甲状腺癌组织差异表达cDNA消减文库的构建与鉴定

目的 :应用抑制性消减杂交技术 ,构建人甲状腺癌与正常甲状腺差异表达的cDNA消减文库。方法 :分别从甲状腺癌及正常甲状腺组织中提取总RNA ,应用高效、灵敏的抑制性消减杂交技术 ,构建成功cDNA消减文库 ,再转染大肠杆菌进行文库扩增。结果 :构建成功具有高消减效率的人甲状腺癌组织cDNA消减文库 ,文库扩增得到了 4 0 0个克隆 ,随机挑取 5 0个制备质粒 ,酶切分析均得到 5 0～ 4 0 0bp大小插入片段 ,这些片段可能就是载有高度特异性的目的片段 ,结论 :人甲状腺癌组织cDNA消减文库的建立为进一步克隆甲状腺癌特异性表达的新基因奠定了基础。

Construction and Identification for Differential Expression cDNA Subtractive Lib rary in Thyroid Carcinoma Tissue

Objective To construct a cDNA subtractive library of human thyroid carcinoma with the technique of suppression subtractive hybridization(SSH).Methods Total RNA were isolated from tissue of thyroid carcinoma and normal thyroid respectively.By using the suppression subtractive hybridization (SSH),The subtractive library which contained the differently expressing cDNAs between thyroid carcinoma and normal thyroid tissue were constructed.Amplification of the library was carried out with the E.coli strain Top 10F'.Results Human thyroid carcinoma subtractive library with high subtractive efficiency was set up successfully.The amplified library contained 400 positive clones.Random analysis of 40 clones with enzyme restriction showed that all plasmids in the clones contained 50-400 bp inserts.Conclusion The constructed cDNA subtractive library of human thyroid carcinoma is a highly efficient one and plays solid foundation for cloning new and specific oncogenes or tumor suppressor genes of thyroid.