Blueberries (Vaccinium corymbosum L.) benefit from increased fruit firmness because of consumer preference and machine harvestability. However, the genetic component of fruit texture and skin thickness and their relationship to firmness have yet to be deciphered. This study used bulked segregant RNA-seq (BSR-seq) for differential gene expression analysis. Previously an F1 population of a cross between firm-fruited southern highbush cv. 'Reveille' and soft-fruited cv. 'Arlen' was developed in our laboratory. The total RNA of the parents, the two softest, and the two firmest progenies, were extracted at the breaker and fully ripe stages. Next-generation sequencing cDNA libraries were constructed and subjected to Illumina short-read sequencing. Subsequently, the short reads were mapped to a blueberry genome, and differentially expressed genes (DEGs) were identified in immature and mature fruit coded for potentially biologically significant proteins such as expansins, polygalacturonase, polygalacturonase-inhibiting protein, and mannosidase. Additionally, DEGs corresponding to cysteine proteases and S-adenyl methyltransferases (SAM-MTases) that were previously reported as candidate genes for blueberry firmness were identified in this study. Our results indicated that BSR-seq is a promising method for identifying major candidate genes controlling complex traits in blueberry. 

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