高级检索+

RT-PCR联合一步法扩增G蛋白α亚基基因的开放阅读框

A COUPLED ONE-STEP METHOD OF REVERSE TRANSCRIPTION(RT)FOR GENERATION OF OPEN READING OF G PROTEIN αSUBUNIT GENE

  • 摘要: 报道逆转录-多聚酶链式反应(RT-PCR)联合一步程序, 扩增大于1kb的拟南芥菜G蛋白α亚基基因的开放阅读框。该程序中, 当RNA模板和引物变性, 并在同一管中加入PCR缓冲液、4种dNTP底物、逆转录酶和TaqDNA聚合酶之后, 就不需其它手工操作步骤, 因而可同时进行大批量样品的操作。实验结果证明AMV(鸟类骨髓性白血病病毒)逆转录酶对TaqDNA聚合酶的活性没有抑制作用。这种RT-PCR联合一步法可从0.5μg的总RNA中快速地扩增长于1.0kb的cDNA片段。

     

    Abstract: It is reported that a one-step reverse transcription and polymerase chain reaction (RT-PCR)procedure for amplification of open reading frame of G-protein αsubunit gene from Arabidopsis thaliana which is larger than 1kb. In the procedure developed, manual work is minimized, larger numbers of samples can be handled. After denaturation of template RNAand the primers and addition of other regents, no further manual steps are needed, It was not found that AMVreverse transcriptase inhibited the activity of Taq-DNA polymerase, so the coupled one-step procedure of RT-PCRrapidly amplifies ORFjust only from 0.5μg amount of total RNA.

     

/

返回文章
返回