Fine Mapping Of Dwarfing Gene In Wheat Mutant Je0105 And Mechanism Analysis Of Gibberellin

Plant height is an important agronomic trait in wheat,which is closely related to lodging resistance and therefore influences yield.Until now,only several dwarfing genes in wheat have been cloned.Mapping and cloning of genes affecting plant height in wheat is of great significance.In this study,we used a dwarf mutant je0105 induced by EMS mutagenesis and the wildtype（WT）Jing 411 as materials,and fine-mapped the dwarfing gene through exome capture sequencing,genome-wide resequencing,and genetic linkage analysis.Meanwhile,the gibberellin mechanism in je0105 was explored through exogenous gibberellin treatment and detection the content of endogenous gibberellin.The main results are as follows:1.By investigation plant height in the field,the result suggested that the plant height of WT and je0105 were 93 cm and 73 cm,respectively.Comparing with WT,the plant height of je0105 was reduced by 27.4%.The yield data showed that there was no significant difference in thousand grain weight between WT and je0105 at Changping and Zhongpuchang field stations,and the grain number per spike in je0105 was significantly increased comparing with that in WT.Taken together,je0105 is an elite dwarfing mutant with no negative impact on yield.2.Exome capture sequencing suggested that the dwarfing gene in je0105 was mapped on the interval of 11.59-81.96 Mb on chromosome 2DS.The dwarfing gene was further mapped within the interval of 13.6-29.5 Mb by development of 7 KASP markers on 2DS.The high density molecular markers were developed through SNPs obtained by genome-wide resequencing,and then the dwarfing gene was fine-mapped to a 173-kb interval between markers dw14 and dw15 by analysis of genotype and phenotype in an enlarged F₂ population.3.Based on Chinese Spring reference genome V1.0,we found no annotated genes in the mapped interval.According to the reference genomes of Jing 411 and Fielder,a candidate gene Traes CSU03G0022100 with multiple copy numbers was found in the mapped interval.Sequencing of this gene indicated that a G-to-A substitution in je0105at position of the 201^th resulted in premature stop codon in Traes CSU03G0022100.According to the mapping interval and sequencing analysis,we found that the dwarf gene was a new allelic variation of Rht8.4.Both WT and je0105 grown in the greenhouse were treated with gibberellin acid（GA₃）and GA synthesis inhibitor paclobutrazol(PP₃₃₃),the plant height of WT and je0105 increased by 24.2%and 30.9%after treatment with GA₃,while that was decreased by 25.3%and 25.7%with PP₃₃₃ treatment,respectively.In the field experiment,the plant height of WT and je0105 spraying with GA₃ increased by 20.9%and 32.2%,respectively.Expression analysis of gibberellin synthesis genes suggested that the expression level of GA13ox was significantly down-regulated in the mutant.Endogenous gibberellin detection indicated that the content of bioactive GA₃ was significantly decreased in je0105.Taken together,these results suggested that je0105 was sensitive to gibberellin acid.The dwarf phenotype in je0105 may result from the decrease of GA13ox expression and thus decrease in bioactive GA₃content.In this study,we identified a candidate gene leading to reduction of plant height in the mutant by gene mapping and sequencing analysis.Exogenous GA treatment,GA content and gene expression analysis suggested that the dwarfism of je0105 was caused by the decrease of bioactive GA₃ content.These results provide basis for molecular mechanism study on regulation of plant height and also facilitate development of wheat varieties with proper height.