A Family-Level Approach to Identify Loci Controlling Soluble Solid Content in Blackberry for DNA-Informed Breeding

Sweeter berries are a target of interest for many blackberry (Rubus L.) breeders. As part of the USDA-NIFA SCRI funded RosBREED project, genes associated with fruit sugar production in blackberry were identified using a family-level candidate gene approach. A total of 279 Rubus transcripts were identified from the Rubus reference transcriptome (https://www.rosaceae.org/analysis/227) using sugar candidate genes from apple (Malus domestica), peach (Prunus persica), and alpine strawberry (Fragaria vesca). These genes were used to design 9,355 Hyb-Seq baits at a 2X tiling density that covered 99.6% of the targeted genes. The baits were used in conjunction with PacBio sequencing to genotype 40 cultivars with high and low soluble solids content (SSC) from the University of Arkansas and USDA blackberry breeding programs. Sequencing and filtering of the captured genomic reads for the 40 samples produced 430,167 high-quality circular consensus (CCS) reads. The number of CCS reads were variable and ranged from 62 to 37,369 reads per individual. The mean and median read lengths were 2,661 and 2,610 nucleotides, respectively. The quality of the CCS reads was high and a mean phred score of 40 was observed. The CCS reads were aligned to the 'Hillquist' v1.0 blackberry and Rubus occidentalis v3.0 genomes, followed by variant identification. Welch's t-test and a Benjamini-Hochberg correction identified 467 and 312 loci from the 'Hillquist' and the R. occidentalis genotype tables, respectively, significantly (alpha = 0.05) associated with SSC. Population structure modeling identified a total of 173 alleles that were significantly (alpha = 0.05) associated with sugar production regardless of population structure. KASP genotyping was conducted on a validation set of blackberries from each breeding program for 92 of the alleles and 48 markers were identified that were significantly associated with SSC. One QTL identified in both breeding programs accounted for a 1.46 Brix increase and is in the intron space of a sucrose synthase gene. This is the first environmentally stable SSC QTL identified in blackberry. Moreover, the approach explored in this study can be used to develop breeding tools for other crops that have not yet benefited from the genomics revolution.