利用HRM功能标记检测黑龙江省水稻种质资源Wx和fgr的基因型分布

水稻直链淀粉含量与香味作为水稻重要的品质性状,一直是水稻研究热点。HRM（高分辨率熔解曲线）技术作为近年来新兴的PCR分析技术,拥有简便、快速及分辨率高等特点,备受关注。为明确黑龙江省种质资源中Wx及fgr基因的分布情况,利用HRM功能标记Wx-a/b和fgr-E7FNP,在67份部分黑龙江省主栽品种及优异种质资源中进行基因分型,结果表明,67份种质均含有纯合Wx^b基因;4份种质含有纯合的fgr基因,分别为哈99352、龙粳香1号、中龙香粳1号及龙交103970,其余63份种质含纯合野生型BAD2基因。     

陕西省水稻Wx基因种质资源遗传多样性分析

稻米品质与稻米中直链淀粉含量(AC)有关,而直链淀粉含量与蜡质基因(Wx)的基因型有关,研究表明Wx基因第一内含子的剪接效率决定直链淀粉含量。明确陕西省主要水稻种质资源中Wx基因的基因型,可为水稻品质育种提供依据。应用CAPS法对112份陕西省水稻主要种质资源Wx基因第一内含子供体+1位碱基G/T进行检测,根据G/T碱基将112份供试材料的Wx基因分为GG、GT、TT等3种基因型。同时,按国标检测3种不同G/T型37份稻米的直链淀粉含量及其他相关性状。结果显示,112份供试材料中有1份材料为GG型,约占1%;25份为GT型,占22%;86份材料为TT型,约占77%。说明Wx基因第一内含子供体+1位碱基G/T的多态性与AC含量有良好的对应关系,与垩白粒率和垩白度有一定的对应关系。     

基于HRM体系的水稻不育系香味和抗稻瘟病基因分型研究

为了解析现有不育系香味及稻瘟病抗性基因背景,本文通过基于高分辨率熔解曲线分析系统(HRM)的水稻香味和抗稻瘟病基因功能型分子标记对四川省近年生产上广泛应用的或新育成的46份三系杂交水稻不育系进行fgr、Pita及Pi-k基因检测及分型。表明具有纯合fgr香味基因型的不育系有10份,分别具有显性纯合抗稻瘟病Pita、Pi-k基因型的不育系有9和29份,同时含有Pita和Pi-k基因型的5份;同时具有fgr、Pita、Pi-k基因型的只有2份。其结果对指导四川水稻香味品质和抗稻瘟病分子育种具有重要作用。     

杂交水稻骨干亲本Wx基因第一内含子+1位碱基多态性分析

【目的】直链淀粉含量是影响稻米品质的重要因素,而编码颗粒结合型淀粉合成酶(granule-bound starch synthetase,GBSS)的蜡质基因(Wx)是影响直链淀粉含量的主效基因。本研究拟分析120份杂交水稻亲本的Wx基因第一内含子+1位碱基的多态性,以期研究其对内含子的剪接效率及Wx的表达的影响,为亲本育种筛选提供理论参考。【方法】选取120份水稻亲本材料,采用CTAB法提取基因组DNA,并以此为模版PCR扩增Wx基因片段;采用限制性内切酶Acc I对PCR产物进行酶切分析,根据琼脂糖电泳结果分析Wx基因第一内含子+1位碱基类型;同时测定其中19份亲本的直链淀粉含量、胶稠度和碱消值。【结果】PCR扩增结果显示所有材料中均可扩增出清晰且单一的目的条带;PCR产物的Acc I酶切分析结果显示,其中81份亲本的Wx基因第一内含子+1位碱基是T,即为T型,占总数的67.5%,38份亲本为G型,占总数的31.67%,而仅有1份亲本为杂合型,即GT型,占总数的0.83%。19份亲本的直链淀粉含量测定结果表明,7份G型亲本的直链淀粉含量比12份T型亲本的高,且其中6个G型亲本的直链淀粉含量平均值均超过20%。另外,测定结果表明T型亲本和G型亲本的胶稠度和碱消值差别不明显。【结论】120份杂交水稻骨干亲本的Wx基因第一内含子+1位碱基类型测定结果表明,与T型亲本相比,G型亲本的直链淀粉含量明显较高,而胶稠度和碱消值没有明显差别。     

