, Conclusions The forward and reverse subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation is successfully constructed.

关 键 词：小鼠 氡染毒 抑制性消减杂交 CDNA消减文库 

分 类 号：Q78[生物学—分子生物学]