Fine mapping and candidate gene analysis of a green-revertible albino gene gra（t） in rice

Green-revertible albino is a novel type of chlorophyll deficiency in rice （Oryza sativa L.）, which is helpful for further research in chlorophyll synthesis and chloroplast development to illuminate their molecular mechanism. In the previous study, we had reported a single recessive gene, gra（t）, controlling this trait on the long arm of chromosome 2. In this paper, we mapped the gra（t） gene using 1,936 recessive individuals with albino phenotype in the F2 population derived from the cross between themo-photoperiod-sensitive genic male-sterile （T/PGMS） line Pei＇ai 64S and the spontaneous mutant Qiufeng M. Eventually, it was located to a confined region of 42.4 kb flanked by two microsatellite markers RM2-97 and RM13553. Based on the annotation results of RiceGAAS system, 11 open reading frames （ORFs） were predicted in this region. Among them, ORF6 was the most possible gene related to chloroplast development, which encoded the chloroplast protein synthesis elongation factor Tu in rice. Therefore, we designated it as the candidate gene of gra（t）. Sequence analysis indicated that only one base substitution C to T occurred in the coding region, which caused a missense mutation （Thr to Ile） in gra（t） mutant. These results are very valuable for further study on gra（t） gene.     

Molecular evolution and functional divergence of HAK potassium transporter gene family in rice （Oryza sativa L.）

The high-affinity K＋ （HAK） transporter gene family is the largest family in plant that functions as potassium transporter and is important for various aspects of plant life. In the present study, we identified 27 members of this family in rice genome. The phylogenetic tree divided the land plant HAK transporter proteins into 6 distinct groups. Although the main characteristic of this family was established before the origin of seed plants, they also showed some differences between the members of non-seed and seed plants. The HAK genes in rice were found to have expanded in lineage-specific manner after the split of monocots and dicots, and both segmental duplication events and tandem duplication events contributed to the expansion of this family. Functional divergence analysis for this family provided statistical evidence for shifted evolutionary rate after gene duplication. Further analysis indicated that both point mutant with positive selection and gene conversion events contributed to the evolution of this family in rice.     

Identification of quantitative trait loci for four morphologic traits under water stress in rice （Oryza sativa L.）

Late season drought coinciding with the rice booting to heading stage affects the development of plant height,panicle exsertion,and flag leaf size,and causes significant yield loss.In this study,a recombinant inbred line population derived from a cross between paddy and upland cultivars was used for data collection of the morphologic traits under well water and drought stress conditions.bought stress was applied at the stage of panicle initiation in the field in 2002 and at the booting stage in PVC pipes in 2003.The data from stress con ditions and their ratios(tait measured under stress condition/trait measured under well water condition)or differences(trait measured under stress condition minus trait measured under well water condition)were used for OTL analysis.Totally,17 and 36 QTLs for these traits were identified in 2002 and 2003,respectively,which explained a range of 2.58%-29.82%Of the phenotypic variation.Among them,six QTLs were commonly identified in the two years,suggesting that the drought stress in the two years was different.The genetic basis of these traits will provide useful information for improving rice late season drought resistance,and their application as indirect indices in rice late season drought resistance screening was also discussed.     

Characterization and mapping of a novel mutant sms1 （senescence and male sterility 1） in rice

Plant senescence plays diverse important roles in development and environmental responses.However,the molecular basis of plant senescence is remained largely unknown.A rice spontaneous mutant with the character of early senescence and male sterility (sms) was found in the breeding line NT10-748.In order to identify the gene SMS1 and the underlying mechanism,we preliminarily analyzed physiological and biochemical phenotypes of the mutant.The mutant contained lower chlorophyll content compared with the wild type control and was severe male sterile with lower pollen viability.Genetic analysis showed that the mutant was controlled by a single recessive gene.By the map-based cloning approach,we fine-mapped SMS1 to a 67 kb region between the markers Z3-4 and Z1-1 on chromosome 8 using 1,074 F2 recessive plants derived from the cross between the mutant sms1 (japonica) × Zhenshan 97 (indica),where no known gene involved in senescence or male sterility has been identified.Therefore the SMS1 gene will be a novel gene that regulates the two developmental processes.The further cloning and functional analysis of the SMS1 gene is under way.     

