Allelic variation in an expansin, MdEXP-A1, contributes to flesh firmness at harvest in apples.

Abstract

Flesh firmness is a core quality trait in apple breeding because of its correlation with ripening and storage. Quantitative trait loci (QTLs) were analyzed through bulked segregant analysis sequence (BSA-seq) and comparative transcriptome analysis (RNA-seq) to explore the genetic basis of firmness formation. In this study, phenotypic data were collected at harvest from 251 F₁ hybrids derived from 'Ruiyang' and 'Scilate', the phenotype values of flesh firmness at harvest were extensively segregated for two consecutive years. A total of 11 candidate intervals were identified on chromosomes 03, 05, 06, 07, 13, and 16 via BSA-seq analysis. We characterized a major QTL on chromosome 16 and selected a candidate gene encoding expansin MdEXP-A1 by combining RNA-seq analysis. Furthermore, the genotype of Del-1166 (homozygous deletion) in the MdEXP-A1 promoter was closely associated with the super-hard phenotype of F₁ hybrids, which could be used as a functional marker for marker-assisted selection (MAS) in apple. Functional identification revealed that MdEXP-A1 positively expedited fruit softening in both apple fruits and tomatoes that overexpressed MdEXP-A1. Moreover, the promoter sequence of TE-1166 was experimentally validated containing two binding motifs of MdNAC1, and the absence of the MdEXP-A1 promoter fragment reduced its transcription activity. MdNAC1 also promotes the expression of MdEXP-A1, indicating its potential modulatory role in quality breeding. These findings provide novel insight into the genetic control of flesh firmness by MdEXP-A1.