Transferring Translucent Endosperm Mutant Gene Wx-mq and Rice Stripe Disease Resistance Gene Stv-bi by Marker-Assisted Selection in Rice (Oryza sativa)

A high-yielding japonica rice variety, Wuyunjing 7, bred in Jiangsu Province, China as a female parent was crossed with a Japanese rice variety Kantou 194, which carries a rice stripe disease resistance gene Stv-bi and a translucent endosperm mutant gene Wx-mq. From F2 generations, a sequence characterized amplified region (SCAR) marker tightly linked with Stv-bi and a cleaved amplified polymorphic sequence (CAPS) marker for Wx-mq were used for marker-assisted selection. Finally, a new japonica rice line, N...     

水稻Wx基因的遗传多态性及其与主要米质指标的相关性分析

利用RM190和484/W2R ACCⅠ标记分析了50个水稻非糯品种的Wx基因的等位性变异。其中,RM190揭示出7种(CT)n纯合变异类型和1种杂合类型,分别为 (CT)20、(CT)19、(CT)18、(CT)17、(CT)14、(CT)11、(CT)10和(CT)11/(CT)18,并以（CT）11和（CT）18两种类型为主;484/W2R ACCⅠ共揭示出2种纯合类型和1 种杂合类型,分别为G/G型、T/T型及G/T型;(CT)11大多为G/G型,(CT)18大多为T/T型,这两个标记可将供试材料划分为10种等位基因变异类型。进一步分析表明,RM190揭示的Wx基因多态性可以解释直链淀粉含量变异的59.3 %;484/W2R ACCⅠ揭示的Wx基因多态性可以分别解释直链淀粉含量及胶稠度变异的56.1%和24.6%;而两标记共同可解释直链淀粉含量变异的72.4%。还对这两个标记在育种实践中的应用前景进行了探讨。     

Transferring Translucent Endosperm Mutant Gene Wx-mq and Rice Stripe Disease Resistance Gene Stv-b by Marker-Assisted Selection in Rice (Oryza sativa)

A high-yielding japonica rice variety, Wuyunjing 7, bred in Jiangsu Province, China as a female parent was crossed with a Japanese rice variety Kantou 194, which carries a rice stripe disease resistance gene Stv-bⁱ and a translucent endosperm mutant gene Wx-mq. From F₂ generations, a sequence characterized amplified region (SCAR) marker tightly linked with Stv-bⁱ and a cleaved amplified polymorphic sequence (CAPS) marker for Wx-mq were used for marker-assisted selection. Finally, a new japonica rice line, Ning 9108, with excellent agronomic traits was obtained by multi-generational selection on stripe disease resistance and endosperm appearance. The utilization of the markers from genes related to rice quality and disease resistance was helpful not only for establishing a marker-assisted selection system of high-quality and disease resistance for rice but also for providing important intermediate materials and rapid selection method for good quality, disease resistance and high yield in rice breeding.     

分子标记辅助选择聚合水稻暗胚乳突变基因Wx-mq和抗条纹叶枯病基因Stv-b~i

以江苏高产粳稻品种武运粳7号为母本,具有条纹叶枯病抗性基因Stv-bi和暗胚乳突变基因Wx-mq的日本粳稻品种关东194作父本配制杂交组合进行聚合育种。利用与Stv-bi基因共分离的SCAR标记及与暗胚乳突变基因Wx-mq紧密连锁的CAPS功能标记对其分离世代进行标记位点的检测,结合田间多代选育、抗性鉴定和籽粒胚乳外观鉴定,将条纹叶枯病抗性基因Stv-bi和暗胚乳突变基因Wx-mq同时转育到高产品种中,筛选、培育出优质、抗病、高产且农艺性状优良的水稻新品系宁9108。利用与水稻优质及抗病基因紧密连锁的分子标记,不仅初步建立了优质、抗病育种的分子标记辅助选择体系,也为水稻优质、抗病、高产育种提供了一种简单、快捷的选择方法和重要的中间材料。     

Genetic Polymorphism of Wx Gene and Its Correlation with Main Grain Quality Characteristics in Rice

