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《Acta Prataculturae Sinica》 2008-04
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Optimization of SRAP-PCR system on centipedegrass(Eremochloa ophiuroides) using orthogonal design and selection of primers

ZHENG Yi-qi1,2,WANG Zhi-yong2,GUO Hai-lin2,XUE Dan-dan2,LIU Jian-xiu2 (1.Department of Horticulture,Nanjing Agricultural University,Nanjing 210095,China;2.Institute of Botany,Jiangsu Province and Chinese Academy of Sciences,Nanjing Botanical Garden Mem.Sun Yat-sen,Nanjing 210014,China)  
An orthogonal design was used to optimize a SRAP-PCR system with 4 factors(Mg2+,dNTP,primer and Taq polymerase) at 3 levels plus the concentration of template DNA.The optimized SRAP-PCR system for E.ophiuroides was:2 μL 10 ×PCR buffer,60 ng template DNA,Mg2+ 1.50 mmol/L,dNTP 260 μmol/L,primer 0.25 μmol/L,Taq DNA polymerase 0.5 U in a total of 20 μL reaction solution.Each factor had a different effect on the results of PCR.Mg2+ concentration had the greatest effect and Taq DNA polymerase had the least effect.The optimized SRAP-PCR system was tested on four E.ophiuroides germplasms and shown to be steady and reliable.26 primer combinations were selected with abundant polymorphism from 45 primer combinations.The optimized SRAP-PCR system and polymorphism primer combinations could be applied to molecular genetics research of E.ophiuroides.
【Fund】: 国家自然科学基金(30670200)资助
【CateGory Index】: S688.4
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