TaRLK-4B, a novel gene regulating grain number per spike in wheat

Wheat is one of the most important food crops in the world, and grain number per spike (GNS) is one of the most important factors affecting wheat grain yield. Therefore, identifying genes controlling GNS is important for wheat production. However, wheat is an allohexaploid species and has a highly complex genome, and the isolation of wheat genes is challenging. In this study, we identified a candidate gene, TraesCS4B02G047100 ( TaRLK-4B ), associated with GNS using both quantitative trait locus (QTL) mapping and genome-wide association study (GWAS) based on RNA-Seq. We obtained three homozygous mutant lines, bb-1 , bb-2 and bb-3 , using the CRISPR/Cas9 gene editing system. Compared with Fielder (wild type, WT), the three mutant lines exhibited significant reductions in GNS, total spikelets per spike (TSS) and spike length (SL). These results indicated that TaRLK-4B positively regulates GNS and its related traits TSS and SL. The TaRLK-4B gene contains two exons and one intron and belongs to the largest subfamily of LRR-RLKs. We performed RNA-Seq analysis using spikes from the WT and bb-2 mutant line at the tillering stage. A total of 1,193 differentially expressed genes (DEGs) were identified, including previously cloned TaSPL17 homologous genes in the three subgenomes and their orthologous gene OsIPA1 ( OsSPL14 ) in rice. Combining RNA-Seq data of TaRLK-4B and DAP-Seq data of the TaSPL17-7D , we identified 136 overlapping genes which are likely downstream targets of TaSPL17-7D . Therefore, we hypothesized that TaRLK-4B represents a novel putative component of the TaSPL17 -centered regulatory network. In addition, haplotype analysis revealed that Haplotype 2 (Hap2) is a favorable haplotype for increasing GNS, but the thousand-grain weight (TGW) was not significantly different between Haplotype 1 (Hap1) and Hap2.