Identification of peach flower genotype (Non-showy/Showy), development of flower type related molecular markers

Abstract：【Objective】The purpose of this study was to supplement the classification of peach flower types, identify the germplasm resources of homozygous non-showy flower patterns, and develop flowertype-related molecular markers in peach, in order to provide new experimental materials and theoretical support for marker assisted breeding of ornamental peach.【Methods】The peach phenotypes of flower types were first investigated, then the genome-wide association analysis (GWAS) based on the structure variations (SV) was performed to locate the key loci identified in 327 peach germplasm resources. Polymerase chain reaction (PCR) amplification of transposons was performed in different populations, after the amplified PCR products were verified using 1% agarose gel electrophoresis. The coefficients of variation were calculated by chi-square test. The mining of the key variant loci was performed by the integrative genomics viewer (IGV) software to identify candidate intervals from 145 peach germplasm materials. The 192 inbreeding populations of Zhongpan 17 were used for kompetitive allele-specific PCR (KASP) experiments to verify the accuracy of genotypes and phenotypes.【Results】The flower-types(d) of peach were mainly divided into Non- showy, Showy and Chrysanthemum- shapes, among which the Non-showy type was dominant to the Showy type. The data of flower type localization analysis carried out so far were SNP mutations, and the flower-type (d) candidate gene was locked in the range of 14- 16 M on chromosome 8. However, GWAS results based on SNPs tended to locate larger intervals with more variant loci within the intervals, and it was still challenging to carry out key gene mining, while the SVs of variants could affect the function of genes over SNPs. A variant ms179 810 transposon was identified at Pp08: 14 518 604-14 521 291 bp, and GWAS localization analysis was performed by the identified SVs, using the resequencing results from 327 peach germplasm materials. The localization results showed that the transposon exhibited insertion in the non- showy locus? and deletion in the Showy locus? A total of 384 individuals concluding Ruiguang 39×08-9-106, Qinglang×Zhongyoupan 7 hybrid populations and the Zhongpan 17 inbreeding population were amplified to verify the accuracy of the transposons The chi-square test showed that the insertion of transposons was not significantly associated with the floral traits. But this did not mean that the error of the localization interval? and did not exclude the existence of other loci with high phenotypic association in the vicinity of the transposon. The resequencing data of 145 peach germplasm materials were used to find the variant loci related to flower type at the range of 200 kb in the transposon, and finally, a SNP variant was identified at 33 980 bp upstream of the transposon. The exact location was Pp: 14 484 624 bp. The locus had three types of variants, including CC, AC and AA. When the flower phenotype was Non-showy, the genotype was CC or AC, and when the flower phenotype was Showy, the genotype was AA. The accuracy of genotypes and phenotypes was 98.62%. However, the phenotypic comparison showed that the corolla diameter of non-showy type CC was smaller than AC, and the Non-showy type might be incompletely dominant to the Showy type. To verify this hypothesis, the inbreeding population of Zhongpan 17 (heterozygous Non-showy) was used for identification, we found that the genotype and phenotype segregation ratio of the homozygous Non- showy: heterozygous non- showy: Showy types in the inbreeding population were close to 1∶2∶1, and the accuracy was 98.44%, which conformed to Mendel's law of genetic segregation. The corolla diameter of the homozygous non-showy type was similar to the sepal diameter, (1.54±0.46) cm, while the corolla diameter of the heterogeneous non-showy type was generally (2.61±0.4) cm and the corolla diameter of Showy type was greater than 3.7 cm. The subsequent cluster analysis of the diameter size also showed that the homozygous and heterozygous non-showy types were more closely related to each other and more distantly related to the Showy type, resulting in the difficulty to accord with a traditional classification that did not separate homozygous and heterozygous non-showy types.【Conclusion】The Nonshowy flowers of peach we subdivided into homozygous and heterozygous Non-showy types for the first time, based on the variant type at Pp08:14 484 624 bp of the peach genome and the findings of the peach flower phenotype, and the molecular markers to identify homozygous non- showy, heterozygous Non- showy and Showy types were developed as well. The identification of homozygous non- showy germplasm resources would provide a clue for marker assisted selection for ornamental peach.