ZHANG Yunfeng,QU Anjiang,FU Xinmeng.Establishment and Optimization of ISSR-PCR System of Pleurotus ostreatus by PAGE Detection Technology[J].Northern Horticulture,2017,41(02):91-95.[doi:10.11937/bfyy.201702022]
基于PAGE检测技术的平菇ISSR-PCR体系的建立和优化
- Title:
- Establishment and Optimization of ISSR-PCR System of Pleurotus ostreatus by PAGE Detection Technology
- 关键词:
- 平菇; ISSR-PCR体系; PAGE
- Keywords:
- Pleurotus ostreatus; ISSR-PCR; PAGE
- 文献标志码:
- A
- 摘要:
- 以平菇“唐平26”基因组DNA为模板，以U836为引物，比较了琼脂糖凝胶电泳和不同浓度聚丙烯酰氨凝胶（PAGE）电泳对ISSR-PCR产物的分离效果，并且采用PAGE电泳技术对影响ISSR反应体系的主要因素进行优化。结果表明：以PAGE电泳技术检测平菇ISSR-PCR产物的效果最好，最佳ISSR-PCR反应体系为，在20 μL 反应体系中，模板20 ng，Mg2+浓度为1.5 mmol?L-1，引物0.8 μmol?L-1，dNTPs 200 μmol?L-1，Taq酶1.0 U，30个扩增循环。
- Abstract:
- The separation efficiency of ISSR-PCR product was detected by agarose gel electrophoresis and different concentrations of polyacrylamide gel (PAGE) electrophoresis，with Pleurotus ostreatus‘Tangping26’genomic DNA as template and with primers U836 as primer.The results showed that the separation of PAGE was significantly better compared to agarose gel.Main factors which had an influence on ISSR reaction system were optimized based on the PAGE detection technology，that the optimum ISSR-PCR reaction system was 20 μL total volume including 20 ng template，1.5 mmol?L-1 Mg2+，0.8 μmol?L-1 primer，200 μmol?L-1 dNTPs，1.0 U Taq enzyme，30 cycles of amplification.
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备注/Memo
第一作者简介：张运峰（1982-），男，硕士，实验师，现主要从事微生物分子遗传学等研究工作。E-mail：zyfikx@126.com.基金项目：河北省自然科学基金资助项目（C2014105067）；唐山师范学院科学研究基金资助项目（2016C05）；河北省大学生创新创业计划资助项目（201510099032）。