摘要
采用CTAB改良法,以苹果梨及其变异单系叶片为材料,进行苹果梨基因组DNA提取及RAPD扩增条件优化。结果表明,提取的DNA质量较高,适宜于RAPD分析;RAPD扩增最佳反应体系为25μL(15 ng DNA模板,1.0 mmol.L-1 dNTP,1.0 mmol.L-1 MgCl2,2.5μL Buffer,15 pmol.L-1引物,1U TaqDNA聚合酶,15.67μL ddH2O)。扩增反应程序为94℃预变性5 min,94℃变性1 min,37℃退火1 min,72℃延伸1.5 min;45个循环;最后72℃延伸7min;终止温度为4℃。
With fresh leaves of Apple-pear and variation strains as the materials, the extraction of genomic DNA and the optimization of RAPD analytic conditions were studied. The results showed that the high-quality genomic DNA was obtained by the improved CTAB method. The optimal PCR system for RAPD analysis was as follows： 15ng template DNA,1.0mmol·L^-1 dNTP,1.0mmol·L^-1 MgCl2,2.5μL Buffer,15pmol·L^-1 random primer,lU Tatl polymerase,15.67μL dd H2O in 25μL reaction system. The program of amplifying reaction was as follows： After pre-denaturing at 94℃ for 5rain,under the condition of dena turing at 94℃ for lmin,annealing at 37℃ for lmin,extension at 72℃ for 1.5min, amplifying for 45cycles,extension at 72℃ for 7min at last, stop at 4℃.
出处
《湖北农业科学》
北大核心
2007年第1期21-23,共3页
Hubei Agricultural Sciences
关键词
苹果梨变异单系
基因组DNA
扩增产物
优化
Apple-pear variation strains
genomic DNA
amplification products
optimization
