Genetic structure and diversity of Oryza sativa L. in Guizhou, China

Preserving many kinds of rice resources and rich variations, Guizhou Province is one of the districts with the highest genetic diversity of cultivated rice （Oryza sativa L.） in China. In the current research, genetic diversity and structure of 537 accessions of cultivated rice from Guizhou were studied using 36 microsetellite markers and 39 phenotypic characters. The results showed that the model-based genetic structure was the same as genetic-distance-based one using SSRs but somewhat different from the documented classification （mainly based on phenotype） of two subspecies. The accessions being classified into indica by phenotype but japonica by genetic structure were much more than that being classified into japonica by phenotype but indica by genetic structure. Like Ding Ying＇s taxonomic system of cultivated rice, the subspecific differentiation was the most distinct differentiation within cultivated rice. But the differentiation within indica or japonica population was different： japonica presented clearer differentiation between soil-watery ecotypes than indica, and indica presented clearer differentiation between seasonal ecotypes than japonica. Cultivated rices in Guizhou revealed high genetic diversity at both DNA and phenotypic levels. Possessing the highest genetic diversity and all the necessary conditions as a center of genetic diversity, region Southwestern of Guizhou was suggested as the center of genetic diversity of O. sativa L. from Guizhou.     

Efficient indica and japonica rice identification based on the InDel molecular method: Its implication in rice breeding and evolutionary research

An efficient molecular method for the accurate and efficient identification of indica and japonica rice was created based on the polymorphisms of insertion/deletion (InDel) DNA fragments obtained from the basic local alignment search tool (BLAST) to the entire genomic sequences of indica (93-11) and japonica rice (Nipponbare). The 45 InDel loci were validated experimentally by the polymerase chain reaction (PCR) and polyacrylamide gel electrophoresis (PAGE) in 44 typical indica and japonica rice varieties, including 93-11 and Nipponbare. A neutrality test of the data matrix generated from electrophoretic banding patterns of various InDel loci indicated that 34 InDel loci were strongly associated with the differentiation of indica and japonica rice. More extensive analyses involving cultivated rice varieties from 11 Asian countries, and 12 wild Oryza species with various origins confirmed that indica and japonica characteristics could accurately be determined via calculating the average frequency of indica- or japonica-specific alleles on different InDel loci across the rice genome. This method was named as the “InDel molecular index” that combines molecular and statistical methods in determining the indica and japonica characteristics of rice varieties. Compared with the traditional methods based essentially on morphology, the InDel molecular index provides a very accurate, rapid, simple, and efficient method for identifying indica and japonica rice. In addition, the InDel index can be used to determine indica or japonica characteristics of wild Oryza species, which largely extends the utility of this method. The InDel molecular index provides a new tool for the effective selection of appropriate indica or japonica rice germplasm in rice breeding. It also offers a novel model for the study of the origin, evolution, and genetic differentiation of indica and japonica rice adapted to various environmental changes.     

Genetic diversity, geographic differentiation and evolutionary relationship among ecotypes of Glycine max and G. soja in China

The knowledge of origin and evolution of cultivated soybeans is one of the basic issues in both biology and agronomy of the crop. In order to investigate the nuclear and cytoplasmic genetic diversity, geographic differentiation and genetic relationship among geographic ecotypes of cultivated （Glycine max） and wild （G. soja） soybeans, the allelic profiles at 60 nuclear simple-sequence repeat （nuSSR） loci and 11 chloroplastic SSR （cpSSR） loci evenly distributed on whole genome of 393 landraces and 196 wild accessions from nation-wide growing areas in China were analyzed. （i） The genetic diversity of the wild soybean was obviously larger than that of the cultivated soybean, with their nuSSR and cpSSR alleles as 1067 vs. 980 and 57 vs 44, respectively. Of the 980 nuclear alleles detected in the cultivated soybean, 377 new ones （38.5%） emerged, while of the 44 chloroplastic alleles in the cultivated soybean, seven new ones （15.9%） emerged after domestication. （ii） Among the cultivated geographic ecotypes, those from southern China, including South-Central China, Southwest China and South China possessed relatively great genetic diversity than those from northern China, while among the wild geographic ecotypes, the Middle and Lower Changjiang Valleys wild ecotype showed the highest genetic diversity. （iii） The analysis of molecular variance, association analysis between geographic grouping and molecular marker clustering and analysis of specific-present alleles of ecotypes demonstrated that the geographic differentiation of both cultivated and wild soybeans associated with their genetic differentiation, or in other words, had their relevant genetic bases. （iv） The cluster analysis of all accessions clearly showed that the wild accessions from Middle and Lower Changjiang Valleys and South-Central ＆ Southwest China had relatively small genetic distances with all cultivated accessions. The UPGMA dendrogram among geographic ecotypes further showed that the genetic distances between all cultivated ecotypes and the Middle and Lower Changjiang Valleys wild ecotype were smaller than those with other wild ones, including their local wild counterparts. Therefore, it is inferred that the wild ancestors in southern China, especially those from Middle and Lower Changjiang Valleys might be the common ancestor of all the cultivated soybeans.     