Novel pleiotropic loci controlling panicle architecture across environments in japonica rice （Oryza sativa L.）

To identify quantitative trait loci (QTLs) controlling panicle architecture in japonica rice, a genetic map was constructed based on simple sequence repeat (SSR) markers and 254 recombinant inbred lines (RILs) derived from a cross between cultivars Xiushui 79 and C Bao. Seven panicle traits were investigated under three environments. Single marker analysis indicated that a total of 27 SSR markers were highly associated with panicle traits in all the three environments. Percentage of phenotypic variation explained by single locus varied from 2% to 35%. Based on the mixed linear model, a total of 40 additive QTLs for seven panicle traits were detected by composite interval mapping, explaining 1.2%-35% of phenotypic variation. Among the 9 QTLs with more than 10% of explained phenotypic variation, two QTLs were for the number of primary branches per panicle (NPB), two for panicle length (PL), two for spikelet density (SD), one for the number of secondary branches per panicle (NSB), one for secondary branch distribution density (SBD), and one for the number of spikelets per panicle (NS), respectively. qPLSD-9-1 and qPLSD-9-2 were novel pleiotropic loci, showing effects on PL and SD simultaneously. qPLSD-9-1 explained 34.7% of the phenotypic variation for PL and 25.4% of the phenotypic variation for SD, respec- tively. qPLSD-9-2 explained 34.9% and 24.4% of the phenotypic variation for PL and SD, respectively. The C Bao alleles at the both QTLs showed positive effects on PL, and the Xiushui 79 alleles at the both QTLs showed positive effects on SD. Genetic variation of panicle traits are mainly attributed to additive effects. QTL × environment interactions were not significant for additive QTLs and additive × additive QTL pairs.     

Genetic analysis of tolerance to photo-oxidative stress induced by high light in winter wheat （Triticum aestivum L.）

High light induced photooxidation (HLIP) usually leads to leaf premature senescence and causes great yield loss in winter wheat. In order to explore the genetic control of wheat tolerance to HLIP stress, a quantitative trait loci (QTL) analysis was conducted on a set of doubled haploid population, derived from two winter wheat cultivars. Actual values of chlorophyll content (Chl), minimum fluorescence level (Fo), maximum fluorescence level (Fm), and the maximum quantum efficiency of photosystem Ⅱ (Fv/Fm) under both HLIP and non-stress conditions as well as the ratios of HLIP to non-stress were evaluated. HLIP considerably reduced Chl, Fm, and Fv/Fm, but increased Fo, compared with that under non-stress condition. A total of 27, 16, and 28 QTLs were associated with the investigated traits under HLIP and non-stress and the ratios of HLIP to non-stress, respectively. Most of the QTLs for the ratios of HLIP to non-stress collocated or nearly linked with those detected under HLIP condition. HLIP-induced QTLs were mapped on 15 chromosomes, involving in 1A, 1B, 1D,2A, 2B, 2D, 3A, 3B, 4A, 4D, 5B, 6A, 6B, 7A, and 7D while those expressed under non-stress condition involved in nine chromosomes, including 1B, 1D, 2A, 2B, 3B, 4A, 5A, 5B, and 7A. The expression patterns of QTLs under HLIP condition were different from that under non-stress condition except for six loci on five chromosomes. The phenotypic variance explained by individual QTL ranged from 5.0% to 19.7% under HLIP, 8.3% to 20.8% under non-stress, and 4.9% to 20.2% for the ratios of HLIP to non-stress, respectively. Some markers, for example,Xgwm192 and WMC331 on 4D regulating Chl, Fo, Fm, and Fv/Fm under HLIP condition, might be used in marker assistant selection.     

Expression of PIN Genes in Rice （Oryza sativa L.）： Tissue Specificity and Regulation by Hormones

Twelve genes of the PIN family in rice were analyzed for gene and protein structures and an evolutionary relationship with reported AtPINs in Arabidopsis. Four members of PIN1 （designated as OsPINla-d）, one gene paired with AtPIN2 （OsPIN2）, three members of PIN5 （OsPIN5a-c）, one gene paired with AtPIN8 （OsPIN8）, and three monocot-specific PINs （OsPIN9, OsPINIOa, and b） were identified from the phylogenetic analysis. Tissue-specific expression patterns of nine PIN genes among them were investigated using RT-PCR and GUS reporter. The wide variations in the expression domain in different tissues of the PIN genes were observed. In general, PIN genes are up-regulated by exogenous auxin, while different responses of different PIN genes to other hormones were found.     

