金龟甲基因组DNA提取方法比较研究

摘 要：为了筛选适宜金龟甲RAPD-PCR的基因组DNA提取方法，于2009年在青岛农业大学实验室内，分别采用改良的SDS法、平衡酚法、裂解液法和CTAB法提取棕色鳃金龟(Holotrichia titanis Reitter)的基因组DNA，通过DNA沉淀性状观察、产量和质量分析，以及随机引物PCR (RAPD-PCR)比较，筛选最优的金龟甲基因组DNA提取方法。研究结果表明，改良的SDS法提取的基因组DNA产量较高，但纯度较低，并且降解严重，RAPD-PCR扩增谱带弥散较严重；裂解液法提取的基因组DNA纯度较高，降解程度较低，但产量最低，RAPD-PCR扩增谱带弥散较严重，并且存在非特异扩增现象；CTAB法提取的基因组DNA不仅产量和纯度较低，降解严重，而且RAPD-PCR扩增结果不稳定，具非特异扩增现象；平衡酚法提取的基因组DNA产量和纯度高，降解轻微，RAPD-PCR扩增结果稳定，扩增谱带弥散轻微。因此，在上述4种方法中，平衡酚法是最适合PAPD-PCR的金龟甲基因组DNA提取方法，裂解液法和改良的SDS法次之，CTAB法最差。

Comparison of DNA Extraction Methods from Scarab

Abstract: In order to select the proper extraction method of genome DNA from scarab suitable for RAPD –PCR, genome DNA of Holotrichia titanis Reitter was extracted by Improved SDS, Balance Phenol, Lysis Solution and CTAB Method in the laboratory of Qingdao Agriculture University in 2009, and the most proper extraction method was selected by observation on the character of DNA deposition, production & quality analyzing and RAPD-PCR. The results showed that the genome DNA extracted by Improved SDS Method was higher in production, but lower in purity, degraded seriously, and could generate seriously dispersed RAPD –PCR fingerprinting when amplifying with it as template. Genome DNA extracted by Lysis Solution Method was higher in purity, degraded slightly, but lowest in production, and could generate seriously dispersed and nontypical RAPD –PCR fingerprinting when amplifying with it as template. Genome DNA extracted by CTAB Method was not only lower in purity and production, degraded seriously, but also could not produce stable RAPD –PCR fingerprinting when using it as template, and there was nontypical amplification too. Genome DNA extracted by Balance Phenol Method was higher in purity and production, degraded slightly, and could produce stable RAPD –PCR fingerprinting without dispersion when using it as template. So among above four methods, Balance Phenol Method is the most suitable method to extract genome DNA from scarab, which is to be used for PAPD-PCR, Lysis Solution and Improved SDS Method take the second place, and CTAB Method is the most ill-fitted one.