Advances in the understanding of inter-subspecific hybrid sterility and wide-compatibility in rice

Hybrid sterility is a major form of postzygotic reproductive isolation and frequently occurs in hybrids between divergent populations, such as the indica and japonica subspecies of Asian cultivated rice （Oryza sativa L.）. It has been a major barrier for utilization of the strong heterosis expressed in hybrids between indica and japonica. A large number of loci for rice inter-subspecific hybrid sterility have been identified by genetic analysis. Cytological studies revealed that male and female gamete abortions and reduced affinity between the uniting gametes all occurred in indica-japonica hybrids, suggesting the complexity of the causes for inter-subspecific hybrid sterility. Two genes conditioning embryo-sac and pollen sterility respectively in indica-japonica hybrids have been cloned recently, providing opportunities for molecular characterization of the indica-japonica hybrid sterility and wide-compatibility. Future studies should aim at cloning more genes for indica-japonica hybrid sterility, characterizing the underlying molecular mechanism, and utilization of the findings for the development of inter-subspecific hybrids to increase rice productivity.     

Experimental validation of inter-subspecific genetic diversity in rice represented by the differences between the DNA sequences of ＇Nipponbare＇ and ＇93-11＇

The pedigrees of three sequenced rice cultivars were analyzed to show that a majority of the genetic composition of ＇Nipponbare＇ originates from japonica cultivars while the minority originates from indica cultivars. In contrast, ＇93-11＇ is derived mainly from indica cultivars with a smaller contribution from japonica cultivars. All ancestors of ＇Guang lu ai 4＇ appeared to be indica lines. A set of molecular markers （46 InDels and 53 SSRs） polymorphic between ＇Nipponbare＇ and ＇93-11＇ were examined in 46 typical indica and 47 typical japonica cultivars selected from 443 accessions according to Cheng＇s index. All cultivars were divided into indica and japonica groups without overlapping when clustered by Cheng＇s index, InDels and SSRs. Much higher InDel and SSR diversity between groups than within groups implies that the marker polymorphisms between ＇Nipponbare＇ and ＇93-11＇ represent a large proportion of inter-subspecific diversity. About 85% of indica cultivars and more than 90% of japonica cultivars were confirmed to have the same PCR banding patterns as ＇93-11＇ and ＇Nipponbare＇, respectively. Some polymorphic loci between ＇Nipponbare＇ and ＇93-11＇ cannot be validated in other indica and japonica cultivars, either as subspecies-specific but not predominant alleles, or alleles not specific between the two groups. It was concluded that molecular markers developed from sequence polymorphism between ＇Nipponbare＇ and ＇93-11＇ often represent inter-subspecific diversity, although some exceptions were sensitive to either particular marker loci or particular cultivars.     

