分子植物育种
分子植物育種
분자식물육충
MOLECULAR PLANT BREEDING
2013年
4期
611-616
,共6页
菜豆%ISSR%体系优化
菜豆%ISSR%體繫優化
채두%ISSR%체계우화
Phaseolus vulgaris L.%ISSR%System optimization
用试剂盒法提取菜豆(Phaseolus vulgaris L.)嫩叶的基因组DNA,并以此为模板对影响ISSR-PCR反应体系的Taq酶量、模板浓度、dNTP浓度、Mg2+浓度、引物浓度5个因素进行了优化。结果表明总体积为25μL的菜豆ISSR-PCR反应最优体系为:45 ng模板DNA,1 U Taq酶,2 mmol/L Mg2+,0.6μmol/L引物和0.1 mmol/L dNTP,在此基础上对循环次数进行优化,最终确定最佳循环次数为45次。利用该体系,选取引物823对8份材料进行扩增,验证了该体系的稳定性。本研究旨在建立菜豆ISSR反应的最优体系,为今后利用该分子标记技术进行有关菜豆种质及遗传方面的研究奠定技术基础。
用試劑盒法提取菜豆(Phaseolus vulgaris L.)嫩葉的基因組DNA,併以此為模闆對影響ISSR-PCR反應體繫的Taq酶量、模闆濃度、dNTP濃度、Mg2+濃度、引物濃度5箇因素進行瞭優化。結果錶明總體積為25μL的菜豆ISSR-PCR反應最優體繫為:45 ng模闆DNA,1 U Taq酶,2 mmol/L Mg2+,0.6μmol/L引物和0.1 mmol/L dNTP,在此基礎上對循環次數進行優化,最終確定最佳循環次數為45次。利用該體繫,選取引物823對8份材料進行擴增,驗證瞭該體繫的穩定性。本研究旨在建立菜豆ISSR反應的最優體繫,為今後利用該分子標記技術進行有關菜豆種質及遺傳方麵的研究奠定技術基礎。
용시제합법제취채두(Phaseolus vulgaris L.)눈협적기인조DNA,병이차위모판대영향ISSR-PCR반응체계적Taq매량、모판농도、dNTP농도、Mg2+농도、인물농도5개인소진행료우화。결과표명총체적위25μL적채두ISSR-PCR반응최우체계위:45 ng모판DNA,1 U Taq매,2 mmol/L Mg2+,0.6μmol/L인물화0.1 mmol/L dNTP,재차기출상대순배차수진행우화,최종학정최가순배차수위45차。이용해체계,선취인물823대8빈재료진행확증,험증료해체계적은정성。본연구지재건립채두ISSR반응적최우체계,위금후이용해분자표기기술진행유관채두충질급유전방면적연구전정기술기출。
The genomic DNA of common bean was extracted from tender leaves by DNA secure plant kit method, and optimized ISSR reaction condition with it as template. We established optimum ISSR-PCR system for common bean using ISSR-PCR amplification with five factors (Taq DNA polymerase, DNA template, dNTP, Mg2+concen-tration and primer). The optimum reaction system with 25μL total reaction volume contains 45 ng DNA template, 1 U Taq DNA polymerase, 2 mmol/L Mg2+, 0.6μmol/L ISSR primer and 0.1 mmol/L dNTP. The suitable cycles was 45 cycles that screened from three different treatments. 8 soybeans were easily amplified with primer 823 under the optimized reaction conditions. This reaction system will provide a technological base for germplasm and genetic study in common bean.