Development and Application of Mitochondrial and Chloroplast Microsatellite Markers for Codonopsis Plants / 中国实验方剂学杂志

ABSTRACT

Objective:To develop the specific molecular markers of <italic>Codonopsis</italic> plants and better identify their germplasm resources considering the significant difference in active ingredients of Codonopsis Radix from various origins and producing areas. Method:Such bioinformatics software as Primer 5.0， NTSYS-pc 2.10e， and PopGene 32 were used for searching the simple sequence repeat （SSR） markers of <italic>C. minima </italic>chloroplast genome， <italic>C. tsinlingensis</italic> chloroplast， and <italic>C. lanceolata </italic>mitochondrial sequences， and 120 pairs of SSR primers were designed by Primer 5.0. Then 16 pairs of cpSSR primers and 10 pairs of mtSSR primers with good screening effect and high polymorphism were selected for analyzing the interspecific versatility of 20 samples. Result:The results showed that 66 cpSSR primer sites and 26 mtSSR sites were identified from the genome sequences， with 86.20% of single nucleotide， 6.9% of dinucleotide， and 6.9% of trinucleotide for <italic>C. minima </italic>chloroplast， 83.78% of single nucleotide，13.51% of dinucleotide， and 2.71% of trinucleotide for <italic>C. tsinlingensis</italic> chloroplast， and 46.15% of single nucleotide and 53.85% of dinucleotide for <italic>C. lanceolata </italic>mitochondria. As demonstrated by polymerase chain reaction （PCR） identification results， the developed 26 pairs of SSR primers had good applicability in the genus<italic> Codonopsis</italic>. The analysis by NTSYS-pc 2.10e revealed that the genetic similarity coefficients of 20 samples were within the range of 0.38-1.00， and they were divided into two subgroups at a threshold of 0.69. Four pairs of polymorphic primers were screened out in the diversity analysis of 20 samples using PopGene 32. The number of observed alleles （<italic>Na</italic>） was 12， and the effective number of alleles （<italic>Ne</italic>） ranged from 1.362 9 to 2.605 9. The percentage of polymorphic loci （PPL） at each site was 100%， and the average values of genetic parameters<italic> Ho</italic>， <italic>He</italic>， and <italic>I</italic> at each site were 0.555 8， 0.444 2， and 0.753 2， respectively， indicating high polymorphism at each site. The screened four pairs of primers were utilized for DNA fingerprinting of the 20 samples， and it was found that the DNA fingerprints enabled the identification of these 20 samples. Conclusion:This study has provided a molecular basis for the study of the genetic relationship between plants in species <italic>Codonopsis</italic> and the intraspecific genetic differentiation.