High-quality reference genes for quantifying the transcriptional responses of Oryza sativa L. (ssp. indica and japonica) to abiotic stress conditions

Rice (Oryza sativa L.) is important to food security and is also an excellent model plant for numerous cereal crops. A functional genomics study in rice includes characterization of the expression dynamics of genes by quantitative real-time PCR (qPCR) analysis; this is a significant key for developing rice varieties that perform well in the face of adverse climate change. The qPCR analysis requires the use of appropriate reference genes in order to make any quantitative interpretations meaningful. Here, the new potential reference genes were selected from a huge public database of rice microarray experiments. The expression stability of 14 candidates and 4 conventional reference genes was validated by geNorm PLUS and NormFinder software. Seven candidates are superior to the conventionally used reference genes in qPCR and three genes can be used reliably for quantitating the expression of genes involved in abiotic stress responses. These high-quality references EP (LOC_Os05g08980), HNR (LOC_Os01g71770), and TBC (LOC_Os09g34040) worked very well in three indica genotypes and one japonica genotype. One of indica genotypes including the Jasmine rice, KDML105 developed in Thailand for which no reference genes have been reported until now.     

Effects of elevated [CO2] on stem and root lodging among rice cultivars

Studies showed that elevated [CO2] would improve photosynthetic rates and enhance yields of rice;however,few studies have focused on the response of rice lodging,which is a major cause of cereal yield loss and quality reduction,under elevated [CO2].In this study,we examined the effects of elevated [CO2] on stem and root lodging using 4 rice cultivars(86Y8,japonica hybrid;LYP9,2-line indica hybrid;variety 9311,type of indica inbred rice,and SY63,3-line indica hybrid) grown under two [CO2] levels:400 and 680 μmol mol-1.Our results indicated that under elevated [CO2],the stem-lodging risk(SLR) of 9311 decreased,while in SY63 the SLR increased,86Y8 and LYP9 were not significantly affected;the risk of root lodging was reduced for all cultivars,because root biomass(instead of root number) and bending strength were increased significantly,and then the increase of anti-lodging ability is far higher than that of self-weight mass moment for all cultivars.These findings suggested that higher [CO2] can enhance the risk of stem-lodging for cultivars with strong-[CO2]-responses,but may not aggravate the root lodging for all rice cultivars.     

SSR标记对籼稻品种的遗传多样性分析

利用500多对SSR标记对广陆矮4号、珍汕97B、佳辐占、明恢86和明恢63 5个籼稻亲本品种进行遗传多样性分析.结果表明,佳辐占与明恢63、明恢86、珍汕97B和广陆矮4号之间的相似系数分别为78.1%、76.1%、72.8%和71.7%.相对而言,其相似系数越小,遗传距离越大,因此利用佳辐占与这些品种组配较有希望选育出品质优良、综合优势明显的新品系.同时研究表明,标记间平均距离为10 cM或小于10 cM时,分析所得结果较为可靠.     

Genetic analysis and fine mapping of the pubescence gene GL6 in rice (Oryza sativa L.)

The pubescence of the leaf blade surface is an important agronomic characteristic for rice morphology and significantly influences rice growth as well as physiological characteristics. This characteristic was analyzed in F1 and F2 plants derived by crossing cultivar 75-1-127 with the indica cultivar Minghui 63, as well as the glabrous cultivar Lemont and indica cultivar 9311. Results indicated that the pubescence of the leaf blade surface was a dominant trait and controlled by a single gene. The GL6 gene was primarily mapped on rice chromosome 6 with recessive F2 population derived from 75-1-127/Minghui 63 by combining bulked segregation analysis and recessive class analysis using the Mapmaker3.0/MapDraw software. The genetic distances between the simple sequence repeat markers RM20491 and RM20547 were 7.2 and 2.2 cM, respectively. The GL6 gene was fine mapped in the interval between InDel-106 and InDel-115 at genetic distances of 0.3 and 0.1 cM, respectively. The large, recessive F2 population was derived from 75-1-127/Minghui 63. A high-resolution genetic and physical map of GL6 was constructed. Derived from the map-based sequences published by the International Rice Genome Sequencing Project, the GL6 gene was localized at an interval of 79 (japonica) and 116.82 kb (9311) bracketed by InDel-106 and InDel-115 within the BAC accession numbers AP008403 and AP005760. Seven annotated genes (japonica) and eight annotated genes (9311) were present. The basis was further set for GL6 cloning and function analysis.     

