摘要
目的 构建金仓鼠神经管cDNA文库,以筛选与神经管发育及神经管畸形发生相关的基因。 方法 用TRIZOLReagent提取金仓鼠神经管总RNA;用LD PCR法反转录合成双链cDNA;经蛋白酶K消化、SfiⅠ酶切、 CHROMASPIN 400柱分离去除小于500bp的片段;将cDNA与载体λTriplEX2按一定比例连接;经体外包装,建立 cDNA的噬菌体表达文库。 结果 cDNA文库容量为1.5×106pfu ml,重组率为99%,平均插入片段大于1kb。 结论 我们初步构建的金仓鼠神经管cDNA文库为高效的cDNA文库。
Objective Constructing golden hamster neural tube cDNA library in order to detect new genes associated with development of neural tube and neural tube defects. Methods Total RNAs were isolated from neural tube using TRIZOL Reagent; Doublestranded cDNAs were synthesized from total RNAs using LD-PCR method; The dsc DNAs were digested by proteinase K and Sfi Ⅰ, less than 500?bp fragments were separated by CHROMA SPIN-400, and then the cDNAs were ligated to λ TriplEX-2 vector.After packaging,the neural tube cDNA library was constructed. Results The titer of neural tube cDNA library was 1.5×10 6?pfu/ml.The recombination rat was 99%.The average insert was larger than 1?kb.Conclusion Our successfully constructed cDNA library is a full-length library with high efficiency,and can be used to detect the genes related to development of neural tube and neural tube defects.
出处
《解剖学报》
CAS
CSCD
北大核心
2005年第2期187-189,共3页
Acta Anatomica Sinica
基金
国家自然科学基金资助项目(30270701)