应用CRISPR/Cas9基因编辑技术获得高直链淀粉水稻种质

【目的】通过CRISPR/Cas9基因编辑技术遗传改良水稻种质,创制高直链淀粉新材料。【方法】以水稻品种中花11为试验材料,利用CRISPR/Cas9编辑系统对水稻淀粉分支酶（Starch branching enzymes, SBE）基因OsSBE3进行靶向编辑,利用PCR技术鉴定无标记纯合突变体,并测定其淀粉含量。【结果】T0代获得10株突变体株系,T1代获得5个无标记纯合突变株系,其中4个株系（sbe3-22-6、sbe3-25-3、sbe3-25-4、sbe3-25-6）的直链淀粉含量和淀粉直支比显著高于野生型。【结论】本研究创制了高直链淀粉含量的水稻新种质,为水稻品质改良提供了参考。     

利用蜡质基因Wx改良稻米品质的研究进展

品质优良的稻米往往含有较低的直链淀粉,其含量决定于蜡质基因(Waxy,Wx)。目前Wx基因的多个等位变异已经被鉴定,并且应用到水稻的分子育种中。本文综述了Wx基因的等位变异类型,分子标记及其在分子育种中的应用,希望为选育低直链淀粉的优质水稻品种提供参考。     

利用Wx基因分子标记辅助选择技术培育中等直链淀粉含量的恢复系

为培育中等直链淀粉含量的水稻恢复系,以HC086为供体亲本(其Wx基因来自美国水稻品种Francis),以优良的水稻恢复系R898、R476、R838、R6547为受体亲本,利用PCR-Accl分子标记检测技术在回交世代进行辅助选择,将供体材料控制直链淀粉的Wx基因导入到不同恢复系中.对后代株系的基因型及其直链淀粉含量的分析结果表明,直链淀粉含量普遍有了显著提高,受体亲本的直链淀粉含量由10.6%～14.4%提高到了18.0%～21.4%.改良株系与广占63S和Y58S的配组实验表明,F1代杂交种的直链淀粉含量较原组合有显著提高,并达到适中直链淀粉含量水平,且主要农艺性状没有发生显著改变.说明分子标记辅助选择是改良水稻品种直链淀粉含量的快速有效的方法.     

反义Wx基因导入我国常规籼稻品种的研究

经农杆菌介导，将自行克隆并构建的反义Wx基因导入4个高直链淀粉含量的常规籼稻品种绿黄占、清芦占11号、三芦占7号和特青中，经潮霉素抗性筛选获得抗性植株。聚合酶链反应(PCR)和Southern杂交检测证明反义Wx基因已整合进转基因水稻植株的基因组中。对成熟种子直链淀粉含量的分析表明，部分转基因水稻植株T1或T2代种子中的直链淀粉含量有不同程度的下降，最低的已降至16．52％，较对照下降了40．5％；在此基础上筛选获得了部分转基因纯合株系。研究结果还表明，盲链淀粉含量的改变会导致相应稻米的胶稠度和糊化温度的变化。     

软米直链淀粉含量的遗传分析

利用引物484／485和CAPs标记PCR-AccI对软米直链淀粉含量进行DNA分子标记检测表明,软米直链淀粉含量的遗传受一个与Wx基因非等位的基因控制。遗传分析表明,软米低直链淀粉含量对高直链淀粉含量为隐性,表现为单基因遗传。为软米直链淀粉含量基因的定位以及利用奠定了基础。     

2个低直链淀粉含量籼稻突变体的遗传分析

对2个空间诱变低直链淀粉含量籼稻突变体XLA-1和XLA-2进行了遗传分析和分子生物学研究.结果表明:XLA-1低直链淀粉遗传特性受2对隐性基因控制,这2对基因具有连锁关系和互补作用,任何1对基因隐性纯合都将导致直链淀粉含量降低;XLA-2低直链淀粉遗传特性受1对隐性主效基因控制,该基因可能为Wx的等位基因,同时受微效基因的修饰.XLA-1和XLA-2蜡质基因(CT)n微卫星多态性与糯稻相同,但其直链淀粉质量分数分别为14.42%和11.59%,说明除Wx基因外确实还有其他影响直链淀粉含量的遗传因素,这与遗传分析结果相符.     