The allelic variation of the Wx gene in 50 non-glutinous rice varieties (lines) was analyzed by using the microsatellite marker RM190 [for (CT)n simple sequence repeat (SSR)] and cleaved amplified polymorphic sequence(CAPS) marker 484/W2R-ACCⅠ[for G/T single nucleotide polymorphism (SNP)]. Six homozygous (CT)n types, namely (CT)20, (CT)19, (CT)18, (CT)17, (CT)16, (CT)14, (CT)11 and (CT)10, and a heterozygous genotype (CT)11/(CT)18 were detected for RM190, of which (CT)11 and (CT)18 were predominant. Two homozygous Wx genotypes (G/G and T/T) and one heterozygous (G/T) were detected using 484/W2R-ACCⅠ . Most of the materials with a RM190 of (CT)11 were G/G for SNP of 484/W2R-ACC I, while T/T for SNP was predominantly appeared in materials with (CT)18. The materials tested could be grouped into 10 categories using the two markers together. Results indicated that 59.3% variance of amylose content was attributed to the polymorphism of Wx gene revealed by RM190, while 56.1% and 24.6% of the variances in amylose content and gel consistency were respectively to the polymorphism of Wx gene revealed by 484/W2R-ACC I. Furthermore, with both SSR and CAPS markers, 72.4% of the variance in amylose content could be explained. In addition, the application prospects of the two markers in breeding were also discussed.     

同源四倍体和二倍体水稻Wx基因与淀粉品质的遗传关系

用Wx 基因的(CT)n微卫星标记和切割的扩增产物多态性序列(cleaved amplified polymorphic sequence, CAPS)分子标记PCR Acc Ⅰ 对40份同源四倍体和14份二倍体水稻Wx基因型进行研究。利用484/485引物检测,Wx基因呈 Wx1、Wx2和Wx3 3种多态性;而用PCR Acc Ⅰ检测,Wx基因表现为G 型和T 型。同时,在测定稻米直链淀粉含量（AC）、胶稠度（GC）和糊化温度（GT）的基础上探讨它们与Wx基因的关系,结果表明AC、GC与Wx基因密切相关,GT与Wx基因相关不显著。综合分析表明,同源四倍体水稻中,92.0%的Wx1基因型材料为G 型,而Wx2基因型村料和800%的Wx3基因型材料为T 型。统计分析表明,Wx基因(CT)n微卫星标记检测与PCR Acc Ⅰ 分子标记检测的相关系数为0842,呈极显著正相关。     

通过分子标记辅助选择培育优良食味水稻新品种

 为了培育食味品质优良、综合丰产性好的粳稻新品种,以高产粳稻品种武香粳14作母本,具有暗胚乳突变基因的优质粳稻品种关东194作父本配制杂交组合。利用与暗胚乳突变基因Wx mq直接相关的单核苷酸差异,设计合成了CAPS标记,该标记能区分含或不含Wx mq基因的纯合基因型和杂合基因型。利用该标记对武香粳14/关东194衍生的F5、F6株系进行辅助选择,筛选到含Wx mq基因的纯合基因型,成熟后对胚乳淀粉性质的调查结果与分子检测结果完全一致,分子标记辅助选择效果达100%。最终将关东194的暗胚乳突变基因Wx mq与高产基因聚合于一体,育成含有暗胚乳突变基因Wx mq的优良食味粳稻新品种南粳46。     

水稻胚乳外观云雾性状形成基础及其快速识别条件分析

以早籼稻新品种(系)、杂交水稻不育系、恢复系及其衍生的部分杂交组合为材料进行研究,发现云雾性状是低表观直链淀粉含量品种（系）特有的胚乳外观,其形成的原因和胚乳内淀粉的形态和结构有关。种子老化试验表明,种子水分和老化是快速识别云雾性状的两个重要条件。育种实践中可采用高温烘干和高温保湿预处理两种方法来快速识别胚乳云雾性状,并以此作为标记,早期特异性剔除低表观直链淀粉材料。     

Identification of Multiple Alleles at the Wx Locus and Development of Single Segment Substitution Lines for the Alleles in Rice

Amylose content (AC) is a key determinant of the cooking and processing quality of rice (Oryza sativa). It has been reported that the amylose content is mainly controlled by the Wx locus [1]. By using RFLP analysis, two alleles, largely corresponding to t…     

利用Wx和fgr基因双功能性标记高效选育优质水稻保持系

对选取的13份籼型保持系进行直链淀粉含量和香味测定及分子标记(484/485和fgr-1F/1R)检测,筛选出宜香1B作为试验用分子育种优质供体亲本。配组Ⅱ-32B/宜香1B,F2代进行该双标记分离分析,结果表明,不仅F2单株间特异性谱带差别明显便于鉴别,且呈现出两对独立遗传基因分离模式。选取F2代484/485和fgr-1F/1R呈Ⅰ型(与宜香B相同的带型)和Ⅲ型(杂合带型)的优良单株,经F3～F6加代,并结合农艺性状(株型、播抽历期等)和稻米品质性状(粒型、垩白度、透明度等)鉴定和选择,在F6再利用484/485和fgr-1F/1R分子标记进行复测和验证,从而获得了一批株系内性状基本稳定、株型好、直链淀粉含量中等、具有香味、外观和蒸煮食用品质优良的高世代株系。经测交鉴定,发现其保持系株系比例约占75.5%。表明利用Wx和fgr基因功能性分子标记的早世代检测和辅助选择,结合常规优质保持系鉴定和选育方法,可以快速获得直链淀粉含量中等、香味、长粒型、透明、低垩白、较易糊化等综合性状较优的保持系。     

