Identification and candidate gene screening of qCIR9.1, a novel QTL associated with anther culturability in rice (Oryza sativa L.)

Abstract

A novel QTL, qCIR9.1, that controls callus induction rate in anther culture was identified on chromosome 9 in rice, and based on RNA-seq data, Os09g0551600 was the most promising candidate gene. Anther culture, a doubled haploid (DH) technique, has become an important technology in many plant-breeding programmes. Although anther culturability is the key factor in this technique, its genetic mechanisms in rice remain poorly understood. In this study, we mapped quantitative trait loci (QTLs) responsible for anther culturability by using 192 recombinant inbred lines (RILs) derived from YZX (Oryza sativa ssp. indica) × 02428 (Oryza sativa ssp. japonica) and a high-density bin map. A total of eight QTLs for anther culturability were detected in three environments. Among these QTLs, a novel major QTL for callus induction rate (CIR) named qCIR9.1 was repeatedly mapped to a ~ 100kb genomic interval on chromosome 9 and explained 8.39-14.14% of the phenotypic variation. Additionally, RNA sequencing (RNA-seq) was performed for the parents (YZX and 02428), low- (L-Pool) and high-CIR RILs (H-Pool) after 16 and 26days of culture. By using the RNA of the bulked RILs for background normalization, the number of differentially expressed genes (DEGs) both between the parents and between the bulked RILs after 26days of culture was drastically reduced to only 78. Among these DEGs, only one gene, Os09g0551600, encoding a high-mobility group (HMG) protein, was located in the candidate region of qCIR9.1. qRT-PCR analysis of Os09g0551600 showed the same results as RNA-seq, and the expression of this gene was decreased in the low-callus-induction parent (YZX) and L-Pool. Our results provide a foundational step for further cloning of qCIR9.1 and will be very useful for improving anther culturability in rice.