Abstract

To provide a fast genetic diversity survey of endangered Polyporus umbellatus (Pers.) Fries for immediate conservation, DNA isolation and optimization of polymerase chain reaction(PCR) assay of sequence-related amplified polymorphism (SRAP) were investigated. Due to high amount of polysaccharide contained in mycelium, three special approaches were adopted to eliminate it during DNA isolation, including adding RNAiso-mate for Plant Tissue buffer to mycelium powder, ethanol to DNA extraction buffer and 5% NaCl solution to the mixture of isoamyl alcohol and DNA deposit solution. Based on screening design, the optimal SRAP-PCR condition was a total volume of 25 μl containing 20 ng of DNA template, forward primer 0.4 μM, reverse primer 0.4 μM, 1× Taq MasterMix and the best annealing temperature was 35/50°C for each primer combination. According to our optimal SRAP-PCR condition, 49 out of 81 primer combinations were chosen for their high clarity and repetition in all samples.

Key words: DNA isolation, sequence-related amplified polymorphism (SRAP), optimalpolymerase chain reaction (PCR) condition, chuling, Polyporus umbellatus.