论文部分内容阅读
以玉米开放群体为试材，对引物种类，复性温度，引物、ｄＮＴＰＳ、Ｍｇ＋＋、缓冲液、Ｔａｑ聚合酶浓度等因素对ＲＡＰＤ带型的影响进行了比较研究。结果表明，在不同复性温度下，有的引物在个体间扩增相同的带型，有的引物在个体间扩增不同的带型。同一引物高浓度比低浓度扩增信号强。高浓度ｄＮＴＰＳ比低浓度ｄＮＴＰＳ扩增效果差。低浓度Ｍｇ＋＋无扩增产物。高浓度Ｍｇ＋＋和高浓度聚合酶引起带纹模糊拖尾。高浓度缓冲液无扩增信号。双引物扩增可增加带数。根据以上研究结果，建立了适合玉米ＲＡＰＤ分析的技术体系：２５μｌ反应液中，含１倍缓冲液，１．５ｍｍｏｌ／ＬＭｇＣｌ２，０．２ｍｍｏｌ／ＬｄＡＴＰ，０．２ｍｍｏｌ／ＬｄＴＴＰ，０．２ｍｍｏｌ／ＬｄＣＴＰ，０．２ｍｍｏｌ／ＬｄＧＴＰ，０．８μｍｏｌ／Ｌ随机引物，１．５ＵＴａｑ酶，６０ｎｇ模板ＤＮＡ。
The effects of different primer types, refolding temperature, primer, dNTPS, Mg ++, buffer and Taq polymerase concentration on the RAPD patterns were compared among the open corn populations. The results showed that at different refolding temperatures, some primers amplified the same banding patterns among individuals, and some of the primers amplified different banding patterns among individuals. The same primer high concentration than low concentration amplification signal. High concentrations of dNTPS are less effective than low concentrations of dNTPS. Low concentration of Mg ++ no amplification products. High concentrations of Mg ++ and high concentrations of polymerases cause smeared smear. High concentration buffer without amplification signal. Double-primer amplification can increase the number of bands. According to the above results, a technical system suitable for RAPD analysis of maize was established. The reaction system consisted of 1-fold buffer, 1.5mmol / L MgCl2, 0.2mmol / LdATP, 0.2mmol / LdTTP, 0.2mmol / LdCTP, 0.2mmol / LdGTP, 0.8μmol / L random primer, 1.5UTaq enzyme, 60ng template DNA.