LA-PCR-RFLP analysis of the chloroplast DNA in some species of Salvia L.

摘要

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the chloroplast DNA (cpDNA) was used to investigate phylogenetic relationships among 54 species of Salvia L. One long fragment of around 10 kilo base pair (kb) of cpDNA from Salvia was amplified using a universal pair of primers and Long Accurate-PCR (LA-PCR) protocol, then the amplified products were digested with each of six restriction enzymes; Eco RI, Hind III, Pvu II, Nde I, Sma I on Mfe I. Four restriction enzymes; Eco RI, Hind III, Pvu II, and Mfe I produced more polymorphic patterns than Nde I or Sma I. Polymorphic fragments were visually detected, documented and used to build a genetic similarity data matrix based on Nei and Li (1979) method. The genetic similarity matrix and PHYLIP software package (Felsenstein 2005) were the bases to construct a phylogenetic tree. The resulted tree divided Salvia species under study into distinct groups based on origin. Salvia roemeriana showed amplified fragment length difference (AFLD) and can be identified using any of the six enzymes. Ocimum species were distinguishable when LA-PCR products were digested with Mfe I. PCR-RFLP was tried in this study due to less labor-intensive, low cost and adequate value in primary phylogenetic studies.