摘要
目的： 构建藏族外周血淋巴细胞全长cDNA文库并鉴定其质量.方法： 分离健康藏族人外周血液的淋巴细胞并提取总RNA, 利用SMART（switching mechanism at 5′ end of RNA transcript）技术, 构建全长cDNA文库并进行文库滴度和重组率的测定.扩增文库并随机挑取噬菌斑进行PCR鉴定插入片段的大小.结果： 构建的cDNA文库的滴度为1.8×109pfu/L, 重组率〉98%, 文库扩增后的滴度达8×1012pfu/L, 插入片段的长度在0.75～2.0 kb之间, 平均长度为1.35 kb.结论： 构建的藏族外周血淋巴细胞cDNA文库具有很高的质量, 为进一步筛选、克隆藏族低氧反应相关基因奠定坚实的基础.
AIM： To construct the full length cDNA library of the peripheral blood lymphocytes in Tibetans and to identify the quality of the library. METHODS： The lymphocytes were separated from healthy Tibetan fresh blood, and total RNA was extracted, The full length cDNA library was constructed by SMART （switching mechanism at 5＇ end of RNA transcript） technique. The unamplified library was titered, and the percentage of recombinant clones were determined. The library was amplified, and the size of the inserts was identified by PCR. RESULTS： The titer of the constructed cDNA library was 1.8 ×10^9 pfu/L with the percentage of recombinant clones above 98%, and that of the amplified library was 8 × 10^12 pfu/L. The insert＇s size ranged from 0.75 to 2.0 kb with the average length of 1.35 kb. CONCLUSION： The constructed cDNA library of peripheral blood lymphocytes of Tibetans has excellent quality, and it lays a solid founclation for further screening and cloning hypoxia-related genes of Tibetans living at high altitudes,
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2006年第2期148-150,共3页
Chinese Journal of Cellular and Molecular Immunology
基金
国家自然科学基金资助项目(No.30260034)