胚乳外观标记与RVA谱理化指标相结合辅助改良早籼稻食用品质

对具有较低和中等表观直链淀粉含量的浙辐504和IR64的杂交后代进行观察,发现胚乳外观存在4种类型：乳白色、云雾状、透明状及各外观混合体。与胚乳外观紧密关联,表观直链淀粉含量(AAC)也相应有4种含量水平：明显低AAC变异型、浙辐504型、IR64型和介于双亲的中间型。RVA谱分析表明,不同胚乳外观的淀粉粘滞特性差异明显。透明状外观尽管AAC相仿,但个体间淀粉粘滞性分化明显,只有个别优良品系的RVA谱与优质亲本IR64相似。表明以胚乳外观标记和RVA谱理化指标辅助选择可有效控制表观直链淀粉含量和淀粉粘滞性,是改良早籼稻米品质的一条有效途径。     

Functional Marker Development and Effect Analysis of Grain Size Gene GW2 in Extreme Grain Size Germplasm in Rice

GW2 is an important gene that regulates grain width and weight. We used c DNA clone to obtain the sequences of GW2 from large- and small-grained rice varieties,TD70 and Kasalath,respectively. Then,we developed a d CAPS(derived cleaved amplified polymorphic sequence) marker on the basis of the sequence difference between functional and nonfunctional GW2 genes to analyze the genotypes and phenotypes of recombinant inbred lines. Results showed that the sequence of GW2TD70 had a single nucleotide deletion at site 316 that generates a termination codon. This codon terminated the GW2 protein in advance. By contrast,the sequence of GW2 Kasalath encoded an intact protein. A novel d CAPS marker was designed in accordance with a base A deletion at site 316 of the sequence. After the PCR product was digested by Apo I,TD70 showed 21 and 30 bp fragments,and Kasalath showed a 51 bp fragment. Up to 82 lines contained GW2TD70,and 158 lines contained GW2 Kasalath. The lines that contained TD70 alleles displayed substantial increases in width and 1000-grain weight. This result suggested that GW2 played a critical role in rice breeding.     

一种水稻香味基因功能标记的开发

为了提高水稻香味基因的分子标记辅助选择的准确性,根据水稻隐性香型品种与非香型品种的BAD2基因（即香味基因）序列之间存在8 bp碱基缺失,设计出香味基因fgr的InDel功能标记GRFM04。利用该标记对粤丰B（香型）/振丰B（非香型）的F2分离群体和其他16份香稻品种和6份非香稻品种进行检测验证。依据其PCR扩增产物电泳带型,可以准确地区分出香型纯合基因型、非香型纯合基因型和杂合基因型3种带型,且3种带型与其植株或品种相应的香味性状表现完全呈一一对应关系。     

Lowering Grain Amylose Content in Backcross Offsprings of indica Rice Variety 057 by Molecular Marker-Assisted Selection

To lower the amylose content （AC） of the indica rice restorer line 057 with high AC, backcrosses were made respectively by using four indica varieties （R367, 91499, Yanhui 559, Hui 527） as low AC donor parents and 057 as the recurrent parent. A molecular marker （PCR-Acc Ⅰ） was used to identify the genotypes （GG, TT and GT） of the waxy （Wx） gene. Plants with GT genotype were selected and used as female parent and crossed with 057 to advance generation. The ACs of rice grains harvested from plants with different Wx genotypes were measured and compared to analyze the efficiency of marker-assisted selection. The ACs of the rice grain, harvested from the plants of Wx genotypes GG, GT and TT, were higher than 20%, in the range of 17.7-28.5%, and less than 18%, respectively. The PCR-Acc Ⅰ marker could be used for efficiently lowering the AC of 057 through backcrossing, and there were some influence of parental genetic background on the AC of rice grains with the same Wx genotype.     

