Genetic Diversity Of The Wild Auricularia Heimuer In China And Its Whole Genome Analyses

A uricularia heimuer F.Wu,B.K.Cui&Y.C.Dai,as one of the most important edible mushrooms in China,is popular with many people due to its various natural compounds containing pharmacologically active ingredients.Since China has the largest production of A.heimuer in the world,evaluation of genetic diversity and structure of the nationwide wild A.heimuer will provide the basis for breeding selection and development of new varieties.Previous studies on A.heimuer were mainly focused on cultivation,breeding and nutrient composition,but there were few researches on the genome of A.heimuer.Sequencing and analyses of the whole genome and primary characterization of its gene funtions were conducted in the present study,therefore playing significant roles for further studies on metabolic pathways and regulatory mechanisms of A.heimuer.In this study,ITS and IGS1 sequences of 118 samples of four Chinese wild Auricularia heimuer populations were analyzed,and their genetic diversity and structure were explored at DNA sequence level.In addition,genome sequencing and characterization of the single-spore strain Dai 13782 were fully investigated.The results of the study are as follows:(1)The genetic diversity of Auricularia heimuer was analyzed based on the sequences of ITS and IGS1.The results showed that the total length of ITS sequence was 530 bp,including 6 mutation sites,which produced 17 haplotypes(Hd=0.8579,Pi=0.0043);while the IGS1 sequence was 818 bp,containing 12 polymorphic sites,and a total of 32 haplotypes were produced(Hd=0.9148,Pi=0.0047).The population of the Chinese wild A.heimuer exhibited a high level of haplotype diversity and a low level of nucleotide diversity,and the haplotypes in different groups was abundant.It was indicated that different geographical populations of A.heimuer showed strong ability to adapt different environments.(2)The genetic structure of Auricularia heimuer were analyzed according to ITS and IGS1 sequences.The Fst of the total population of A.heimuer was 0.2470 and 0.1219,respectively,suggesting that its degree of genetic differentiation was indistinct.However,there was a significant genetic differentiation between the northeast(NEC)population and other three populations.The results of genetic differentiation coefficient(Gst>Nst,P<0.5)showed that A.heimuer had no geographical lineage structure at species level.AMOVA variance analysis demonstrated that the variation of A.heimuer populations mainly existed within group,and the gene exchange between groups was frequent and the difference was relatively small.Neutrality tests and mismatch distribution analyses showed that the population of A.heimuer did not experience large-scale expansion events and stayed in dynamic equilibrium during the evolutionary history.(3)The whole genome sequencing of the high-throughput sequencing technology combined with Illumina Hiseq 4000 and Pacbio RSI1 longread sequencing platform of the single-spore strain Dai 13782 was performed without the reference genome.The genome size was 49.76 Mb,with the GC content of 56.98%.A total number of 16,244 genes were obtained,and the total length of protein coding region was 22.01 Mb,with a total intron length of 6.83 Mb.15,135 genes accounted for 93.17%of the total genes were functionally annotated,which paved the way for functional gene screening and analysis in depth(4)Of the Auricularia heimuer genome,six gene clusters composed of five terpene synthases clusters and one non-ribosomal peptide synthetases gene cluster were identified to participate in the biosynthesis of secondary metabolite.None of these gene cluster involved in PKS The reasons for this phenomenon are still unknown and need to be further studied.Filtering and bioinformatics analyses of genes involved in the synthesis of polysaccharides and terpenoids revealed that A.heimuer genome existed a large number of polysaccharides and terpenoids synthesis related genes.A total of 33 genes were predicted to participate in polysaccharide biosynthesis,22 genes were predicted to be involved in terpenoid backbone biosynthesis,3 genes related to sesquiterpenoid and triterpenoid biosynthesis,and 27 genes participated in biosynthesis of steroidal.The results provide basic information for the intensive study of the synthesis of polysaccharides and terpenoids in A.heimuer at different stages of growth.The future research could center on the analyses of its metabolic pathways and regulatory mechanisms to furnish theory evidence for improving the nutritional value of A.heimuer.(5)Through the cluster analysis of gene family,a total of 18,059 gene families were found.There were 8,167 gene families in the Auricularia heimuer and 268 were unique gene families,which may be related to its species-specificity.A.heimuer genome had 1,039 single-copy homologue genes,1,198 multi-copy genes and 1,005 genes in the specific gene families.There were 1,411 unique genes which unclustered in gene families.The KEGG annotation was performed on the genes in the specific gene families of A.heimuer.The result showed that 11 genes was annotated by KEGG,in which the gene(GLEAN 0002067)was involved in the biosynthesis of terpenoids,and this gene was the encoding gene of 624 sterol reductase.