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cje ›› 2010, Vol. 29 ›› Issue (02): 401-406.

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Optimization of SSR detection system forArabidopsis thaliana.

GAO Ha-rong1;ZHONG Ming1|LIU Wan2|LI Pei-jun2;WANG Lei1|ZHOU Qi-xing2   

  1. 1Key Laboratory of Agricultural Biotechnology of Liaoning Province, Shenyang Agricultural University, Shenyang 110161, China;2Key Laboratory of Terrestrial Ecological Process, Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang 110016, China
  • Online:2010-02-10 Published:2010-02-10

Abstract: Taking Arabidopsis thaliana as test material and by the methods of single factor selection and L16(45) orthogonal design, the amplification component, program, and electrophoresis detection in polymerase chain reaction (PCR) in simple sequence repeat (SSR)detection system were optimized. The optimized 25 μl reaction system contained 1×PCR Buffer, 20 ng DNA template, 1.5 mmol·L-1 of Mg2+, 0.3 μmol·L-1 of primers, 150 μmol·L-1of dNTPs, and 1.0 U of Taq DNA polymerase. The PCR amplification procedures consisted of an initial denaturing for 5 min at 94 ℃, followed by 30 cycles of denaturation for 30 s at 94 ℃, annealing for 30 s at 57 ℃, and an extension for 45 s at 72 ℃, with an ad
ditional extension period of 10 min at 72 ℃. By using nondenaturing polyacrylamide gel electrophoresis with ethidium bromide (EB) staining, the amplification products bands of SSR were easy to be identified. In the SSR system, amplification bands were clear and stable, and there were fewer nontarget bands.

Key words: Tea orchard, Arthropod assemblage, Diversity, Stability