蜡梅科(Calycanthaceae)若干植物nrDNA ITS序列PCR反应条件分析

Exploration of the Optimal PCR Reaction Conditions of nrDNA's ITS Region Among Some Members in Calycanthaceae

该文旨在探讨适合于扩增蜡梅科植物ITS序列的PCR反应条件。试验中设计了2个新引物来扩增整个核糖体RNA基因(nrDNA)的ITS序列,进行了4种引物组合、2种扩增程序、7种Mg^(2+)浓度、4种dNTP浓度和3种二甲基亚砜浓度的比较。结果表明:当反应体系中Mg^(2+)浓度为1.8 mmol/L,dNTP为2 mmol/L,含5%二甲基亚砜,扩增时先进行两步预扩增,反应效果最好。文中还对引物的设计进行了探讨,认为当所选用的植物可能有寄生真菌时,最好不用与真菌同源的引物。最后提出了几点引物设计的注意事项。

The purpose of this experiment is to find out the optimal PCR reaction conditions of nrDNA's ITS region for detecting the phylogenetic relationship among members of Calycanthaceae.Two new primers were de- signed to amplify the whole nuclear rRNA genes' internal transcribed spacer.Four pairs of primers,two amplifi- cation procedures,several kinds of Mg^(2+),dNTP,DMSO concentration levels were used to attain the expected re- sults.As well,the authors come up with several points concerning primer design.