药用野生稻转育后代一个抗白叶枯病新基因的定位

从药用野生稻渗入后代选育的水稻株系B5表现为高抗褐飞虱、白背飞虱和白叶枯病。对B5与籼稻品种明恢63杂交组合的187个重组自交系(RILs)进行了抗白叶枯病接种鉴定,采用分离集团分析法(Bulked Segregant Analysis,BSA),在第1染色体上筛选到与水稻抗白叶枯病基因相连锁RFLP分子标记。利用RILs抗病性表现型鉴定资料和构建的分子标记连锁图谱,将抗白叶枯病基因定位在第1染色体短臂的C904和R596之间,这两个分子标记间遗传距离为1．3cM。该基因对RILs群体抗病性变异的贡献率为52．96％,是一效应值较大的主效基因。这一抗白叶枯病基因不同于已报道的抗白叶枯病基因的位点,因此将其命名为Xa29(t)。     

利用SSR定位籼稻品种Kaharamana中抗褐飞虱基因Bph9

褐飞虱是危害水稻生产最重要的害虫之一,利用寄主抗性被认为是防治褐飞虱最经济而有效的方法。斯里兰卡水稻品种Kaharamana对东亚和东南亚的褐飞虱种群均表现抗虫性,利用分子遗传学的方法对其携带的Bph9基因进行了SSR定位。所用的遗传群体为来源于Kaharamana和02428的含有180个单株的F2分离群体,每个F2单株套袋自交获得F2：3家系。利用苗期集团鉴定埘F2：3家系进行抗褐飞虱鉴定,以推测相应F2单株的基因型。连锁分析表明,Bph9位于第12染色体上的两个SSR标记RM463和RM5341之间,分别与之相距6．8cM和9．7cM。该标记有助十将Bph9用于分子标记辅助选择育种研究。     

水稻抗白叶枯病基因Xa12区间连锁图的构建

利用SSR标记对143株Java14/珍珠矮F₂随机群体进行分析,构建了水稻第4染色体上抗白叶枯病基因Xa12高饱和度的SSR标记区间连锁图。该分子图谱整合了SSR标记RM349、RM348、RM255、MRG4611,为进一步利用SSR标记在Java14/珍珠矮F₂群体中精细定位Xa12以及克隆该基因奠定了基础。     

体细胞突变体HX-3抗水稻白叶枯病基因的鉴定

以感病杂交稻恢复系明恢63的成熟胚为外植体,利用离体筛选技术获得了抗水稻白叶枯病细胞突变体HX-3。连续8年以我国长江流域白叶枯病代表菌析浙173（IV型）对HX-3的R1到R9代进行抗性鉴定,HX-3的抗病性可以稳定遗传。抗性遗传分析表明HX-3的抗性由1对显性核基因控制。1999～2000年连续两年利用我国、菲律宾和日本的32个水稻白叶枯病菌株,测定HX-3及IRBB1等13个具不同显性抗病基因的近等基因系抗性,HX-3抗谱广,且与已知显性抗病基因的抗谱不同。在此基础上,以抗白叶枯病近等基因系IRBB4、IRBB7、CBB12和IRBB21和HX-3杂交,进行等位性分析,4个杂交组合的F2代均出现抗、感分离,说明HX-3与这4个基因不等位。综合以上研究结果,HX-3具有1个新的抗白叶枯病基因,暂命名为Xa-25(t）。     

水稻长穗颈基因eui紧密连锁SSR标记获得

02428h是从半矮秆材料02428体细胞培养后代中发现的隐性高秆突变体, 其株高性状由1对长穗颈基因eui和1对半矮秆基因sd-1共同控制。以02428h与半矮秆材料南京11杂交的F₂为作图群体, 利用Gramene公布的SSR标记和根据NCBI中的BAC序列自己新开发的SSR标记, 将eui基因定位在第5染色体上的RM3673和RM0012之间, 两侧遗传距离分别为0.3 cM和1.0 cM, 为该基因的分子标记辅助选择奠定了基础。     

