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Bulletin of Botanical Research ›› 2020, Vol. 40 ›› Issue (1): 66-72.doi: 10.7525/j.issn.1673-5102.2020.01.010

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Optimization of SCoT-PCR System and Screening of Effective Primers for Castanopsis wenchangensis,an Extremely Small Population Endemic to Hainan Province

YANG Li-Rong1,2, SUN Xiu-Xiu1,2, CHEN Jia-Li1,2, YUN Yong1,2, CHEN Xuan1,2, ZHENG Dao-Jun1,2   

  1. 1. Institute of Tropical Horticulture Research in Hainan Academy of Agricultural Sciences, Haikou 571100;
    2. The Key Laboratory of Tropic Special Economic Plant Innovation and Utilization, Haikou 571100
  • Received:2019-09-03 Online:2020-01-05 Published:2020-02-21
  • Supported by:
    Key research and development Program in Hainan Province(ZDYF2018076);The Project for Technology Development of Hainan Provincial Scientific Research Institutions,2019(Collection,conservation and evaluation of tropic special economic plant germplasm resources)

Abstract: The effects of DNA, primers and dNTPs, Taq DNA polymerase on the SCoT-PCR amplification results of Castanopsis wenchangensis were analyzed by single factor test and orthogonal test, and the SCoT-PCR system of C.wenchangensis was optimized. The polymorphism primer combination was screened based on the established reaction system, which provided the conditions for the study of SCoT molecular markers in the study of C.wenchangensis germplasm resources. The results showed that the influence of each factor on SCoT-PCR amplification was DNA > dNTPs > primer > Taq enzyme, and the optimum reaction system was as follows:when the total system was 20 μL, DNA content was 2.5 ng, primer concentration was 0.8 μmol·L-1, dNTPs concentration was 0.2 mmol·L-1, Taq content was 1 U, and the amplified products were clear and stable. In addition, 15 primers with good polymorphism and suitable for C.wenchangensis were screened out from 80 SCoT primers by this system, which provided theoretical basis and technical support for the related research of C.wenchangensis and other plants of Fagaceae by SCoT molecular marker technique in the future.

Key words: Castanopsis wenchangensis, SCoT, reaction system optimization, effectiveness primer

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