Journal of Pharmaceutical Practice 2020;38(1):42-46

doi:10.3969/j.issn.1006-0111.201909048

Establish and optimization of inter-simple sequence repeat PCR reaction system of Gnaphalium affine

Shenggui JIANG 1 ; Zhiying DONG 2 ; Qing LI 3 ; Yingkui ZHAO 4 ; Doudou HUANG 4 ; Lianna SUN 5 6

Affiliations

+expand

Keywords

Gnaphalium affine; inter-simple sequence repeat PCR; system optimization; primer screening

Country

China

Language

Chinese

Abstract

Objective To provide the experimental basis for the subsequent genetic diversity research through establishing and optimizing the inter-simple sequence repeat PCR (ISSR-PCR) reaction system of Gnaphalium affine. Methods The single-factor experimental method and full experimental method were used to optimize the ISSR-PCR reaction system of Gnaphalium affine. Under the optimal system, after screening primers and corresponding annealing temperatures, the systematic feasibility was verified. Results The optimal ISSR-PCR reaction system was consisted of 10 μl Premix Taq DNA polymerase, 0.3 μmol/L primer, 10 ng DNA template, and sterilized water added to 20 μl. Finally, 10 primers were screened from 100 universal primers, and verification results indicated the system had high stability, good reproducibility, and the selected primers had good polymorphism. Conclusion The ISSR-PCR amplification system of Gnaphalium affine was established for the first time and the primers with appropriate annealing temperatures were filtered out, which provided a reference for the subsequent genetic diversity research of Gnaphalium affine.