The genetic diversity of Gaozhou wild rice analyzed by SSR

LOCATED AT 21°42′34″―22°18′48″N AND 110°36′48″― 111°22′45″E IN THE TRANSITION REGION FROM SOUTH SUB- TROPICS TO NORTH SUBTROPICS, GAOZHOU WILD RICE (GWR) IS COMMON WILD RICE (ORYZA RUFIPOGON GRIFF.) AND THE WIDEST WILD RICE POPULATION IN GUA     

Genetic diversity and construction of core collection in Chinese wheat genetic resources

Genetic diversity among 5029 accessions representing a proposed Chinese wheat core collection was analyzed using 78 pairs of fluorescent microsatellite （SSR） primers mapped to 21 chromosomes. A stepwise hierarchical sampling strategy with priority based on 4×10^5 SSR data-points was used to construct a core collection from the 23090 initial collections. The core collection consisted of 1160 accessions, including 762 landraces, 348 modern varieties and 50 introduced varieties. The core accounts for 23.1% of the 5029 candidate core accessions and 5% of the 23090 initial collections, but retains 94.9% of alleles from the candidate collections and captures 91.5% of the genetic variation in the initial collections. These data indicate that it is possible to maintain genetic diversity in a core collection while retaining fewer accessions than the accepted standard, i.e., 10% of the initial collections captured more than 70% of their genetic diversity. Estimated genetic representation of the core constructed by preferred sampling （91.5%） is much higher than that by random sampling （79.8%）. Both mean genetic richness and genetic diversity indices of the landraces were higher than those of the modern varieties in the core. Structure and principal coordinate analysis revealed that the landraces and the modern varieties were two relatively independent subpopulaUons. Strong genetic differentiation associated with ecological environments has occurred in the landraces, but was relatively weak in the modern culUvars. In addition, a mini-core collection was constructed, which consisted of 231 accessions with an estimated 70% representation of the genetic variation from the initial collections. The mini-core has been distributed to various research and breeding institutes for detailed phenotyping and breeding of genetic introgression lines.     

利用ISSR分子标记研究樱桃李野生类型的遗传多样性及亲缘关系

利用ISSR分子标记技术对产于中国天山的40个樱桃李(Prunus divaricata Ldb.)野生类型遗传多样性进行研究。通过遗传多样性研究及UPGMA聚类分析显示,野生樱桃李种下类群间遗传多样性有差异,平均多样性指数(I)为0.488,遗传相似系数(Hs)介于0.504~0.851之间,平均为0.678,遗传距离(GD)的范围在0.127 0~0.604 2之间,等位基因数(Na)和有效等位基因数(Ne)分别为1.908和1.571,而平均Nei基因多样性指数为0.17,平均Shannon信息指数(I)为0.22则显示其群体遗传多样性较低;分子标记下的聚类结果,可将中国天山产的40个樱桃李野生类型归为4大类,结合形态学特征分析,研究表明中国天山的40个野生樱桃李可划分为4个变种,即矮生樱桃李P.divaricata var.humila(N.R.Cui et L.Wang)D.F.Cui,comb.nov.、黑果樱桃李P.divaricata var.nigrooarp(N.R.Cui et L.Wang)D.F.Cui,comb.nov.、红果樱桃李P.divaricata var.phoenicea(Z.Xu)D.F.Cui,comb.nov.和黄果樱桃李P.divaricata var.luteola(N.R.Cui)D.F.Cui,comb.nov.。     

Introduction of antisensewaxy gene into main parent lines ofindica hybrid rice

Amylose content in rice endosperm is one of the key determinants of rice eating and cooking quality, and the poor quality ofindica hybrid rice is closely related to the high amylose level in rice grains. In order to improve the grain quality of theindica hybrid rice by genetic engineering, an antisense fragment of ricewaxy gene, driven by the 5′-franking sequences of the ricewaxy gene, was successfully introduced into three major parent lines ofindica hybrid rice, all contain a high amylose level in the grains, viaAgrobacterium, and more than 100 hygromycinresistant plants were regenerated. The analysis of PCR amplification and Southern blots indicated that the T-DNA containing the antisensewaxy gene had been integrated into the genome of transgenic rice plants. Most of the primary transgenic rice plants grew normally, and the mature seeds from these transgenic plants were performed for analysis of the amylose content. The results showed that the amylose content in the endosperm of some grains was reduced and the lowest reached 7.02% in one homozygous transgenic line, 72.4% lower than that of the wild type. The influence of the altered amylose content on the gelatinization temperature and gel consistency was also observed in several homozygous transgenic rice plants. The two authors contributed equally to this work.     

