Optimization of SRAPPCR reaction system for Harmonia axyridis using orthogonal design
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Key Words：Harmonia axyridis, SRAP, orthogonal design, molecular marker, natural enemy
Abstract：The orthogonal design was used to optimize SRAP-PCR reaction system for Harmonia axyridis (Pallas) using five factors (Taq DNA polymerase, Mg2+, DNA template, dNTPs and primer) at four levels, respectively. The results of PCR were analyzed by DPS and MINITAB and a most suitable SRAP_PCR system for H. axyridis was established. The 20 μL volume system contained 50～100 ng DNA template, 0.25 μmol/L primers, 0.1 mmol/L dNTPs, 1.5 U Taq DNA polymerase and 0.375～0.625 mmol/L Mg 2+. The optimal annealing emperature for SRAP-PCR was 50.3℃ based on gradient PCR. This system provided the opportunity for analysis of diversity, construction of genetic map and gene location in H. axyridis.