摘要
以鱼呼肠孤病毒双链RNA为模板 ,建立一种快速、简便、高效的cDNA合成及克隆策略。在一定量模板条件下 ,采用随机六聚体为引物合成cDNA第一链。以退火方式形成双链cDNA ,直接通过琼脂糖凝胶电泳检测其cDNA合成产量。双链cDNA经两端补齐后 ,平头连接于具有阳性选择标记的载体中 ,以高效电转化的方式进行快速克隆。
A rapid, simple and very efficient method for synthesis and cloning of cDNA is described , by taking fish reovirus dsRNA as template. The technique include: Under the fixed quantity of template, the first strand of cDNA is synthesesed by using random hexamers as primer. The two strands of cDNA are annealed and checked on 1% agarose gel directly. The ds-cDNA is blunt-end ligated into the pZErO2.0 vector which has positive selective mark, then cloned by electroporation transformation with high efficiency.
出处
《生物技术》
CAS
CSCD
2000年第1期6-10,共5页
Biotechnology
基金
国家自然科学基金!(39570035)
关键词
双链RNA
CDNA合成
CDNA文库
基因克隆
double-stranded RNA
random hexamers
synthesis of cDNA
highly efficient cloning