Subspecies-specific intron length polymorphism markers reveal clear genetic differentiation in common wild rice （Oryza rufipogon L.） in relation to the domestication of cultivated rice （O. sativa L.）

It is generally accepted that Oryza rufipogon is the progenitor of Asian cultivated rice （O. sativa）. However, how the two subspecies of O. sativa （indica and japonica） were domesticated has long been debated. To investigate the genetic differentiation in O. rufipogon in relation to the domestication of O. sativa, we developed 57 subspecies-specific intron length polymorphism （SSILP） markers by comparison between 10 indica cultivars and 10 japonica cultivars and defined a standard indica rice and a standard japonica rice based on these SSILP markers. Using these SSILP markers to genotype 73 O. rufipogon accessions, we found that the indica alleles and japonica alleles of the SSILP markers were predominant in the O. rufipogon accessions, suggesting that SSILPs were highly conserved during the evolution of O. sativa. Cluster analysis based on these markers yielded a dendrogram consisting of two distinct groups： one group （Group I） comprises all the O. rufipogon accesions from tropical （South and Southeast） Asia as well as the standard indica rice; the other group （Group II） comprises all the O. rufipogon accessions from Southern China as well as the standard japonica rice. Further analysis showed that the two groups have significantly higher frequencies of indica alleles and japonica alleles, respectively. These results support the hypothesis that indica rice and japonica rice were domesticated from the O. rufipogon of tropical Asia and from that of Southern China, respectively, and suggest that the indica-japonica differentiation should have formed in O. rufipogon long before the beginning of domestication. Furthermore, with an O. glaberrima accession as an outgroup, it is suggested that the indica-japonica differentiation in O. ruffpogon might occur after its speciation from other AA-genome species.     

Genetic analysis and gene mapping of a new rolled-leaf mutant in rice (Oryza sativa L.)

To understand the development of rice leaf blades,we identified a new rolled-leaf mutant,w32,from indica cultivar IR64 through EMS mutagenesis. The mutant showed a stable rolled-leaf phenotype throughout the life cycle. Two F2 populations were developed by crossing w32 to cultivar IR24 and PA64. Genetic analysis showed that the rolled-leaf phenotype was controlled by a single recessive gene. To determine the location of the gene,bulked segregant analysis was carried out using mutant and wild-type DNA pools ...     

Molecular cloning and structural analysis of a novel Rac gene osRACB in rice (Oryza sativa L.)

Rac is a subfamily of small GTP-binding protein family. Its molecular weight is between 20 and 30 kilodaltons. As a signal protein, Rac directly or indirectly participates in many physiological processes, such as the regulation of cytoskeleton and the transduction of stress-induced signal. So Rac is also named ?molecular switch? The switch is based on the cycle from a GTP-bound 憃n?to a GDP-bound 憃ff?state[1]. In the superfamily of GTP-binding protein, only heterotrimeric G protein, Ra…     

Genetic analysis and gene mapping of a rice few-tillering mutant in early backcross populations (Oryza sativa L.)

A rice mutant, G069, characteristic of few tiller numbers, was found in anther culture progeny from the F1 hybrid between an indica-japonica cross, Gui630×02428. The mutant has another two major features: delayed tillering development and yellowing apex and margin on the mature leaves. As a donor parent, G069 was further backcrossed with the recurrent parent, 02428, for two turns to develop a BC2F2 population. Genetic analysis in the BC2F2 population showed that the traits of few-tillering and yellowing apex and margin on the mature leaves were controlled by one recessive gene. A pool of equally mixed genomic DNA, from few-tillering individual plants in BC2F2, was constructed to screen polymorphism with simple sequence repeat (SSR) markers in comparison with the 02428 genome. One SSR marker and three restriction fragment length polymorphism (RFLP) markers were found possibly linked with the recessive gene. By using these markers, the gene of few-tillering was mapped on chromosome 2 between RFLP marker C     

Development and mapping of 2240 new SSR markers for rice (Oryza sativa L.).

A total of 2414 new di-, tri- and tetra-nucleotide non-redundant SSR primer pairs, representing 2240 unique marker loci, have been developed and experimentally validated for rice (Oryza sativa L.). Duplicate primer pairs are reported for 7% (174) of the loci. The majority (92%) of primer pairs were developed in regions flanking perfect repeats > or = 24 bp in length. Using electronic PCR (e-PCR) to align primer pairs against 3284 publicly sequenced rice BAC and PAC clones (representing about 83% of the total rice genome), 65% of the SSR markers hit a BAC or PAC clone containing at least one genetically mapped marker and could be mapped by proxy. Additional information based on genetic mapping and "nearest marker" information provided the basis for locating a total of 1825 (81%) of the newly designed markers along rice chromosomes. Fifty-six SSR markers (2.8%) hit BAC clones on two or more different chromosomes and appeared to be multiple copy. The largest proportion of SSRs in this data set correspond to poly(GA) motifs (36%), followed by poly(AT) (15%) and poly(CCG) (8%) motifs. AT-rich microsatellites had the longest average repeat tracts, while GC-rich motifs were the shortest. In combination with the pool of 500 previously mapped SSR markers, this release makes available a total of 2740 experimentally confirmed SSR markers for rice, or approximately one SSR every 157 kb.     