水稻抗褐飞虱基因bph2的SSR定位和标记辅助选择

利用综合性状较好对褐飞虱敏感的粳稻恢复系C418为父本,以含有bph2基因的抗褐飞虱品种ASD7为母本构建了包含134个F23家系的群体,利用苗期鉴定法对F2：3家系进行抗性鉴定：用SSR标记技术,将bph2基因定位在第12染色体长臂上,标记RM7102和RM463之间,其遗传距离分别为7．6cM和7．2cM。在进行表型选择的同时,利用与bph2基因连锁的SSR标记RM7102和RM463对BC1F1和BC2F1进行了标记辅助选择,选择效率分别为89．9％和91．2％,为培育高抗褐飞虱水稻品种奠定了基础。     

基于CSSL的水稻抽穗期QTL定位及遗传分析

抽穗期是水稻(Oryza sativa)品种的重要农艺性状之一, 适宜的抽穗期是获得理想产量的前提。鉴定和定位水稻抽穗期基因/QTL, 分析其遗传效应对改良水稻抽穗期至关重要。以籼稻品种9311(Oryza sativa ssp. indica ‘Yangdao 6’)为受体,粳稻品种日本晴(Oryza sativa ssp. japonica ‘Nipponbare’)为供体构建的94个染色体片段置换系群体为材料, 以P≤0.01为阈值, 对置换片段上的抽穗期QTL进行了鉴定。采用代换作图法共定位了4个控制水稻抽穗期的QTL, 分别位于第3、第4、第5和第8染色体; QTL的加性效应值变化范围为–6.4 – –2.7, 加性效应百分率变化范围为–6.4%– –2.7%; qHD-3和qHD-8加性效应值较大, 表现主效基因特征。为了进一步定位qHD-3和qHD-8, 在目标区域加密16对SSR引物, qHD-3和qHD-8分别被界定在第3染色体RM3166–RM16206之间及第8染色体RM4085-RM8271之间, 其遗传距离分别为13.9 cM和6.4 cM。研究结果为利用分子标记辅助选择改良水稻抽穗期奠定了基础。     

基于CSSL的水稻抽穗期QTL定位及遗传分析

抽穗期是水稻（Oryza sativa）品种的重要农艺性状之一,适宜的抽穗期是获得理想产量的前提。鉴定和定位水稻抽穗期基因/QTL,分析其遗传效应对改良水稻抽穗期至关重要。以籼稻品种9311（Oryzasativa ssp.indica‘Yangdao 6’）为受体,粳稻品种日本晴（Oryza sativa ssp.japonica‘Nipponbare’）为供体构建的94个染色体片段置换系群体为材料,以P≤0.01为阈值,对置换片段上的抽穗期QTL进行了鉴定。采用代换作图法共定位了4个控制水稻抽穗期的QTL,分别位于第3、第4、第5和第8染色体;QTL的加性效应值变化范围为–6.4––2.7,加性效应百分率变化范围为–6.4%––2.7%;qHD-3和qHD-8加性效应值较大,表现主效基因特征。为了进一步定位qHD-3和qHD-8,在目标区域加密16对SSR引物,qHD-3和qHD-8分别被界定在第3染色体RM3166–RM16206之间及第8染色体RM4085–RM8271之间,其遗传距离分别为13.9cM和6.4cM。研究结果为利用分子标记辅助选择改良水稻抽穗期奠定了基础。     

水稻白色中脉Oswm2的遗传分析与分子标记定位

从T-DNA突变体库中获得一份以中花11为遗传背景的白色中脉突变体。该突变体剑叶以下叶片的中下部中脉表现为白色, 白色中脉附近的叶色微黄, 并且伴随株高等农艺性状的改变, 暂时将其定名为Oswm2(Oryza sativa white midrib 2)。遗传分析表明该突变性状受一对隐性单基因控制, 以Oswm2和粳稻02428杂交的F2分离群体作为定位群体, 将OsWM2基因定位在水稻第7染色体的SSR标记RM21478和RM418之间, 遗传距离分别为8.7和15.9 cM。     

