Study Of Genetic Variation By Genomics In Pear Population

Pear germplasms have widely genetic variations,which are important bases for formatting different phenotype traits.In order to analyze genetic variations of diffenrent pear germplasms totally and explore the important traits about potentially regulated genes to serve the molecular breeding,this research carried out pear genomic DNA level,mitochondrial DNA and chloroplast DNA level,and expressed sequence level to reveal pear genetic variations by genomics research methods.The main results were as follows:At first,we used 312 pears(Pyres py.)for whole genomic resequencing,and then achived a total of 1,273,308 SNPs.Our natural population of 312 pears had two subpopulations based on the population structure analysis,principle content analysis and phylogram.GWAS was performed on agronomic traits of three continuous years,and twelve agronomic traits had significantly associated loci.Ten traits were about fruit quality(fruit shape,flesh color,number of fruit russet,core position,broad base of fruit stem,furrows on fruit surface,fruit-stem status,fruit weight,flesh texture and number of stone cells),another two were climatic traits(date for fruit mature and days of fruit development).Then the associated loci were identified,and genes distributed in that regions were annotated.What was more,it was found that MLM with considering kinship and PCA was the most appropriate method for our pear population.The second,the transcriptomes of five main representative pear cultivars,'Hosui'(Pyrus pyrifolia),'Yali'(Pyrus bretschneideri),'Kuerlexiangli'(Pyrus sinkiangensis),'Nanguoli'(Pyrus ussuriensis),and 'Starkrimson'(Pyrus communis)were sequenced at seven key fruit developmental stages,from fruit setting to maturation and fruit senescence after harvesting.In total,33,136 genes that could be mapped by reads,were analyzed.Most gene expression cluster models showed a steadily decreasing trend.Gene expression patterns had obvious differences according to maturity type,that is,post-ripening cultivars were still vigorous at maturity,and showed a higher proportion of up-regulated genes;non post-ripening cultivars had a gradually decreasing tendency during fruit maturation.Meanwhile,differentially expressed genes related to fruit quality and development,such as stone cells,sugar,acid and hormones,were identified.Co-expression analysis revealed that several ethylene synthesis genes and polyphenoloxidase-related genes interacted with each other directly,and an indirect relationship was reflected between ethylene synthesis genes and ethylene response genes.In addition,the highly diverse SNPs represented the great differences between oriental and occidental pears.Transcriptome of 'Dangshansuli' from 12 different tissues were newly assembled in pear and a total of 17,353 SSR loci were identified.We developed 194 EST-SSR makers from the assembled sequences to test on seven pear varieties from different genetic backgrounds.A total of 132 EST-SSR primers had amplifications on these genotypes.There were 108 primers with dinucleotide and trinucleotide motifs,these primers detected a total of 311 alleles,with an average per primer of 2.88 and 2.86,respectively,and displayed better quality than that of tetranucleotides,pentanucleotides and hexanucleotides.Fifty polymorphic markers divided 36 pear genotypes into two groups with a similarity coefficient of 0.62.Results showed that Japanese pears had a comparatively close relationship with Chinese pears in dendrogram,this indicated great potential application of noval EST-SSR makers in future research on evaluation of pear germplasm.Further more,54 EST-SSR markers were selected to test transferability to other 96 Rosaceae cultivars from eight species including Japanese apricot,plum,apricot,peach,cherry,apple,strawberry and loquat.Seventeen EST-SSR primers had PCR amplification products in at least one of the 96 accessions,and the highest transferability was from pear to apple at 29.63%.Transferability from pear to Japanese apricot,plum,apricot,peach,cherry,strawberry and loquat was 9.26%,14.81%,11.11%,12.96%,5.56%,3.7%and 12.96%,respectively.Therefore,the newly developed EST-SSR markers can contribute to exploring the genetic relationship,evolution and comparative genomics,and gene introgressions in the Rosaceae family.At last,the complete pear mitochondrial genome was assembled,and genome size is 458,897 bp,with G+C contents of 45.2%.A total of twenty-eight open reading frames in the assembled mitochondrial genome were annotated,including fifteen electron transport complex,one intron maturase,one transport membrane,five cytochrome C biogenesis.What is more,twenty tRNA,three rRNA genes and eight ribosomal subunits were also presented.The co-linearity of mitochondrion from distinct species varied a lot,but the gene region had a relatively high co-linearity.There were 129 pear samples distributed around the world to reveal the pear classification and wild origin in our study.Phylogenic tree of chloroplast presented that occidental pears manifested great differences from the oriental pears in the genetic level,and the same species had genetic variation because of geographic distribution;while mitochondrial phylogenic tree mainly showed the pear inheritance trend.Maternal inheritance should be the main inheriting method for pear mitochondrion and chloroplast except of few pears.Furthermore,we suppose pear mitochondrial genome is less diverse than the chloroplast genome based on mutation rates and tandom repeats.Comparative mitochondrial gene expression revealed a potential tRNA gene perhaps responsible for pear post mature character.