Differential expressions among five Waxy alleles and their effects on the eating and cooking qualities in specialty rice cultivars

Eating and cooking qualities(ECQs) of rice are important attributes due to its major influence on consumer acceptability. To better understand the molecular mechanism of the variation in ECQs, we investigated and compared the expressions among different alleles of the Waxy(Wx) gene and its effect on ECQs in specialty rice cultivars. The results showed that the accumulation of amylose was positively and significantly correlated to the level of mature Wx m RNA and granule-bound starch synthase I(GBSS I) in developing rice grain at 12 days after flowering. The amount of GBSS I and its activity together are the main factors controlling amylose synthesis. Differences in ECQs among five Wx allele types were investigated in samples from 15 rice varieties. The apparent amylose content(AAC) and gel consistency(GC) were similar in each type of Wx allele. The AAC followed the order, Wxa typeWxin typeWxb typeWxmq typewx. Contrary to this, the GC showed an opposite trend compared to AAC. There was a wide variation in rapid visco analyzer(RVA) profile among five Wx allele types, while varieties sharing a specified Wx allele had basically the similar RVA profile, although there was a slight difference in some RVA parameters, peak, hot paste and cool paste viscosities.     

稻米品质的遗传研究及分子育种进展

水稻是我国最主要的粮食作物之一,高产优质不仅是水稻基础研究的重点,更是水稻育种应用的主要方向.稻米品质主要包含加工碾磨品质、外观品质、蒸煮食味品质和营养品质,是受遗传与环境因素共同影响的复杂性状.本文对稻米品质性状的分类、影响因素、遗传研究进展和稻米品质育种改良现状进行了综述,并对稻米品质研究进行总结和展望,以期为稻米...     

水稻不育系槟榔红A的选育

【目的】为了增加特种稻种质资源,解决白米不育系配组红香米杂交稻F1易混杂白米,影响米质的问题。【方法】采用常规育种方法,将低直链淀粉含量的亲本材料与带香味的亲本材料进行杂交、回交、转育等。【结果】育成直链淀粉含量为11．07％、香味明显、食用口感佳的水稻三系不育系槟榔红A,为红香米三系杂交稻的配组应用提供了新的种质资源。     

水稻品种直链淀粉的积累特性与W_x蛋白的关系

本文研究了中国水稻品种直链淀粉的积累特性及其与W_x蛋白的关系。结果表明:非糯品种直链淀粉的含量随籽粒的发育而增加;糯性品种花后7～14天直链淀粉含量上升,以后渐渐降低。糯性品种缺乏W_x蛋白;非糯品种的W_x蛋白水平与直链淀粉含量呈很好的线性关系,W_x蛋白的出现和变化情况则因直链淀粉含量的不同而异,其变化主要是受W_x基因顺序表达的影响。     

水稻不育系槟榔红A的选育

【目的】为了增加特种稻种质资源,解决白米不育系配组红香米杂交稻F1易混杂白米,影响米质的问题。【方法】采用常规育种方法,将低直链淀粉含量的亲本材料与带香味的亲本材料进行杂交、回交、转育等。【结果】育成直链淀粉含量为11.07%、香味明显、食用口感佳的水稻三系不育系槟榔红A,为红香米三系杂交稻的配组应用提供了新的种质资源。     

Determination of Amylose Content and Its Relationship with RVA Profile Within Genetically Similar Cultivars of Rice(Oryza sativa L.ssp.japonica)