矮泰引-3中半矮秆基因的分子定位

矮泰引-3的矮生性状受两对独立遗传的半矮秆基因控制,利用SSR标记将这两个矮秆基因分别定位到第1和第4染色体上。等位性测交的结果表明,位于第1染色体上的矮秆基因与sd1是等位的,所以仍然称其为sd1;而位于第4染色体上的矮秆基因是一个新基因,暂命名为sdt2。利用SSR标记将sd1定位于RM297、RM302和RM212的同一侧,而与OSR3共分离,它们之间的位置关系可能是RM297-RM302-RM212-OSR3-sd1,遗传距离分别为4．7cM、0cM、0．8cM和0cM,这与sd1在第1染色体长臂上的确切位置是基本一致的。利用已有的SSR标记和拓展的SSR标记将sdt2定位于SSR332、RM1305和RM5633、RM307、RM401之间,它们的排列位置可能是SSR332-RM1305-sdt2-RM5633-RM307-RM401,它们之间的遗传距离分别为11．6cM、3．8cM、0．4cM、0cM和0．4cM。     

Tagging and mapping of rice sheath blight resistant gene

Sheath blight (Rhizoctonia solani Kühn) is one of the severe rice diseases worldwide. In this study, an F(2) population from a cross between "4011" and "Xiangzaoxian19" is used to identify molecular markers linked with the resistant trait. "4011" was a transgenic rice cultivar carrying a resistant gene to sheath blight, while "Xiangzaoxian19" is a highly susceptible one. As a result, five molecular markers, including three RFLP markers converted from RAPD and AFLP markers, and two SSR markers were identified to link with the sheath blight resistant gene. This dominant resistant gene was named as R sb 1 and mapped on rice chromosome 5. The linkage distance between the markers (E-AT:M-CAC(120), E-AT:M-CTA(230), OPN-16(2000), RM164(320) and RM39(300)) and R sb 1 was 1.6 cM, 9.9 cM, 1.6 cM, 15.2 cM and 1.6 cM, respectively.     

Genetic and physical mapping of a new gene for bacterial blight resistance in rice

The inheritance of resistance for bacterial blight, caused by Xanthomonas oryzae pv. oryzae ( Xoo), was studied in Minghui 63, an elite restorer line for a number of widely used rice hybrids in China. A new dominant gene against a Chinese Xoo strain JL691 in both the seedling and adult stages was identified in Minghui 63 and designated as Xa26( t). Using a total of 477 highly susceptible individuals from an F(2) population, the Xa26( t) locus was mapped to a region of about 1.68 cM. This locus co-segregated with marker R1506 and was 0.21 cM from marker RM224 on one side and 1.47 cM from marker Y6855RA on the other side, in rice chromosome 11. A contig map, composed of five non-redundant bacterial artificial chromosome (BAC) clones and spanning approximately 500 kb in length, was constructed. Analysis of recombination events in the Xa26( t) region with the highly susceptible F(2) individuals anchored the gene locus to a region covered by three overlapped BAC clones. Assay of the lines showing a double crossover in marker loci flanking Xa26( t), in a population of recombinant inbred lines carrying Xa26( t), further delineated the gene to a 20-kb fragment. The Xa26( t) locus is tightly linked to another bacterial blight resistance gene locus, Xa4.     

A SNP haplotype associated with a gene resistant to Xanthomonas axonopodis pv. malvacearum in upland cotton (Gossypium hirsutum L.)

An F_4:5 population of 285 families with each tracing back to a different F₂ plant, derived from a cotton bacterial blight resistant line ‘DeltaOpal’ and a susceptible line ‘DP388’, was artificially inoculated with bacterial blight race 18 (Xanthomonas axonopodis pv. malvacearum) to assay their resistance or susceptibility to the disease. The segregation in the F_4:5 population indicates that the resistance was conditioned by a single dominant gene designated B _12. Simple sequence repeat (SSR) markers identified as putatively linked to the resistance gene by bulked segregant analysis were confirmed on the entire F_4:5 population. Three SSR markers, CIR246, BNL3545 and BNL3644 on chromosome 14, were found closely linked to B ₁₂ . The association between CIR246 and B ₁₂ was validated among 354 plants of 16 diverse varieties. Based on Monsanto SSR/single nucleotide polymorphism (SNP) consensus map, SNP markers closely linked to CIR246 were used to screen ‘DeltaOpal’ and ‘DP388’ for polymorphism. The polymorphic SNP markers were run on the F_4:5 population and the four SNP markers spanning 3.4 cM were found to flank the resistance gene on chromosome 14. The linkage between B ₁₂ and the 4-SNP marker haplotype was validated using 18 elite cotton lines. This 4-SNP marker haplotype can be used for marker assisted selection for bacterial blight resistance breeding programs or for screening germplasm collections for this locus rapidly.     

