High-density Genetic Linkage Map Construction And QTL Indentification For Important Fruit Quality Traits In Apple

Molecular markers and genetic linkage maps are efficient tools for molecular breeding in woody perennials such as apple. In this study, the F1population derived from ’Jonathan’ x’Golden Delicious’ and their two parents were used for genetic map construction and SNP markers development via molecular biological techniques, RADseq and re-sequencing.Both EcoRI and HindⅢ were used to digest the genomic DNA in RADseq. An LOD score of6.0was set to distinguish linkage groups, and Kosambi’s mapping function in regression mapping algorithm was used to estimate genetic distance between markers. Finally, a high-density genetic map with3441SNP markers from297individuals was generated. Of the3441markers,2017were mapped to ’Jonathan’ with a length of1343.4cM and a density of0.67cM per marker,1932were mapped to ’Golden Delicious’ with a length of1516.0cM and a density of0.78cM per marker. Basd on the map, twelve significant QTLs for important fruit quality traits were mapped in apple. In which, six QTLs linked to fruit acidity were mapped on J08（3）, G08（1）, J07and J15; two QTLs of fruit weight were located on J03and J05; two fruit firmness associated QTLs were detected on J11; also two QTLs for sugar content were mapped to JO1.Among these QTLs, eight were major QTLs with more than20%variance explaination. QTLs linked to malic, citric and total acid were all mapped to J08and associated with marker emC08.14121899. The peak LOD score of qtlma.j8and qtlta.j8were both7.7, and explained39.1%and39.2%of the phenotypic variance, respectively, while qtlc.j8can explain27.2%of the phenotypic variance with an LOD of5.5. Another major QTL for citric acid was located on J15linked with marker huC15.29186826, with an LOD score of3.6and a20.2%variance explaination. Acetic acid related QTL, qtla.j7, was associated with marker euLG07₀14, and the peak LOD score was3.7with a20.1%explanation of variance, qtlf.j1（LOD=4.3,28.8%）, which contributed to fruit fructose content, was mapped to J01, and the nearest marker was huC01.18233570. There were two QTLs related to fruit firmness located on J11, qtlff.j11a （LOD=4.4,27.5%） was associated with euC11.7408490and huC11.10309880, while qtlff.j11b （LOD=3.9,21.6%） was between marker hmC11.4173064and euC11.5719084.Later, candidate genes were searched in five QTL regions which were less than5cM and have corresponding positins on reference genome of apple. Based on gene annotation,80,64and17genes related to fruit weight, fruit firmness and fruit acidity, respectively, were obtained. Among the candidate genes associating fruit acidity, changes in the expression of MDP0000582174, an MYB4gene, were coordinated with changes in the fruit acidity of apple.In order to dig the SNP variation in two parents, a whole genome re-sequencing project was performed on both ’Jonathan’and’ Golden Delicious’. Pair-end sequencing strategy was used on Illumina Hiseq2000platform. The read length was100bp with an average of500bp insertion fragment, In total,4950346SNPs were detected in’ Jonathan’, with a density of210bp per marker, and2222816SNPs were found in’ Golden Deicious’, the marker density was293bp per marker. Compared the heterozygosity with ’Golden Deicious’（237bp per locus）, it is higher in ’Jonathan’（206bp per locus）.Combined the data of genentic maps and re-sequencing, the phenomenon of cross-chromosomal translocation between Chr09and Chrl3in ’Jonathan’ has been proved preliminarily using PCR, which is the Chr13fragment insert into Chr09after replication. In addition, sequence variations of candidate genes between two parents were analyzed which can lay a foundation for their validation in future.