小鼠短指/缺指畸形肢体cDNA消减文库的构建

被引：1

作者：

陈蓉芳

张天宝

印木泉

朱江波

郭苗莉

机构：

[1] 第二军医大学卫生毒理学教研室,第二军医大学卫生毒理学教研室,第二军医大学卫生毒理学教研室,第二军医大学卫生毒理学教研室,第二军医大学卫生毒理学教研室上海,上海,上海,上海,上海

来源：

中国药理学与毒理学杂志 | 2003年 / 04期

关键词：

短指; 少指; 基因表达; 基因文库;

D O I：

暂无

中图分类号：

R346 [];

学科分类号：

1001 ;

摘要：

目的　为筛选、克隆小鼠短指 /缺指畸形肢体的特异表达基因 ,构建了GD18小鼠畸形肢体 (短指 /缺指 )与正常肢体的差异表达的cDNA消减文库。方法　应用抑制消减杂交技术 (SSH ) ,结合PCRcDNA合成法 ,将从总RNA中合成的双链cDNA ,酶切成平均大小为 4 0 0～ 6 0 0bp的片段 ,经接头连接 ,两次消减杂交和两次抑制性PCR后 ,将产物与T/A载体连接构建cDNA消减文库 ,并转染大肠杆菌进行文库扩增 ,菌液PCR筛选有插入片段的SSH克隆。结果　成功地构建了GD18小鼠畸形肢体(短指 /缺指 )差异表达的cDNA消减文库 ,从中挑取2 75个克隆进行菌液PCR分析 ,结果显示其中 2 5 5均得到 2 0 0～ 6 0 0bp插入片段。结论　SSH与PCR合成双链cDNA ,是一种自微量总RNA中构建高特异性的差异表达cDNA消减文库的有效方法。消减文库的构建 ,有助于筛选、克隆小鼠短指 /缺指畸形肢体的特异表达基因

引用

页码：281 / 285

页数：5

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