Liu Huiyun, Wang Ke, Xiao Lele, Wang Shunli, Du Lipu, Cao Xinyou, Zhang Xiaoxiang, Zhou Yang, Yan Yueming, Ye Xingguo
Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Key Laboratory of Genetics and Biotechnology, College of Life Science, Capital Normal University, Beijing, 100048, China.
PLoS One. 2016 Jan 14;11(1):e0146933. doi: 10.1371/journal.pone.0146933. eCollection 2016.
High molecular weight glutenin subunits (HMW-GSs) are important seed storage proteins in wheat (Triticum aestivum) that determine wheat dough elasticity and processing quality. Clarification of the defined effectiveness of HMW-GSs is very important to breeding efforts aimed at improving wheat quality. To date, there have no report on the expression silencing and quality effects of 1Bx20 and 1By20 at the Glu-B1 locus in wheat. A wheat somatic variation line, AS208, in which both 1Bx20 and 1By20 at Glu-B1 locus were silenced, was developed recently in our laboratory. Evaluation of agronomic traits and seed storage proteins by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and reversed-phase high performance liquid chromatography (RP-HPLC) indicated that AS208 was highly similar to its parental cultivar Lunxuan987 (LX987), with the exception that the composition and expression of HMW-GSs was altered. The 1Bx20 and 1By20 in AS208 were further identified to be missing by polymerase chain reaction (PCR) and quantitative real-time RT-PCR (qRT-PCR) assays. Based on the PCR results for HMW-GS genes and their promoters in AS208 compared with LX987, 1Bx20 and 1By20 were speculated to be deleted in AS208 during in vitro culture. Quality analysis of this line with Mixograph, Farinograph, and Extensograph instruments, as well as analysis of bread-making quality traits, demonstrated that the lack of the genes encoding 1Bx20 and 1By20 caused various negative effects on dough processing and bread-making quality traits, including falling number, dough stability time, mixing tolerance index, crude protein values, wet gluten content, bread size, and internal cell structure. AS208 can potentially be used in the functional dissection of other HMW-GSs as a plant material with desirable genetic background, and in biscuit making industry as a high-quality weak gluten wheat source.
高分子量谷蛋白亚基（HMW - GSs）是小麦（普通小麦）中重要的种子贮藏蛋白，决定着小麦面团的弹性和加工品质。明确HMW - GSs的特定功效对于旨在提高小麦品质的育种工作非常重要。迄今为止，尚无关于小麦Glu - B1位点上1Bx20和1By20的表达沉默及品质效应的报道。最近我们实验室培育出了一个小麦体细胞变异系AS208，其中Glu - B1位点上的1Bx20和1By20均被沉默。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS - PAGE）和反相高效液相色谱（RP - HPLC）对农艺性状和种子贮藏蛋白进行评估，结果表明AS208与其亲本品种轮选987（LX987）高度相似，只是HMW - GSs的组成和表达发生了改变。通过聚合酶链反应（PCR）和定量实时逆转录PCR（qRT - PCR）分析进一步确定AS208中缺失了1Bx20和1By20。基于AS208与LX987中HMW - GS基因及其启动子的PCR结果推测，AS208在离体培养过程中1Bx20和1By20被缺失。使用揉混仪、粉质仪和拉伸仪对该品系进行品质分析以及面包制作品质性状分析，结果表明编码1Bx20和1By20的基因缺失对面团加工和面包制作品质性状产生了各种负面影响，包括降落数值、面团稳定时间、混合耐受性指数、粗蛋白值、湿面筋含量、面包体积和内部细胞结构。AS208作为具有理想遗传背景的植物材料，有可能用于其他HMW - GSs的功能解析，并且在饼干制作行业中作为优质弱筋小麦来源。
