高大山羊草中优质HMW-GS编码基因向小麦中的转移和品质效应分析

将高大山羊草中蕴含的优良HMW-GS基因1Sx2.3*和1Sy16*转入小麦，对于改良小麦品质具有重要意义。本项目拟利用携带1Sx2.3*和1Sy16*基因的小麦-高大山羊草1Sl(1B)代换系与小麦品种杂交，通过无性系变异途径诱导小麦1B染色体与高大山羊草1Sl染色体间的易位；利用分子标记和SDS-PAGE筛选1SlL•1BS纯合易位系，对纯合易位系进行GISH和FISH鉴定。同时，构建1Sx2.3*和1Sy16*基因过表达载体，将1Sx2.3*和1Sy16*基因转入小麦，获得纯合稳定转基因株系。最后，对易位系和转基因株系进行面包加工品质、相关基因表达模式和蛋白体积累趋势等分析，解析1Sx2.3*和1Sy16*基因的品质效应及作用机制，比较染色体工程和基因工程途径向小麦中导入野生种优良基因的优劣。研究结果对于培育小麦优质材料、丰富小麦优质基因资源、改良小麦面包加工品质具有理论和实践价值。

It is very important to transfer the two potential available genes 1Sx2.3* and 1Sy16* in Aegilops longissima encoding high molecular weight glutenin subunits (HMW-GS) into wheat for improving wheat bread-making quality. In this application project, a substitution line 1Sl(1B) between wheat and Ae. longissima in which wheat chromosome 1B was replaced by Ae. longissima chromosome 1Sl carrying 1Sx2.3* and 1Sy16* genes will be used to be crossed with a wheat cultivar, and translocation between chromosomes 1Sl and 1B will be induced through somatic variation by culturing the immature hybrid embryos on callus production and differentiation media in order. Comprehensively using specific markers to chromosome arms 1SlL, 1SlS, 1BL, and 1BS, respectively, homozygous translocation lines of 1SlL•1BS will be screened out in the somatic offspring. Then, the homozygous translocation lines will be identified by genomic in situ hybridization (GISH) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) for integration and expression of 1Sx2.3* and 1Sy16*. At the same time, 1Sx2.3* and 1Sy16* genes will be amplified in Ae. longissima and constructed onto expression vector, and transferred into wheat by Agrobacterium-mediated transformation. Stable transgenic wheat lines containing 1Sx2.3* and 1Sy16* genes will be achieved by molecular test. Finally, the homozygous translocation lines and stable transgenic lines will be analyzed by Farinograph, Mixograph, quantitative real-time polymerase chain reaction (qRT-PCR), and transmission electron microscope (TEM) for flour processing quality, expression of associated genes with the quality formation, and development of protein bodies to understand the effect on bread-making and functional mechanism of 1Sx2.3* and 1Sy16* genes, and to compare the advantages of the two technologies of chromosome engineering and genetic engineering in transferring potential genes into wheat from its wild relative species. The outputs obtained in this program will have theoretical and practical values for developing wheat materials with good quality, enriching gene resource for wheat quality improvement, and enhancing the bread-making property of wheat flour.