【作者】 王涛；

【导师】 任正隆；

【作者基本信息】 四川农业大学 ， 生物化学与分子生物学， 2004， 博士

【摘要】 小麦的胚乳贮藏蛋白是决定其面粉烘烤品质的主要因素，谷物化学和遗传学的研究表明，限制小麦品种品质遗传潜力的最重要因素是其中的麦谷蛋白特别是高分子量的麦谷蛋白亚基(HMW-GS)，其中由Glu-D1位点编码的亚基5+10对小麦面粉烘烤品质的贡献最大。到目前为止，还没有在六倍体普通小麦中发现亚基5’+12的报道，本研究采用种间杂交技术，获得了具有新亚基5’+12的六倍体普通小麦新种质，并对其进行了品质和分子生物学的研究。进入省级区试和国家区试的小麦品系是各个育种单位新近选育的最优品系，也是今后小麦育种的重要亲本资源和生产上主要推广品种来源，为了为今后小麦育种提供优质亲本，为生产上优质小麦的推广布局提供依据，因而深入研究这些品系的遗传组成和品质特性具有十分重要的意义。本研究选取(2001-2002，2002-2003共2个年度)参加四川省区试、国家预试、国家区试的全部品系，通过高分子量谷蛋白亚基、1RS．1BL易位检测、SDS-沉降值测试，分析和比较其遗传多样性，进一步了解和掌握四川省小麦品质改良现状和发展趋势，并根据分析结果，找出四川小麦品质存在的问题，进一步提出改良策略。通过研究分析，本文得出以下结论： 1．通过种间杂交(硬粒小麦×普通小麦)，本研究获得了具有高分子量谷蛋白新亚基5’+12的小麦新种质。利用Glu-Dl位点特异引物对亚基5’+12新种质基因组DNA进行PCR扩增，结合SDS-PAGE对该种质进行了鉴定，发现新亚基5’+12中的5’亚基与5+10中的5亚基具有相同的电泳迁移率。但是采用不同的Dxs基因特异引物进行扩增时，发现5’+12中的5’亚基基因与5+lO中的5亚基基因在重复区起始处存在有差异，这表明本研究中获得的HMW一GSS’+l2是一个新的亚基组合，其中的Dxs’基因是一个与Dxs基因相似的新基因。亚基5’+l2在杂种一代呈现共显性遗传，在杂种后代的遗传规律因亲本类型不同而不同。通过对单个亚基在整个HMW一GS中的相对含量进行分析，结果表明新亚基5’+l2中y型亚基12的相对含量明显高于亚基5+10中y型亚基10的相对含量。亚基5’+l2新种质的沉降值、湿面筋含量、粉质图形成时间、稳定时间及评价值等品质指标显著优于其具有优质亚基5+10的姊妹系，表明5’+l2亚基具有增加面筋强度，提高面筋质量的作用，初步认为本研究中获得的新亚基组合5’+l2优于目前国际公认的优质亚基5+lO，具有罕见亚基5’+l2的新种质在遗传上已经纯合，且农艺性状稳定，亚基5，+12小麦新种质的创造，将为小麦品质育种提供新的优质遗传资源。2.SDS一PAGE分析表明，四川小麦区试中的参试品系具有较丰富的HMW-GS类型和亚基组合类型，Glu一BI位点亚基13+16、14+15、6+8等是近几年四川小麦中新出现的亚基，Glu一Dl位点5+lO亚基的频率己经超过2+l2的频率，表明近几年四川小麦育种将优质亚基5+l0导入Glu一Dl位点，从而改良该位点的亚基组成取得了较好的效果。本研究表明四川小麦育种对来自地方品种的N，7十8，2+12的人工改良取得了明显的成效，四川省的小麦育种思路发生了明显的变化，越来越重视优质亚基的选择，四川小麦品质育种有了较大进展。本研究认为还需要进一步改良四川小麦Glu一Al、Glu一Dl亚基组成，在Glu一Al位点导入亚基l或者2*，在Glu一Dl位点增加优质亚基5+l0等的频率。3.A一PAGE分析结果表明，四川小麦中具有极高的的IRS.IBL频率(两年四川省区试的IRS.IBL易位系的平均频率为69.7%，两年国家区预试的IRS.IBL易位系的平均频率为735%)。本研究发现具有IRs.IBL参试材料的SDS一沉降值达为53.Oml，比不含IRS.IBL参试材料的SDS一沉降值(平均57.9ml)低9.2%，表明IRS.IBL对小麦品质具有较大的副效应，该易位对在小麦育种过程中选育高品质小麦是不利的，四川小麦中存在高频率的IRS.IBL易位问题应该引起高度重视。认为可通过以下途径改良四川小麦中IRS.IBL易位的品质副效应问题:通过改善IRS.IBL易位系的遗传背景，可消除IRS的品质负效应;通过增加高分子或低分子量谷蛋白亚基的含量来增加谷蛋白聚合体的大小和数量，从而在一定程度补偿由IRS.IBL导致的Glu一B3位点低分子量谷蛋白亚基的缺失，那么IRS.IBL小麦的品质就能提高;通过利用不含Sec一1位点的新IRS.IBL易位，既能利用黑麦IRS上的有益基因，又能避免See一1位点的品质副效应。更多还原