Anther culture pure lines with antisense Wx gene were generated by Agrobacterium tumefaciens-mediated co-transformation followed by anther culture in transgenic rice(Oryza sativa L.ssp.japonica).The antisense Wx transgenic pure lines were used to analyze the differences and the relationship between RVA eigenvalues among the lines with different amylose contents.77 pure lines of anther culture having antisense Wx gene were studied for the amylose content of its offspring cultivar,Wuyunjing 7.It was found that levels of amylose content were similar to the control in 19 lines(16.0%);1-5% lower than control in 50 lines,of which,within 11.0-13.9% in 40 lines;and 3.1-4.0% in 8 lines.The effect of the amylose content on RVA eigenvalue was analyzed through the RVA profile graphic comparison and test of significance for RVA eigenvalues.The analysis of RVA profiles with different amylose contents indicated that there were two kinds of RVA profiles within genetically similar cultivars.The lines with 3.1-4.0% of amylose content were distinctly different from other lines and conventional glutinous rice.The comparison of the similarity curve of RVA profile of the lines with different amylose contents showed that the lines with lower amylose content had a distinguished RVA profile with the curve increasing gradually,but not exceeding the first apex.The test of significance for eigenvalues of the RVA profile in the lines with different amylose contents indicated that there were five eigenvalues remarkably different among the lines with 3.1-4.0% of amylose content compared with other two groups.Further,there were three eigenvalues remarkably different among the lines with 11.0-13.5% of amylose content compared with control group.An optimized mathematical model,which characterizes the relationships between the amylose content and RVA eigenvalue,has been established in this study.From this model,it was clearly indicated that the parameters PeT and SBV were more related to amylose content.It was concluded that the differences in amylose content not only affected the eigenvalues of RVA but also caused different RVA profiles within genetically similar cultivars.The introduction of antisense Wx gene could lead to reduce the amylose content and lay theoretical foundation for quality improvement in rice breeding programs.     

Determination of Amylose Content and Its Relationship with RVA Profile Within Genetically Similar Cultivars of Rice (Oryza sativa L.ssp.japonica)

Anther culture pure lines with antisense Wx gene were generated by Agrobacterium tumefaciens-mediated co-transformation followed by anther culture in transgenic rice (Oryza sativa L.ssp.japonica).The antisense Wx transgenic pure lines were used to analyze the differences and the relationship between RVA eigenvalues among the lines with different amylose contents.77 pure lines of anther culture having antisense Wx gene were studied for the amylose content of its offspring cultivar,Wuyunjing 7.It was found that levels of amylose content were similar to the control in 19 lines (16.0%);1-5%lower than control in 50 lines,of which,within 11.0-13.9% in 40 lines;and 3.1-4.0% in 8 lines.The effect of the amylose content on RVA eigenvalue was analyzed through the RVA profile graphic comparison and test of significance for RVA eigenvalues.The analysis of RVA profiles with different amylose contents indicated that there were two kinds of RVA profiles within genetically similar cultivars.The lines with 3.1-4.0% of amylose content were distinctly different from other lines and conventional glutinous rice.The comparison of the similarity curve of RVA profile of the lines with different amylose contents showed that the lines with lower amylose content had a distinguished RVA profile with the curve increasing gradually,but not exceeding the first apex.The test of significance for eigenvalues of the RVA profile in the lines with different amylose contents indicated that there were five eigenvalues remarkably different among the lines with 3.1-4.0% of amylose content compared with other two groups.Further,there were three eigenvalues remarkably different among the lines with 11.0-13.5% of amylose content compared with control group.An optimized mathematical model,which characterizes the relationships between the amylose content and RVA eigenvalue,has been established in this study.From this model,it was clearly indicated that the parameters PeT and SBV were more related to amylose content.It was concluded that the differences in amylose content not only affected the eigenvalues of RVA but also caused different RVA profiles within genetically similar cultivars.The introduction of antisense Wx gene could lead to reduce the amylose content and lay theoretical foundation for quality improvement in rice breeding programs.     

湖南稻瘟病菌生理小种的组成及其致病性

利用来自日本、国际水稻研究所(IRRI)和中国的10个稻瘟病菌单基因鉴别品种,将59个湖南稻瘟病菌株分别划分为20个生理小种,优势小种为H531.1、H511.1和H501.1,对抗性基因Pi–CO39、Pita、Pi3、Pi12、Pia具有致病性。各个稻区生理小种组成和优势小种存在差异,其中,湘东、湘中稻区稻瘟病菌生理小种多样性指数较高,优势小种的变化最为明显,说明这2个稻区的抗源基因正逐渐变得匮乏,甚至丧失。鉴别品种C101LAC(Pi1)和C101A51(Pi2)对59个湖南稻瘟病菌菌株抗性频率均为100.0%,说明这些抗性品种(基因)可作为湖南稻区稻瘟病的抗源材料(基因),Pi1、Pi2可直接作为分子标记辅助选择育种的供体抗源基因。品种关东51(Pik)、C105TTP–4L23(Pi4–b)和C101PKT(Pita)在湖南各稻区的抗性水平表现存在差异,可以作为选择性抗源材料(基因)加以应用。