桃SRAP-PCR反应体系建立及与半矮生型相关标记的研究

试验采用常规CTAB法提取了桃叶片基因组DNA,并以半矮生型桃基因组DNA为模板,通过对影响扩增结果的Mg2+浓度、dNTP浓度、Taq酶浓度及引物浓度的梯度试验,对桃SRAP-PCR反应体系进行了优化.结果表明:桃SRAP-PCR的最佳反应体系为DNA模板25～30ng,10×Buffer2μL,Mg2+浓度2.5mmol·L-1,dNTP浓度0.25mmol·L-1,Taq酶1.1U,引物浓度0.3μmol·L-1.通过BSA法建立半矮生型和普通型基因池,从266对SRAP引物中筛选出21对与桃半矮生性状相关的SRAP标记,并通过对18个个体的扩增检测,发现了1个与半矮生性状相关的标记.

Establishment of SRAP-PCR reaction system and preliminary research on the marker linked to semi-dwarf phenotype in peach

Total genomic DNA was isolated from fresh leaves of peach using CTAB method and it was as template for PCR amplification. The concentrations of Mg2+, dNTPs, Taq DNA polymerase and primers considering the most important four factors affecting the SRAP-PCR reactions were optimized. In the present study the optimum system of 20 mL volume was as follows:template DNA 25-30 ng,10×Buffer 2 mL,Mg2+ 2.5 mmol·mL-1,dNTPs 0.25 mmol·L-1,Taq DNA polymerase 1.1 U, primer 0.3 mmol·L-1.An efficient, convenient and high resolution method for silver staining was also established. Using BSA method, 21 pairs of SRAP primer related were selected from 266 ones and only one marker linked to the semi-dwarf phenotype of peach based on a progeny of 18 individuals.