ISSR markers based on GA and AG repeats reveal genetic relationship among rice varieties tolerant to drought, flood, or salinity

Drought, flood, salinity, or a combination of these limits rice production. Several rice varieties are well known for their tolerance to specific abiotic stresses. We determined genetic relationship among 12 rice varieties including 9 tolerant to drought, flood, or salinity using inter-simple sequence repeat (ISSR) markers. Based on all markers, the nine tolerant varieties formed one cluster distinct from the cluster of three control varieties. The salt-tolerant varieties were closest to two flood-tolerant varieties, and together they were distinct from the drought-tolerant varieties. (GA)₈YG was the most informative primer, showing the highest polymorphic information content (PIC) and resolving power (Rp). The drought-, flood-, and salt-tolerant varieties grouped in three distinct clusters within the group of tolerant varieties, when (GA)₈YG was used. Sabita was the only exception. The two aus varieties, Nagina22 and FR13A, were separated and grouped with the drought- and flood-tolerant varieties, respectively, but they were together in dendrograms based on other primers. The results show that ISSR markers associated with (GA)₈YG delineated the three groups of stress-tolerant varieties from each other and can be used to identify genes/new alleles associated with the three abiotic stresses in rice germplasm.     

Geographical genetic diversity and divergence of common wild rice （O. rufipogon Griff.） in China

Using 36 SSR markers and 889 accessions of common wild rice in China, the genetic diversity and the divergence among different geographical populations are investigated. Guangdong Province has the largest number of alleles, which account for 84% of the total alleles detected in the study, followed by Guangxi Province. The Nei＇s gene diversity indices, from high to low, are in the sequence of Hainan, Guangdong, Guangxi, Fujian, Hunan, Jiangxi, and Yunnan provinces. Two genetic diversity centers of Chinese common wild rice are detected on the basis of geographic analysis, i.e., the region covering Boluo, Zijin, Lufeng, Haifeng, Huidong and Huiyang counties of Guangdong Province and the region covering Yongning, Longan, Laibin and Guigang counties of Guangxi Province. The common wild rice in Yunnan, Hunan, Jiangxi, and Fujian provinces are diverged into respectively independent populations with relatively large genetic distances, whereas, those in Hainan, Guangdong and Guangxi provinces have relatively low genetic divergence. Under the condition of geographic separation, natural selection is considered as one of the primary forces contributing to the divergence of common wild rice in China.     

Molecular evolution of rice S 5 n and functional comparison among different sequences

The wide compatibility gene, S ₅ ⁿ , can overcome embryo sac sterility between indica and japonica subspecies of rice. Therefore, it is very important to characterize the features of the S ₅ ⁿ sequence to reveal the origin and evolution of S ₅ ⁿ . In this paper, 26 cultivated rice haplotypes and 22 wild rice accessions harboring S ₅ ⁿ were used to sequence S ₅ ⁿ . The results showed that 15 genotypes among the 48 materials were fully consistent with control cultivar 02428 (CK). The other 33 accessions had different degrees of variation in the S ₅ ⁿ sequence. Variations in the coding region mainly occurred in the second exon and eight materials showed a 10-bp deletion at 1710?C1719 bp, including wild (O. nivara) and cultivated rice, such as IRW501 and Yuetai B. S ₅ ⁿ sequences were not biased and evolved neutrally. The 48 materials could be divided into 4 categories using a phylogenetic tree of the amino acid sequences. Most of the wild rice clustered together, and the cultivated rice clustered into another group. Eight cultivated rice and O. nivara (wild rice) clustered in another group, which were found to lack 10 consecutive bases in exon 2. Eight rice varieties with high numbers of differences in their S ₅ ⁿ coding regions were crossed with testers (typically indica and japonica) to produced test cross F₁ populations. The F₁s were examined for their ability to overcome indica-japonica hybrid sterility. The result showed that the embryo sac fertility of S ₅ ⁿ -containing hybrids increased significantly compared with control hybrids, but there were no differences among the materials with divergent sequences, indirectly proving that S ₅ ⁿ is a non-functional gene.     

