Optimization of ISSR and RAPD-PCR Reaction Systems for Alternaria Small-spored Species(PDF)

《北方园艺》[ISSN:1001-0009/CN:23-1247/S]

Title:: Optimization of ISSR and RAPD-PCR Reaction Systems for Alternaria Small-spored Species

Author(s):: LIU Zhenya1; XIONG Renci1; 2; ZHU Tiansheng1; 2; （1.College of Plant Science，Tarim University，Alar，Xinjiang843300；2.Southern Xinjiang Key Laboratory of IPM of Tarim University,Alar，Xinjiang843300）

Keywords:: Alternaria small-spored species; ISSR; RAPD; orthogonal design; system optimization

Abstract:: Using13 Alternaria small-spored species as materials，the orthogonal design were used to optimize ISSR and RAPD reaction system for Alternaria small-spored species，four influding factors on ISSR and RAPD，including Mg2+，dNTPs，TaqDNA polymerase and primer at four levels.The results showed that the optimal ISSR and RAPD reaction system for Alternaria small-spored species were established as follows：in 25 μL ISSR-PCR system were 10×PCR Buffer 2.50 μL，2.50 mmol/L Mg2+，0.10 mmol/L dNTPs，0.30 μmol/L primer，1.25 U Taq DNA polymerase；in ISSR-PCR system were 10×PCR Buffer 2.50 μL，2.00 mmol/L Mg2+，0.15 mmol/L dNTPs，0.60 μmol/L primer，1.25 U Taq DNA polymerase；decided the annealing temperature of optimal reaction system，then ISSR primer UBC808 and RAPD primer OPE07 were used to carry out stable and adaption in the optimal ISSR and RAPD reaction system at the optimal annealing temperature，indicating that the optimal ISSR and RAPD reaction system were stable and adapted.

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Last Update: 2015-08-14