Creation Of Novel Early Flowering Germplasm Using TaLATE1 Genes In Wheat

Wheat（Triticum aestivum）is one of the important food crops in the world,and various adverse weather disasters have seriously affected the production of wheat.Therefore,timely heading and harvesting are particularly important for wheat production.The flowering time of wheat is an important trait that has a significant impact on wheat adaptation to environments,as well as on the timely maturation and quality of seeds.We found that the heading time of wheat variety Jimai 22 treated with vernalization for26 days is significantly later than that of wheat treated with vernalization for 33 and 40 days,indicating that the duration of vernalization will affect the flowering time of Jimai 22.RNA-seq sequencing analysis of vernalized stem tips revealed a group of wheat genes with higher expression levels at 26 days of vernalization than at 33 and 40 days of vernalization,and we found that this group of genes are homologous to the LATE FLOWERING（LATE）gene of Arabidopsis thaliana.This group of wheat genes were named as Ta LATE1-5A,Ta LATE1-4B,and Ta LATE1-4D.Then,we performed a preliminary analysis of Ta LATE1s genes.The results are as follows:The phylogenetic tree analysis of the LATE1 homologous proteins in wheat and other plants revealed that Ta LATE1s were closely related to the LATE1 proteins of gramineous plants such as T.turgidum and T.urartu.Both Arabidopsis LATE and wheat Ta LATE1s have conserved C₂H₂domains and EAR motifs with similar distribution positions.Ta LATE1s were expressed in various tissues of wheat,but the expression level is higher in leaves and stem apex,suggesting that Ta LATE1s may play a role in leaf and stem apex in wheat.Under long-day and short-day conditions,the expression of Ta LATE1s may not be affected by circadian rhythm.Transient expression of 35S::Ta LATE1-4D-GFP in tobacco leaves revealed that Ta LATE1-4D was localized in the nucleus.We edited the genome sequences of Ta LATE1s in Fielder through the CRISPR/Cas9 gene editing system and obtained three independed mutant lines,and two mutant lines with AAbbdd genotype were selected for phenotypic observation and agronomic traits statistics.Under long-day conditions,compared to the wild type Fielder,the heading time of the talate1-3 and talate1-14 mutants was significantly advanced.Compared to the wild type Fielder,the plant height and tiller number of the talate1-3 mutant were significantly reduced,and the number of florets per spike was significantly reduced.Compared with those of the wild type,the spikelet number per spike and the number of florets per spike of the talate1-14mutant were significantly reduced,and the hundred-grain weight significantly increased.The talate1-3 and talate1-14 mutants exhibited early heading phenotype under long-day conditions,while the heading time of them was same with that of the wild type under short-day conditions,indicating that the Ta LATE1-4B and Ta LATE1-4D are involved in the regulation of wheat early-flowering time under long-day conditions.In summary,we created a novel wheat early-flowering germplasm through editing Ta LATE1s,providing genetic resources for molecular design and breeding of wheat.