Genome-wide association analysis revealed that GhD6PKL2 regulates cold tolerance at seed germination and seedling emergence in cotton

Low temperature (12 ℃) is an abiotic stress factor that adversely affects cotton yield and fiber quality. Exposure to low-temperature stress during seed germination and seedling emergence significantly reduces crop productivity. In this study, 383 cotton germplasms were subjected to low-temperature stress, and the variation in seven seedling emergence-related phenotypic traits, including germination rate, was determined. Through genome-wide association analysis (GWAS), we identified 331 significantly associated single nucleotide polymorphism (SNP) loci, which were mapped to traits including hypocotyl length, total seedling length, dry weight, and germination rate. Notably, a quantitative trait locus (QTL) associated with hypocotyl length was identified on chromosome A08 (51.91–51.93 Mb). Further analysis revealed that this significant genomic region harbors polymorphic sites within the promoter of GhD6PKL2 . Transcriptome sequencing and quantitative real-time PCR (qRT-PCR) further revealed significant differences in this gene's expression levels across distinct haplotypes. Virus-induced gene silencing (VIGS) of GhD6PKL2 demonstrated that TRV2: GhD6PKL2 plants exhibited hypocotyl elongation under low-temperature treatment, accompanied by reduced cotton germination rates,indicating that this gene positively regulates cold resistance in cotton. This study provides valuable theoretical foundations for the breeding and improvement of low-temperature tolerant cotton varieties. This study investigated the genetic basis of low-temperature tolerance in cotton by analyzing 383 germplasms under 12°C stress, which revealed 331 significant SNP loci associated with seedling emergence traits. A major QTL on chromosome A08 (51.91–51.93 Mb) was correlated with hypocotyl length, and the GhD6PKL2 gene was identified as a critical regulator. • Prior cotton cold resistance research has relatively few studies on 12 ℃. • A total of 383 cotton germplasms was employed for GWAS of cold tolerance. • A novel QTL(51.91–51.93 Mb) located on chromosome A08 was identified.