基于InDel分子标记的20个水稻品种的籼粳属性分析

水稻品种的籼粳属性分析是培育高产优质杂交水稻亲本选择的重要依据之一。本研究以93-11与日本晴分别为籼粳稻参考标准,利用均匀分布于水稻基因组12条染色体并具有较高籼粳特异性的37对In Del分子标记,对20份水稻育种材料进行In Del标记的籼粳属性分析。结果表明:所选取的20份水稻材料中,有8份为典型籼稻,3份为典型粳稻,4份为偏籼类型,另有5份为偏粳类型。NTsys聚类分析结果显示,在遗传相似系数为0.43处,20份水稻材料被分为两大类群;在遗传相似数为0.70处,所有水稻材料则被分为4个类群。     

Genetic diversity of rice cultivars （Oryza sativa L.） in China and the temporal trends in recent fifty years

Public concern is often expressed at cultivars because the domestication and modern plant breeding have led to a reduction in the genetic diversity of crops and loss of genes, which could result in crops' genetic vulnerability to changes in the spectrum of pestssity of varieties in this zone is very important to the whole rice production in China. REZV, a important japonica rice production areas with more than 278 thousands ha rice which was about 71% of rice area in north China, accounted fo…     

A draft sequence of the rice (Oryza sativa ssp. indicd) genome

The sequence of the rice genome holds fundamental information for its biology, including physiology, genetics, development, and evolution, as well as information on many beneficial phenotypes of economic significance. Using a “whole genome shotgun” approach, we have produced a draft rice genome sequence ofOryza sativa ssp.indica, the major crop rice subspecies in China and many other regions of Asia. The draft genome sequence is constructed from over 4.3 million successful sequencing traces with an accumulative total length of 2214.9 Mb. The initial assembly of the non-redundant sequences reached 409.76 Mb in length, based on 3.30 million successful sequencing traces with a total length of 1797.4 Mb from anindica variant cultivar93-11, giving an estimated coverage of 95.29% of the rice genome with an average base accuracy of higher than 99%. The coverage of the draft sequence, the randomness of the sequence distribution, and the consistency of BIG-ASSEMBLER, a custom-designed software package used for the initial assembly, were verified rigorously by comparisons against finished BAC clone sequences from bothindica andjapanica strains, available from the public databases. Over all, 96.3% of full-length cDNAs, 96.4% of STS, STR, RFLP markers, 94.0% of ESTs and 94.9% unigene clusters were identified from the draft sequence. Our preliminary analysis on the data set shows that our rice draft sequence is consistent with the comman standard accepted by the genome sequencing community. The unconditional release of the draft to the public also undoubtedly provides a fundamental resource to the international scientific communities to facilitate genomic and genetic studies on rice biology. These authors contributed equally to this work.     

分光光度法检测不同地区和类型稻种功能性成分的初步研究

 为研究不同地区和类型水稻品种糙米中γ-氨基丁酸和总黄酮等功能性成分,本实验采用分光光度法准确测定了中国、日本、韩国、国际稻作所和云南品种及云南野生稻γ-氨基丁酸和总黄酮的含量.结果表明:① 用分光光度法测定水稻籽粒中γ-氨基丁酸和总黄酮含量的方法是可靠的;② 不同地区和类型水稻品种功能成分含量存在差异;③ 相关分析表明,氨基丁酸与总黄酮的相关系数较高(r=0.575^**),在育种上可以实现多个相关性状间的遗传改良;④ 鉴定出墨江黄皮饵块谷、明恢86、墨江饵块谷、疣粒野生稻等18份高GABA和高总黄酮的水稻品种,可作为选育高黄酮和γ-氨基丁酸品种的亲本或直接作为功能稻开发利用.首次报道疣粒野生稻总黄酮质量分数达0.751%,在所有水稻品种中黄酮含量最高.