水稻显性早熟材料D64B的发现、遗传分析和分子标记定位

D64B是从籼型杂交稻保持系D63B中发现的一个无色早熟突变株。用不育系、保持系、恢复系以及早稳型水稻品种与之杂交,F1的抽穗期多数与早熟亲本D64B相同或相近,部分偏向早熟亲本。这些结果表明D64B具有显性早熟特性。将D64B在海南陵水短日照和温江长日照下分期种植,观察到两地点因生长发育期间温度变化引起的抽穗期的变化的程度是一致的,并且在一定范围内随着生长发育期间温度升高,D64B抽穗缩短,可知D64B不感光,感温性中等。种植D64B与蜀恢527的正反交F2和回交一代BC1,三者的抽穗期均呈双峰分布,并且峰谷处于同一位置,以峰谷值103d为转折点进行分组,早熟与迟熟植株的分离比经x^2检验分别符合3：1和1：1,表明D64B的早熟特性主要受一对显性早熟核基因控制。用356对微卫星引物对亲本D64B和蜀恢527进行多态性分析,并用多态性引物扩增蜀恢527／D64B的F2早熟和迟熟近等基因池,找到多态引物RM279,进一步用RM279附近的微卫星引物扩增F2早熟和迟熟近等基因池、迟熟植株,筛到多态性引物RM71。用MAPMAKER／EXP3．0软件分析,将该早熟基因定位于第2染色体的短臂端,位于RM179和RM71之间,遗传距离分别为12．6cM和13．3cM,该基因拟名EF-3(t)。在育种实践中用D64B育成早熟不育系D64A。     

水稻米粒延伸性的遗传剖析

以籼稻ZYQ8与粳稻JX17为亲本的DH群体作为研究材料,考察DH群体及双亲的米粒延伸率相关性状,并使用该群体的分子连锁图谱进行QTL分析.共检测到14个与稻米延伸性有关的QTL,包括2个粒长QTL、7个饭粒长QTL和5个米粒延伸率QTL,分别位于第1、2、3、5、6、7、10、11和12染色体.所有QTL的LOD值介于2.26～9.25,分别解释性状变异的5.31%～17.21%.在第3染色体上的G249～G164、第6染色体上的G30～RZ516和第10染色体上的G1082～GA223区间同时检测到控制饭粒长和米粒延伸率的QTL.米粒延伸性受多基因控制,Wx基因与位于第6染色体上的qCRE-6的G30～RZ516区间相近,对米饭的延伸性具重要影响.     

RFLP mapping of the genes controlling hybrid breakdown in rice (Oryza sativa L.)

 Complementary recessive genes hwd1 and hwd2 controlling hybrid breakdown (weakness of F₂ and later generations) were mapped in rice using RFLP markers. These genes produce a plant that is shorter and has fewer tillers than normal plants when the two loci have only one or no dominant allele at both loci. A cultivar with two dominant alleles at the hwd1 locus and a cultivar with two dominant alleles at the hwd2 locus were crossed with a double recessive tester line. Linkage analysis was carried out for each gene independently in two F₂ populations derived from these crosses. hwd1 was mapped on the distal region of rice genetic linkage map for chromosome 10, flanked by RFLP markers C701 and R2309 at a distance of 0.9 centiMorgans (cM) and 0.6 cM, respectively. hwd2 was mapped in the central region of rice genetic linkage map for chromosome 7, tightly linked with 4 RFLP markers without detectable recombination. The usefulness of RFLP mapping and map information for the genes controlling reproductive barriers are discussed in the context of breeding using diverse rice germplasm, especially gene introduction by marker-aided selection.     

不同环境条件下稻米透明度的发育遗传分析

采用条件和非条件数量遗传分析方法,研究了不同环境条件下灿稻稻米透明度的发育遗传规律。稻米4个发育时期两年数据的分析结果表明,开花受精后第8～21天的灌浆中期和后期遗传效应表达对稻米透明度的影响最大。控制稻米透明度性状表现的三倍体胚乳核基因、细胞质基因和二倍体母体植株核基因效应在不同环境下存在着表达水平上的差异。条件遗传分析的结果还表明一些遗传效应存在着发育时期间断表达的现象。基因加性效应和细胞质效应以及相应的环境互作效应在稻米透明度性状发育中起着主要作用,选择可以取得良好的改良效果。浙南3号和1391等亲本可以提高后代的稻米透明度。