抗稻瘟病基因Pi9的STS连锁标记开发及在分子标记辅助育种中的应用

 利用带有广谱抗稻瘟病基因Pi9的籼稻品系75 1 127作为抗病基因供体亲本,用于扬稻6号和R6547抗病性的回交育种。通过比较75 1 127、扬稻6号和日本晴的Pi9基因位点的DNA序列,开发出了与Pi9基因紧密连锁的共显性STS（序列标记位点）标记PB9 1,用于Pi9基因的分子标记辅助选择。结合田间农艺性状选择和分子标记辅助选择,培育出8个Pi9基因纯合的回交后代株系。其中,具有扬稻6号和R6547遗传背景的株系各4个。经湖北恩施和宜昌的病圃鉴定,携带有抗病基因株系的稻瘟病抗性水平较受体品种扬稻6号和R6547有不同程度的提高。具有R6547遗传背景的株系08C893配制的杂交组合在上述病区的抗性表现也明显优于对照品种扬两优6号。上述结果说明,共显性标记PB9 1在Pi9抗稻瘟病基因分子标记辅助育种中具有应用价值,并且Pi9基因作为稻瘟病抗源之一可以在湖北稻区进行有效利用。     

Optimization of a Reaction System of Sequence Related Amplified Polymorphism and Segregation of Polymorphic Loci in an F_2 Population of Rice

An effective PCR protocol for detecting the sequence related amplified polymorphism (SRAP) in rice was developed.One hundred and ten pairs of SRAP primers were used for segregation analysis in an F2 population derived from a cross between Shennong 606 and Lijiangxintuanheigu.Among the 110 primer pairs,35 pairs generated 143 polymorphic bands with an average of 4.09 polymorphic bands per primer pair,and 24 pairs (16.78%) showed the genetic distortion (P＜0.05).Of the 24 primer pairs,12 pairs deviated toward the male parent Shennong 606 and 11 pairs toward the female parent Lijiangxintuanheigu,only one toward heterozygote.It was found that the segregation distortion might be caused by the joint gametic and zygotic effects.     

Optimization of a Reaction System of Sequence Related Amplified Polymorphism and Segregation of Polymorphic Loci in an F2 Population of Rice

An effective PCR protocol for detecting the sequence related amplified polymorphism （SRAP） in rice was developed. One hundred and ten pairs of SRAP primers were used for segregation analysis in an F2 population derived from a cross between Shennong 606 and Lijiangxintuanheigu. Among the 110 primer pairs, 35 pairs generated 143 polymorphic bands with an average of 4.09 polymorphic bands per primer pair, and 24 pairs （16.78%） showed the genetic distortion （P〈0.05）. Of the 24 primer pairs, 12 pairs deviated toward the male parent Shennong 606 and 11 pairs toward the female parent Lijiangxintuanheigu, only one toward heterozygote. It was found that the segregation distortion might be caused by the joint gametic and zygotic effects.     

抗稻瘟病基因Pi9的STS连锁标记开发及在分子标记辅助育种中的应用

利用带有广谱抗稻瘟病基因Pi9的籼稻品系75-1-127作为抗病基因供体亲本,用于扬稻6号和R6547抗病性的回交育种。通过比较75-1-127、扬稻6号和日本晴的Pi9基因位点的DNA序列,开发出了与Pi9基因紧密连锁的共显性STS(序列标记位点)标记PB9-1,用于Pi9基因的分子标记辅助选择。结合田间农艺性状选择和分子标记辅助选择,培育出8个Pi9基因纯合的回交后代株系。其中,具有扬稻6号和R6547遗传背景的株系各4个。经湖北恩施和宜昌的病圃鉴定,携带有抗病基因株系的稻瘟病抗性水平较受体品种扬稻6号和R6547有不同程度的提高。具有R6547遗传背景的株系08C893配制的杂交组合在上述病区的抗性表现也明显优于对照品种扬两优6号。上述结果说明,共显性标记PB9-1在Pi9抗稻瘟病基因分子标记辅助育种中具有应用价值,并且Pi9基因作为稻瘟病抗源之一可以在湖北稻区进行有效利用。     

抗水稻白叶枯病新基因Xa-25(t)的RAPD分析

利用RAPD标记对从体细胞突变体HX-3中鉴定出的抗水稻白叶枯病新基因Xa- 25 (t)进行了研究,在306个随机引物中,两个引物S1269和S1327在亲本和抗感池（均由12个高抗或高感植株组成）之间表现多态性。用这两个引物对龙特甫A和HX-3的F2群体的114个单株进行连锁分析,表明 S1269和 S1327与Xa-25 (t)的连锁距离分别为5.3和23.7 cM,且位于Xa-25 (t)的两侧。     

广西杂交稻组合抗白叶枯病基因Xa-4的分子标记检测

采用SSR分子标记技术,对广西推广面积较大的45份三系杂交稻组合的Xa-4基因进行了分子检测,结果表明,33份(73.3%)材料含有Xa-4纯合基因,9份(20%)材料含有Xa-4杂合基因.在广西杂交水稻育种中应注意避免抗性基因单一化.