Identification of Candidate Genes Conferring Cold Tolerance to Rice (Oryza sativa L.) at the Bud-Bursting Stage Using Bulk Segregant Analysis Sequencing and Linkage Mapping

Abstract

Low-temperature tolerance during the bud-bursting stage is an important characteristic of direct-seeded rice. The identification of cold-tolerance quantitative trait loci (QTL) in species that can stably tolerate cold environments is crucial for the molecular breeding of rice with such traits. In our study, high-throughput QTL-sequencing analyses were performed in a 460-individual F₂ _: ₃ mapping population to identify the major QTL genomic regions governing cold tolerance at the bud-bursting (CTBB) stage in rice. A novel major QTL, <i>qCTBB9</i>, which controls seed survival rate (SR) under low-temperature conditions of 5°C/9 days, was mapped on the 5.40-Mb interval on chromosome 9. Twenty-six non-synonymous single-nucleotide polymorphism (nSNP) markers were designed for the <i>qCTBB9</i> region based on re-sequencing data and local QTL mapping conducted using traditional linkage analysis. We mapped <i>qCTBB9</i> to a 483.87-kb region containing 58 annotated genes, among which six predicted genes contained nine nSNP loci. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis revealed that only Os09g0444200 was strongly induced by cold stress. Haplotype analysis further confirmed that the SNP 1,654,225 bp in the Os09g0444200 coding region plays a key role in regulating the cold tolerance of rice. These results suggest that Os09g0444200 is a potential candidate for <i>qCTBB9</i>. Our results are of great significance to explore the genetic mechanism of rice CTBB and to improve the cold tolerance of rice varieties by marker-assisted selection.