Genetic Analysis and Molecular Mapping of a Novel Gene Conferring Resistance to Rice Stripe Virus

Rice stripe virus (RSV) is one of the most damaging diseases affecting rice in East Asia. Rice variety 502 is highly resistant to RSV, while variety 5112 is extremely susceptible. Field statistical data revealed that all “502 × 5112” F₁ individuals were resistant to RSV and the ratio of resistant to susceptible plants was 3:1 in the F₂ population and 1:1 in the BC₁F₁ population. These results indicated that a dominant gene, designated RSV1, controlled the resistance. Simple sequence repeat (SSR) analysis was subsequently carried out in an F₂ population. Sixty SSR markers evenly distributed on the 12 rice chromosomes were screened and tested. Two markers, RM229 and RM206, showed linkage with RSV1. Based on this result, six SSR markers flanking RM229 and RM206 were further selected and tested. Results indicated that SSR markers RM457 and RM473E were linked to RSV1 with a genetic distance of 4.5 and 5.0 cM, respectively. All of the four SSR markers (RM229, RM473E, RM457 and RM206) linked to RSV1 were all located on chromosome 11, therefore RSV1 should be located on chromosome 11 also. In order to find some new markers more closely linked to the RSV1 gene, sequence-related amplified polymorphism (SRAP) analysis was performed. A total of 30 SRAP primer-pairs were analyzed, and one marker SR1 showed linkage with RSV1 at a genetic distance of 2.9 cM. Finally, RSV1 gene was mapped on chromosome 11 between SSR markers RM457 and SRAP marker SR1 with a genetic distance of 4.5 cM and 2.9 cM, respectively.     

Tagging and combining bacterial blight resistance genes in rice using RAPD and RFLP markers

Four genes of rice,Oryza sativa L., conditioning resistance to the bacterial blight pathogenXanthomonas oryzae pv.oryzae (X. o. pv.oryzae), were tagged by restriction fragment length polymorphism (RFLP) and random amplified polymorphic DNA (RAPD) markers. No recombinants were observed betweenxa-5 and RFLP marker lociRZ390, RG556 orRG207 on chromosome 5.Xa-3 andXa-4 were linked to RFLP locusXNpb181 at the top of chromosome 11, at distances of 2.3 cM and 1.7 cM, respectively. The nearest marker toXa-10, also located on chromosome 11, was the RAPD locusO07 ₂₀₀₀ at a distance of 5.3 cM. From this study, the conventional map [19, 28] and two RFLP linkage maps of chromosome 11 [14, 26] were partially integrated. Using the RFLP and RAPD markers linked to the resistance genes, we selected rice lines homozygous for pairs of resistance genes,Xa-4 +xa-5 andXa-4 +Xa-10. Lines carryingXa-4 +xa-5 andXa-4 +Xa-10 were evaluated for reaction to eight strains of the bacterial blight pathogen, representing eight pathotypes and three genetic lineages. As expected, the lines carrying pairs of genes were resistant to more of the isolates than their single-gene parental lines. Lines carryingXa-4 +xa-5 were more resistant to isolates of race 4 than were either of the parental lines (quantitative complementation). No such effects were seen forXa-4 +Xa-10. Thus, combinations of resistance genes provide broader spectra of resistance through both ordinary gene action expected and quantitative complementation.     

