杂草稻叶绿体psbA-trnH片段的PCR扩增条件优化

Optimization on the PCR Amplification Conditions of Weedy Rice Chloroplast DNA psbA-trnH Fragment

对杂草稻叶绿体psbA-trnH片段的PCR扩增条件进行了优化,建立的最优体系为:20μL反应体积含1μL模板DNA,0.125mM dNTPs,0.5μM正反向引物,1U Taq酶,2.0μL 10×Taq PCR buffer;退火温度为61℃,在此条件下充分保证了psbA-trnH PCR产物的质量和纯度要求。对该片段进行了直接测序,结果为510bp左右,在NCBI中比较分析表明其测序结果准确可靠,经ClustW分析表明含有丰富的系统学信息。

The PCR conditions of weedy rice chloroplast psbA-trnH fragment were optimized. The PCR reactions were carried out in the conditions of 1μL genomie DNA, 0.125mM dNTPs, 0.5μM forward and reverse primer, 2μLIO×PCR reaction buffer, adjusted to 20μLwith sterilized H2O. The annealing temperature was 61℃. The quality and purity of PCR products were assured using these optimal conditions. The sequencing length of the psbA-trnH fragment was about 510bp using the direct sequencing methods and consistent with the NCBI Genebank data. The Clust W results showed that the abundant phylogenetic information was contained.