Establishment and Optimization of ISSR-PCR Reaction System of Ligustrum lucidum Ait.
doi: 10.15886/j.cnki.rdswxb.2010.02.012
- Received Date: 2010-03-29
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Key words:
- Ligustrum lucidum Ait. /
- ISSR /
- influential factors /
- optimization
Abstract: Single factor experiment was used to test the effect factors of ISSR-PCR reaction system.After the optimization,a suitable ISSR-PCR reaction system of Ligustrum lucidum Ait.was established as follows:10 × buffer 2.5 μL,2.0 mmol·L-1 MgCl2,200 μmol·L-1 dNTPs,1.5 U Taq polymerase,0.4 μmol·L-1 primers,10 ng DNA template,which were contained in 25 μL reaction solution.The optimized amplification program was that predenaturing at 94 ℃ for 4 min,denaturing at 94 ℃ for 40 s,annealing at 52 ℃—60 ℃ for 45 s,extension at 72 ℃ for 120 s,for 35 cycles,extension at 72 ℃ for 8 min at last.The products were stored at 4 ℃.The optimized ISSR-PCR experiment system was applied in the amplification of 12 varieties of germplasm materials of Ligustrum lucidum Ait and the results indicated that the rich and stable bands can be amplified.
Citation: | LI Chuan-dai, CHEN Fei, WANG Tian-di, LIU Guo-min. Establishment and Optimization of ISSR-PCR Reaction System of Ligustrum lucidum Ait.[J]. Journal of Tropical Biology, 2010, 1(2): 99-104. doi: 10.15886/j.cnki.rdswxb.2010.02.012 |