Genetic Effects And Breeding Selection Of TaBT1,the Gene Encoding Wheat Endosperm ADP-glucose Transporter

The increase in Chinese wheat production mainly depends on yield,and thousand kernel weight?TKW?contributes the most among the three components.As starch occupies 65-70%of wheat grain,starch synthesis can significantly affect wheat TKW and yield.The transmembrane transport of ADP-glucose,the substrate of starch synthesis,mediated by ADP-glucose transporter?BT1?is an important rate-limiting step in the process of starch synthesis.In this study,the genetic effects of TaBT1 were studied by interfering the expression of TaBT1 and conducting correlation analysis between TaBT1 and yield-related traits.Moreover,the selection of global wheat breeding process on TaBT1 was also analyzed by scanning 1305 modern cultivars from six major wheat production regions.The main results are summarized as follows:1.Three homoeologous genes of TaBT1 were cloned and mapped on wheat chromosome 6A,6B and 6D,respectively with Chinese Spring nulli-tetrasomic lines.TaBT1 were preferentially expressed in young spikes and developing grains,with the expression peaked in developing grains at 10 days post anthesis?DPA?.The expression level was different among three homologous genes,i.e.TaBT1-6A exhibited the highest level,-6D was slightly lower and-6B was substantially lower.2.Compared to the wild type,the expression of BT1 in three RNA interference?RNAi?transgenic lines at 10 DPA was significantly decreased at 20.8%?50.3%and 69.5%,respectively.The proportion of starch granules were also altered in transgenic RNAi lines,with the ratio of typeA/typeB obviously decreased under the same visual field.Grains after maturation became smaller,with the TKW decreased at 15.7%,31.1%and 47.3%,respectively.Moreover,the grain size also changed significantly,with grain length decreased at 0.7%,4.5%and 5.6%,grain width at 8.1%,13.2%and 23.3%,and grain thickness at10.1%,15.4%and 17.1%,respectively.In addition,the above verified that BT1 could significantly affect starch synthesis and wheat TKW.3.Thirty-six common wheat cultivars were used to resequence the TaBT1 gene.Sequence analysis showed that there was no polymorphism in neither TaBT1-6A nor TaBT1-6D loci of the hexaploid wheat,whereas 24 polymorphic sites were detected at TaBT1-6B.These variations formed three haplotypes,TaBT1-6B-Hap1,TaBT1-6B-Hap2 and TaBT1-6B-Hap3?Hereinafter referred to as Hap1,Hap2 and Hap3?.Two molecular markers,designated as CAPS-1664 and InDel-2029,were developed to discriminate the three haplotypes.Association analysis between markers and phenotypes indicated that Hap1 and Hap2 were favored haplotypes associated with higher TKW.4.The confirmatory results with introgression lines showed that the average TKW of the progenies of Hap1 was 50.6 g,which was 10.5%higher than that of Hap3,and the difference reached a significant level at P<0.05.Expression analysis showed that the expression abundance of TaBT1-6B in varieties of Hap1 was 1.6-fold and 4.3-fold higher than that of Hap2 and Hap3,respectively?P<0.01?.GUS activity assay showed that promoters of Hap1 was the strongest,which was 1.5-fold and 3.1-fold higher than that of Hap2 and Hap3,respectively?P<0.01?.The above demonstrated that variations in the promoter region may alter gene expression,which finally brought changes to the TKW differences among haplotypes.Furthermore,the promoter of favored haplotypes possessed relatively higher activity.5.The favored haplotypes,Hap1 and Hap2,underwent strong and positive selection from Chinese landraces to modern cultivars.Moreover,frequencies of the two favored haplotypes increased gradually with the advancement of breeding ages.Moreover,the high frequency of Hap1 and Hap2 in six major wheat production regions revealed that the selection to higher TKW was global.6.Sequence analysis of BT1 in common wheat ancestors Triticum uratu?AA?,Aegilops tauschii?DD?and tetraploid?AABB?ancestors showed that the polymorphism at BT1-6B declined obviously(?_DI/?_TE=7.18,P<0.05).The significant reduction in polymorphism at BT1-6B fell in the promoter region within 1 kb upstream of the ATG.Nucleotide polymorphism analysis of the tetraploid revealed that selection effect from the Dicoccum to the Durum wheat(?_DM/?_DR=11.9)was stronger than the domestication effect from the Dicoccoides to the Dicoccum(?_DI/?_DM=1.3).Study on hexaploid wheat demonstrated that breeding selection also caused decline of polymorphism at BT1-6B.7.In modern Chinese cultivars,the favored haplotypes of TaBT1-6B and TaAGP have certain additive effect,the more the number of favored haplotypes was,the higher the average TKW was.Either the production process of ADP-glucose catalyzed by ADP-glucose pyrophosphorylase or the transmembrane transport of ADP-glucose mediated by TaBT1 protein was weakened will affect starch synthesis,and further affect TKW.8.This study confirms that TaBT1 does have strong genetic effects on TKW.Two functional markers developed in this study could be applied to marker assisted selection?MAS?breeding.