Genetic analysis and molecular mapping of a novel recessive gene xa34(t) for resistance against Xanthomonas oryzae pv. oryzae

A new bacterial blight recessive resistance gene xa34(t) was identified from the descendant of somatic hybridization between an aus rice cultivar (cv.) BG1222 and susceptible cv. IR24 against Chinese race V (isolate 5226). The isolate was used to test the resistance or susceptibility of F₁ progenies and reciprocal crosses of the parents. The results showed that F₁ progenies appeared susceptibility there were 128R (resistant):378S (susceptible) and 119R:375S plants in F₂ populations derived from two crosses of BG1222/IR24 and IR24/BG1222, respectively, which both calculates into a 1R:3S ratio. 320 pairs of stochastically selected SSR primers were used for genes?? initial mapping. The screened results showed that two SSR markers, RM493 and RM446, found on rice chromosome 1 linked to xa34(t). Linkage analysis showed that these two markers were on both sides of xa34(t) with the genetic distances 4.29 and 3.05?cM, respectively. The other 50 SSR markers in this region were used for genes?? fine mapping. The further results indicated that xa34(t) was mapped to a 1.42?cM genetic region between RM10927 and RM10591. In order to further narrow down the genomic region of xa34(t), 43 of insertion/deletion (Indel) markers (BGID1-43) were designed according to the sequences comparison between japonica and indica rice. Parents?? polymorphic detection and linkage assay showed that the Indel marker BGID25 came closer to the target gene with a 0.4?cM genetic distance. A contig map corresponding to the locus was constructed based on the reference sequences aligned by the xa34(t) linked markers. Consequently, the locus of xa34(t) was defined to a 204?kb interval flanked by markers RM10929 and BGID25.     

Genetic Analysis and Molecular Mapping of a Rolling Leaf Mutation Gene in Rice

A rice mutant with rolling leaf, namely γ-rl, was obtained from M2 progenies of a native indica rice stable strain Qinghuazhan （QHZ） from mutagenesis of dry seeds by γ-rays. Genetic analysis using the F2 population from a cross between this mutant and QHZ indicated the mutation was controlled by a single recessive gene. In order to map the locus for this mutation, another F2 population with 601 rolling leaf plants was constructed from a cross between y-rl and a japonica cultivar 02428. After primary mapping with SSR （simple sequence repeats） markers, the mutated locus was located at the short arm of chromosome 3, flanked by RM6829 and RM3126. A number of SSR, InDel （insertion/deletion） and SNP （single nucleotide polymorphism） markers within this region were further developed for fine mapping. Finally, two markers, SNP121679 and InDe1422395, were identified to be flanked to this locus with genetic distances of 0.08 cM and 0.17 cM respectively, and two SNP markers, SNP75346 and SNPl10263, were found to be co-segregated with this locus. These results suggested that this locus was distinguished from all loci for the rolling leaf mutation in rice reported so far, and thus renamed rl10（t）. By searching the rice genome database with closely linked markers using BLAST programs, an e-physical map covering rl10（t） locus spanning about a 50 kb region was constructed. Expression analysis of the genes predicted in this region showed that a gene encoding putative flavin-containing monooxygenase （FMO） was silenced in γ-rl, thus this is the most likely candidate responsible for the rolling leaf mutation.     

Identification of a YAC clone carrying the Xa-1 allele, a bacterial blight resistance gene in rice

Map-based cloning methods have been applied for isolation of Xa-1, one of the bacterial blight resistance genes in rice.Xa-1 was previously mapped on chromosome 4 using molecular markers. For positional cloning of Xa-1, a high-resolution genetic map was made for theXa-1 region using an F₂ population of 402 plants and additional molecular markers. Three restriction fragment length polymorphism (RFLP) markers, XNpb235, XNpb264 and C600 were found to be linked tightly to Xa-1, with no recombinants, and U08 ₇₅₀ was mapped 1.5 cM from Xa-1. The screening of a yeast artificial chromosome (YAC) library using theseXa-1-linked RFLP markers resulted in the identification of ten contiguous YAC clones. Among these, one YAC clone, designated Y5212, with an insert of 340 kb, hybridized with all three tightly linked markers. This YAC was confirmed to possess the Xa-1 allele by mapping the Xa-1 gene between both end clones of this YAC (Y5212R and Y5212L).