Development and Application of Abundance SNP Markers for Auricularia heimuer
The genome of Auricularia heimuer Dai13782 was used as the reference genome,while the genomes of A.subglabra SS5,A.auricula-judae B14-8,A.cornea CCMJ2827,and A.cornea ACW001 from the NCBI database were utilized as the test data.The genome variation data of A.heimuer and its related species were analyzed using Bwa,Samtools and Bcftools.The genet-ic relationship matrix was obtained using Tassel 5 software,and the genetic relationships among five strains were visualized using the pheatmap package.VCFtools-0.1.16 software was used to filter and extract abundant single nucleotide polymorphisms(aSNPs),specifically aSNP regions with a SNP count equal to or greater than 50 in a 300 bp sliding window.Subsequently,47 407 pairs of primers were designed using Primer 3 software.E-PCR results showed that 1 020 pairs of primers met the experi-mental design,involving 8 aSNP regions.Each region retained 1 pair of specific primers,which were distributed in 6 contig re-gions of the reference genome.In this experiment,the commonly used ITS and LSU were employed as reference markers,and the genetic relationship of 13 A.heimuer strains was analyzed using aSNP markers.The results showed that the phylogenetic relation-ships revealed by aSNP markers were generally consistent with the results of ITS and LSU markers,and even better able to distin-guish closely related A.heimuer strains.On the whole,it reflects that A.heimuer strains are relatively close to A.corne,and far-thest from A.subglabra.This result is consistent with the genetic relationship revealed by the whole-genome SNP analysis,which also confirms the reliability of the results based on aSNP sequence analysis.The aSNP marker sequence interval developed in the experiment contains numerous mutation sites,which is expected to serve as a beneficial supplement to traditional target analysis se-quences such as ITS,Rpb2,and Ef1α,providing assistance for the systematic development and genetic identification of A.heim-uer and other germplasm resources.
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