Establishment of ISSR-PCR in Lycoris radiata

YANG Zhi-ling; FENG Gang-li; TAN Zi-feng; LUAN Qi-fu; WANG Cheng-nan

In this paper,the influencing factors of inter-simple sequence repeat(ISSR) were analyzed by using genomic DNA of Lycoris radiata.By adjusting the concentration of template DNA,Mg2+,dNTPs and primer,the contents of Taq DNA polymerase and the different temperature,the stable and reproducible optimum reaction system of ISSR-PCR amplification and PCR amplification parameters were established for L.radiata.The optimal system contains 20 ng template DNA,1.0 U Taq DNA polymerase,2.0 mmol·L-1 Mg2+,150 μmol·L-1 dNTPs and 0.5 mmol·L-1 primer in 25 μL reaction volume.The amplification program was: after l cycle initial denaturation at 94 ℃ for 5 min,each 45 cycles consist of 94 ℃ 45 s,55 ℃ 50 s,72 ℃ 2 min,then final extension was at 72 ℃ for 7 min.

1. Research Institute of Subtropical Forestry, CAF, Fuyang 311400, Zhejiang, China

2. College of Resources and Environment of CSFU, Changsha 410004, Hu'nan, China

3. College of Resources and Environment of CSFU, Changsha 410004, Hu'nan, China

Received Date: 2005-08-29

Abstract: In this paper,the influencing factors of inter-simple sequence repeat(ISSR) were analyzed by using genomic DNA of Lycoris radiata.By adjusting the concentration of template DNA,Mg2+,dNTPs and primer,the contents of Taq DNA polymerase and the different temperature,the stable and reproducible optimum reaction system of ISSR-PCR amplification and PCR amplification parameters were established for L.radiata.The optimal system contains 20 ng template DNA,1.0 U Taq DNA polymerase,2.0 mmol·L-1 Mg2+,150 μmol·L-1 dNTPs and 0.5 mmol·L-1 primer in 25 μL reaction volume.The amplification program was: after l cycle initial denaturation at 94 ℃ for 5 min,each 45 cycles consist of 94 ℃ 45 s,55 ℃ 50 s,72 ℃ 2 min,then final extension was at 72 ℃ for 7 min.

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Reference (20)

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Establishment of ISSR-PCR in Lycoris radiata

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