【Abstract】 The endosperm storage proteins of bread wheat are the major determinants of baking quality of flour. Biochemical and genetic studies have revealed that the wheat glutenin, particularly, HMW-GS(High Molecular Weight Glutenin Subunit), are the major limiting factor for breeding potential of various varieties. Among the various HMW-GS, the 5+10 encoded at the Glu-Dl locus contributes the most to the high baking quality of wheat flour. To our knowledge, there have no reports for the presence of the 5’+12 HMW-GS in hexaploid wheat. A novel wheat line having a new HMW-GS 5’+12 was developed using inter-specific cross in this study. The molecular characterization of this new HMW-GS5’+12 was conducted, and the quality of flour from this novel wheat line was determined. In addition, Wheat lines chosen for provincial and national field tests are some of the best ones newly bred by various institutes, and are important germplasm for breeding and excellent candidates to be promoted for large-scale production. It is, thus, vital to study their genetic characteristics related to grain quality. This study characterized the genetic composition and diversity of HMW-GS, and their impact on grain quality in all wheat from the tests in Sichuan province, national initial and final tests, using techniques such as SDS-PAGE, SDS-sedimentation test, identification of 1RS.1BL translocation. Through these analyses, deficiencies in grain quality related to HMW-GS were identified in various wheat varieties,particularly in those grown in Sichuan province. Strategies for quality improvement in various varieties were proposed. Based on the results from this study, the following conclusions were drawn.1. A novel wheat line possessing new high-molecular-weight glutenin subunits(HMW-GS) 5’+12 was developed by using interspecific cross (T.durum Desf XT.aestivum L.) in this study. The SDS-PAGE analyses demonstrated that this new germplasm possessing HMW-GS 5 and 12 at Glu-Dl loci. PCR based assays were also used to identify the Glu-Dl locus, there are small difference of DNA sequences was found between the Dx5’ gene in HMW-GS 5’+12 and the Dx5 gene in HMW-GS 5+10 by using defferent specific PCR primers to Dx5 gene, indicating the Dx5’ was a new gene at Glu-Dl locus and the HMW-GS5’+12 is a new combination which have never been report. The inheritance of HMW glutenin subunits 5’+12 in F1 is consistent with the co-dominant type, and the segregation regularity in F2 differing from the parent, subunits 5+12, 2+12 and 2+5+12 have the same frequency in F2 when the cross between subunits 5’+12 and 2+12, however subunits 5+12 have higher frequency than 5+10 and 5+10+12 in F2 when the cross between subunits 5’+12 and 5+10.The relative content of mono-subunits in total HMW glutenin subunit was analyzed by using computer, it was found that the relative content of y-type subunit 12 in subunit combination 5’+12 was higher than y-type subunit 10 in subunit combination 5+10. There are various studies showing that HMW glutenin subunits 5+10 is associated with the best bread-making quality. However, Lines possessing HMW-GS 5’+12 subunits were superior in many quality parameters, sedimentation value, wet gluten content, development time and stability time of farinograph as compared to their siblines containing HMW-GS 5+10, This indicate the HMW-GS 5’+12 can result in stronger gluten and improve the gluten quality. This novel line also has high yielding characters in multi-florets, and can be an extremely useful genetic resources for breeding wheat varieties with good grain quality.2. There are rich diversity of HMW-GS and their combinations in wheat varieties chosen from Sichuan provincial tests. The HMW-GS at Glu-Bl locus, 13+16,514+15 and 6+8, were among the novel ones that were introduced into varieties in Sichuan province in recent years. The occurrence frequency of the HMW-GS of 5+10 at Glu-Dl locus in these wheat varieties has exceeded that of the 2+12 HMW-GS, indicating that the superior 5+10 have been bred更多还原