The Function Of TaSBEⅠ Gene On Starch Synthesis In Grains Of Wheat(Triticum Aestivum L.)

Starch,the most important component in wheat grains,not only determines the grain weight,but also has a close relationship with the processing quality of cooked flour products.Our laboratory previously found that TaSBEI,an amylopectin synthesis gene,which was significantly induced during the development of wheat grains,suggesting its important role in starch synthesis.Therefore,the overexpression vector of TaSBE Ⅰ gene was constructed and transferred into wheat cultivar Zhengmai 7698,which is one of the main varieties planted in Henan Province.In this study,homozygous transgenic plants were screened by the detection of different progenies(T2～T4)of TaSBE Ⅰ transgenic lines.Furthermore,the yield,starch content and other main agronomic traits of T4 generation homozygous TaSBE Ⅰ gene transgenic wheat plants were investigated.It was found that overexpression of TaSBE Ⅰ gene significantly increased wheat yield and amylopectin contents,and improved starch properties.At the same time,amylopectin fine structure,such as chain length distribution and branching degree changed significantly.These results indicated that TaSBE Ⅰ gene can affect the amylopectin structure of wheat,and improve starch quality and increase wheat yield.In addition,TaSBE Ⅰ gene was re-sequenced in 355 varieties to identify the favored haplotype of TaSBE Ⅰ gene,which provided a theoretical basis for high TKW and superior starch wheat breeding.The main results are as follows:(1)The homozygous transgenic wheat lines with overexpressing TaSBE Ⅰ were obtained.PCR specific detection method was used to detect and screen positive transgenic plants and Bar free wheat plants of TaSBE Ⅰ gene in T2-T4 generation plants.The results showed the proportion of positive grains without screening marker genes in T2-T4 generation was 5.6%,20.2%and 62.7%,respectively,and the separation ratio increased year by year.In addition,semi quantitative and RT-PCR were used to detect the expression level of TaSBE Ⅰ gene in three homozygous transgenic lines of T4 generation at RNA level.The results showed that the expression level of TaSBE Ⅰ gene in homozygous transgenic lines was significantly higher than that of wild type at 10 and 25 days after anthesis.It indicates that the homozygous overexpression of TaSBE Ⅰgene transgenic plants without screening markers were obtained in this study,which is beneficial to reduce the risk of transgenic wheat.(2)The main agronomic characters and starch properties of homozygous transgenic wheat lines with overexpressed TaSBE Ⅰ were determined.Compared to wild type,the thousand grains weight of homozygous TaSBE Ⅰ transgenic wheat plants were all significantly higher;while there was no significant difference in plant height,average number of panicles per plant and average number of spikelets per ear.The results for determination of starch content and gelatinization characteristics,showed that overexpression TaSBE Ⅰ gene increased the total starch and amylopectin contents,decreased amylose contents and ratio of amylose contents to amylopectin contents,and significantly increased the peak viscosity,breakdown,hold through,setback,pasting time and swelling power of starch.The results showed that overexpression of TaSBE Ⅰwas beneficial to increase grain weight and improve starch quality.(3)Cytological changes in grains of homozygous transgenic wheat plants with overexpressing TaSBE Ⅰ were observed.The starch granules of TaSBE Ⅰ transgenic wheat and wild type were scanned by scanning electron microscope(SEM).It was found that the number of starch granules of B-type in transgenic plants was significantly more than that of wild type.The results of the determination of number,volume and surface area of starch granules showed that the number of starch granules distribution was a single-peak curve,and the volume and surface area distribution showed a doublepeak curve.While,the number,volume and surface area of A-type starch grains in transgenic lines were less than those in wild type;but the number of B-type starch granules was higher than that of wild type,especially the small starch granules(<0.5μm);and the volume and surface area of B-type starch granules were also higher than that of wild type.It showed that the improvement of starch quality of homozygous transgenic plants with overexpressing TaSBE Ⅰ closely related to the increase of Btype starch grains(4)The fine structure of starch in grains of homozygous transgenic wheat plants with overexpressing TaSBE Ⅰ detected.It was found that the general trend of chain length distribution of amylopectin in transgenic lines was consistent with that of wild type,with the highest chain length at DP11,and most of the chain length distributions were clustered in DP 11-24.Compared with the wild type,the branching degree(2.332.65)and proportion of short chains(6 ≤DP≤10)of transgenic lines were significantly increased,while the proportion of medium chain and long chain(DP≥11)decreased.The results showed that the improvement of starch quality of transgenic plants was due to the increase of(6 ≤DP ≤10)short chains.(5)In order to identify the allelic loci associated with wheat grain weight and superior starch quality and to develop related molecular markers,this study was conducted by resequencing wheat varieties.And the results showed that TaSBE I-7A could be divided into three haplotypes,and the average TKW of Hap2 and Hap3 was higher than Hap 1;TaSBEI-7B could be divided into two haplotypes,the TKW of Hap1 was higher than that of Hap2;and TaSBE I-7D had no SNP.