摘要
以树莓优良野生种质插田泡(Rubus coreanus)为材料,比较了SDS法、CTAB法和改良CTAB法提取树莓基因组DNA的效果,结果发现三种方法均能提取到树莓DNA,但改良CTAB法优于CTAB法和SDS法,其所得的DNA质量最好,杂质最少。通过正交试验设计,对Taq酶,primer,dNTP和DNA用量进行了筛选,建立了树莓ISSR技术最优反应体系,即25μl反应体系中含有1×PCR Buffer,2mmol/LMg2+,1.5UTaq酶,0.2μmol/L primer,0.4mmol/L dNTP和20ngDNA。采用引物UBC835和UBC873分别对10个树莓品种和8个树莓野生资源进行检验发现,该体系工作效果良好。
Using one of the excellent germplasm resources of bramble Rubus coreanus as material, three different DNA-extracting methods （SDS method, CTAB method and modified CTAB method） were employed to be compared in order to isolate high-quality genomic DNA from young leaves. The results showed that the SDS method, the CTAB method and the modified CTAB method can extract DNA. But the modified CTAB method was better than the other two methods, because it could obtain high quality DNA with less impurity. By orthogonal design, a satisfactory ISSR reaction system was established, i.e. a total volume of 25 μl system, it contained 1 ×PCR Buffer, 2 mmol/L Mg^2＋, 1.5 U Taq DNA polymerase, 0.2 μmol/L primer, 0.4 mmol/L dNTP and 20 ng DNA template. We tested this reaction system and it worked well with primer UBC835 using in ten bramble cuhivars and UBC873 using in eight bramble wild materials.
出处
《中国农学通报》
CSCD
北大核心
2009年第14期27-31,共5页
Chinese Agricultural Science Bulletin
基金
国家自然科学基金"西南地区树莓植物遗传多样性研究与特异资源的筛选"(30671454)
关键词
树莓
ISSR
DNA提取
体系优化
bramble, ISSR, DNA extraction, reaction system optimization
