猪鼻腔中细菌菌落PCR-DGGE引物条件优化

The Optimization of Bacterial Colonies PCR-DGGE Primers in Pig Nasal Cavity

变性梯度凝胶电泳(Denaturing gradient gel electrophoresis,DGGE)技术是最先用于DNA突变检测的一种电泳技术。为获得最佳的PCR-DGGE上样模板,本实验分别采用十六烷基三甲基溴化铵(CTAB)法和Power Soil DNA Isolation Kit试剂盒抽提细菌基因组DNA,分别以Primer(338F/518R)/Primer(P2/P3)为16Sr DNA V3扩增引物,采用温度梯度PCR对引物扩增条件进行优化。结果显示:在Tm 56℃,通过CTAB法抽提细菌基因组DNA,采用primer(338F/518R)扩增16Sr DNA V3目的产物特异性最佳。

Denaturing gradient gel electrophoresis （DGGE） technology is the first electrophoresis technique to detect DNA mutations .In order to obtainthe optimalof PCR-DGGE template, cetyltrimethyl ammonium bromide （CTAB） method and the Power Soil DNA Isolation Kitare used to extractbacterialmetagenome. Besides, Primer（338F/518R）and Primer（P2/P3）were respectivelyused for 16Sr DNA V3 amplification, temperature gradient PCRwere optimizedfor primer amplification conditions. Results showed thatextracting bacteriametagenome by CTAB and usingprimer A （338 f / 518 r） , are the best way of 16Sr DNA amplification productin Tm 56℃.