75K γ-黑麦碱基因的分子鉴定与转移  

Molecular Identifying and Transferring of 75K γ-secalin in Rye into Wheat

作　　者：王庆[1,2] 袁中伟[2] 王延谦[2] 刘登才[2] 

机构地区：[1]长江师范学院,重庆涪陵408000 [2]四川农业大学小麦研究所,四川温江611130

出　　处：《四川农业大学学报》2012年第4期373-378,共6页Journal of Sichuan Agricultural University

基　　金：国家科技计划863项目(2011AA100103);国家自然科学基金项目(30700495)

摘　　要：【目的】研究75K γ-黑麦碱基因的系统进化关系,并将秦岭黑麦75Kγ-黑麦碱基因导入普通小麦遗传背景中。【方法】利用75K γ-secalin亚基的特异引物secF/secR对5种黑麦进行扩增、克隆、测序及序列分析。以普通小麦(中国春)为母本,黑麦为父本,获得杂种。自杂种后代中,采SDS-PAGE技术分析杂交后代的75K γ-secalins蛋白亚基。用根尖制片进行体细胞染色体数目观察和原位杂交分析。【结果】序列分析结果表明,我国秦岭黑麦(AS156)与美国Wrens黑麦(CIse35)、土耳其4041黑麦(PI168199)、南非的Polko黑麦(PI412949)、智利278黑麦(PI436188)的栽培品系均具有75Kγ-黑麦碱典型的一级结构,但是在重复区中序列差异较大(包括缺失/插入和SNP)。聚类分析表明,我国的秦岭黑麦与同在北纬的土耳其和美国黑麦亲缘关系近,而与位于南纬的南非、智利的两种黑麦关系较远。以中国春为遗传背景,将位于秦岭黑麦2R染色体上的75Kγ-黑麦碱基因导入六倍体小麦,获得结实正常,农艺性状稳定的品系,编号WYQ122。通过SDS-PAGE技术和基因测序分析,已确认WYQ122中携带75Kγ-黑麦碱基因,同时原位杂交也验证黑麦染色体的存在。细胞学鉴定表明,WYQ122植株间染色体数目有41条和42条两种类型。【结论】黑麦的75Kγ-黑麦碱基因存在差异。以染色体代换系的方式将该基因转入普通小麦中。[Objective] In this study, we aimed to study the genetic phylogenetic relationship of 75K 丫-secalin gene in rye （Secale cereale L） and transfer the gene in Chinese rye line Qinling into common wheat. [Method] The experiments were contained of amplification, cloning, sequen- cing, and sequence analysis of five kinds rye using specific primers secF/secR of 75K 丫-secalin subunit. We obtained hybrids of the ordinary wheat （Chinese Spring） as female parent and the rye as male parent. We analyzed 75K丫-secalins protein subunits of hybrids by the SDS-PAGE from the progenies. We observed somatic chromosome number and analyzed in situ hybridization with apical producer. [Results] Sequence analysis showed that the cultivar had typical primary struc- ture of 75K 丫-secalin in Qinling （AS156） from China, Wrens （CIse35） from USA, 4041 （PI168199） from Turkey, Polko （PI412949） from South Africa, and 278 （PI436188） from Chile, respectively. However, they showed a lot variations, including insertion/delection and SNP in re- peat region. Moreover, the sequence from AS156 had a deletion of Cys in C-terminal. A clade wasclustered by the rye sequences from north latitude of China, USA, and Turkey and the other clade from south latitude of South Africa and Chile. The 75K 丫-secalin gene on chromosome 2R in Chi- nese rye line Qinling was transferred in common wheat line Chinese Spring. A F8 line, WYQ122, with this gene shows good fertility and consistent character in agronomy. This gene in WYQ122 was verified by SDS-PAGE analysis and sequencing data of 75K 丫-secalin gene. Cytological obser vation indicated that WYQ122 has 41 or 42 chromosomes. GISH proved this line containing rye chromosomes. [Conclusion] There was genetic diversity for 75K 丫- secalin gene among rye lines. This gene was transferred into common wheat via chromosomal substitution.

关 键 词：黑麦 75K γ-黑麦碱 分子鉴定 转移 

分 类 号：S512.1[农业科学—作物学]