Identification on mating types of monokaryons and their hybrids of Lentinula edodes by AS-PCR

Abstract:

[Background] The mating types and mon-mon hybrids of edible mushrooms are usually identified based on the presence of clamp connections or not as observed by microscopy, which is time-consuming, labor-costing, and prone to false results. [Objective] A molecular-assisted breeding technique was established for identifying the monokaryon mating type and mon-mon hybrids of Lentinula edodes, aiming to provide technical support for improving the breeding efficiency. [Methods] Typing primers were designed based on the SNP loci in the conserved sequence of mating-type factors, and then allele specific (AS)-PCR was employed to identify the mating type of the single spore isolates of L. edodes strains L808 and YX7 and their mon-mon hybrids. [Results] Among the 38 single spore isolates of L808, there were 6, 13, 8, and 11 monokaryons of mating types A1B1, A2B2, A1B2, and A2B1, respectively. Among the 45 single spore isolates of YX7, there were 15, 8, 8, and 12 monokaryons of mating types A3B3, A4B4, A3B4, and A4B3, respectively, and 2 heterokaryons of the mating type A3A4B3B4. The 12 mon-mon hybrids included 10 heterozygotes and 2 non-heterozygotes. The identification results obtained based on the conventional method were consistent with those obtained by AS-PCR, except that the former was prone to misjudging heterokaryons as monokaryons. [Conclusion] The AS-PCR based on SNP loci can identify the monokaryon mating type and mon-mon hybrids of L. edodes and distinguish between monokaryons and heterokaryons, with high precision and efficiency. It serves as an ideal tool for molecular-assisted breeding of L. edodes.