Analysis of Starch Gene Diversity in the Wild Relatives of Oryza sativa

Genetic loci influencing traits important to humans have been selected during crop domestication. The starch properties of rice influence the ease of cooking and attractiveness of rice as a human food. Starch biosynthesis genes likely to influence starch properties in the grain were compared in wild and domesticated rice genotypes. Sequence variation was investigated in starch biosynthesis gene exons that have been reported to have a direct influence on rice amylose content, gelatinization temperature, and amylopectin chain length. Exons 6 and 10 of GBSSI, exon 8 of SSIIa and exons 11, 13, 14 and 16 of SBEIIb were amplified and sequenced from 13 wild Oryza species encompassing genome types AA to HHJJ. Thirty two single nucleotide polymorphisms (SNPs) were identified in the exons of GBSSI; 176 in exon 8 of SSIIa, and 43 in SBEIIb, giving a total of 251 SNPs among the species. Eighty six of these SNP caused changes in the encoded amino acid, of which 28 were missense mutations that affected highly conserved amino acids within the protein sequence of GBSSI, SSIIa or SBEIIb. Two indels were identified in Potamophila parviflora, a close relative of Zizania palustris, a North American native wild rice. Most of the nucleotide variations and non-conservative changes were observed in the genomes other than the AA genome species. This represents a genetic resource for use in rice starch manipulation. The impact of human selection at these loci can be deduced by comparison of modern cultivated genotypes with their wild progenitors.     

Genomic structure and evolution of the Pi2/9 locus in wild rice species

Rice blast, caused by the fungal pathogen Magnaporthe oryzae, is a devastating disease of rice worldwide. Among the 85 mapped resistance (R) genes against blast, 13 have been cloned and characterized. However, how these genes originated and how they evolved in the Oryza genus remains unclear. We previously cloned the rice blast R-genes Pi2, Pi9, and Piz-t, and analyzed their genomic structure and evolution in cultivated rice. In this study, we determined the genomic sequences of the Pi2/9 locus in four wild Oryza species representing three genomes (AA, BB and CC). The number of Pi2/9 family members in the four wild species ranges from two copies to 12 copies. Although these genes are conserved in structure and categorized into the same subfamily, sequence duplications and subsequent inversions or uneven crossing overs were observed, suggesting that the locus in different wild species has undergone dynamic changes. Positive selection was found in the leucine-rich repeat region of most members, especially in the largest clade where Pi9 is included. We also provide evidence that the Pi9 gene is more related to its homologues in the recurrent line and other rice cultivars than to those in its alleged donor species O. minuta, indicating a possible origin of the Pi9 gene from O. sativa. Comparative sequence analysis between the four wild Oryza species and the previously established reference sequences in cultivated rice species at the Pi2/9 locus has provided extensive and unique information on the genomic structure and evolution of a complex R-gene cluster in the Oryza genus.     

Variation in leaf photosynthetic characteristics in wild rice species

Variations in leaf gas-exchange characteristics, leaf pigment content, and other important leaf traits were investigated in seven wild Oryza species, five hybrids, and five improved varieties. The significant variations were observed in photosynthetic pigment contents amongst different species of Oryza. The mean chlorophyll (Chl) content was higher in O. sativa (varieties and hybrids), while O. eichengeri showed the lowest Chl content. The mean carotenoid (Car) content in O. sativa (varieties and hybrids) was higher than in other wild rice species. O. eichengeri and O. barthii had significantly lower Car contents than other rice species. Significant differences were noticed in the rate of photosynthesis (P _N), stomatal conductance (g _s), transpiration rate (E), internal CO₂ concentration (C _i), specific leaf mass (SLM), and leaf thickness amongst different Oryza species. The mean P _N was the highest in O. nivara followed by O. eichengeri. The mean P _N was the lowest in O. glumaepatula, which was lower than that of cultivated varieties and hybrids of O. sativa. High rates of photosynthesis were observed in O. nivara (ACC. No. CR 100097), O. rufipogon (ACC.No. CR 100267), and O. nivara (ACC.No. CR 100008). The O. nivara and O. rufipogon genotypes with high P _N might be used in rice improvement programmes for an increase of leaf photosynthesis in rice. Multiple correlations performed between different gas-exchange characteristics and other physiological traits revealed that the rate of photosynthesis was not dependent on the leaf pigment content or the leaf thickness. A strong positive correlation between P _N and the P _N/C_i ratio, which represents the carboxylation efficiency, indicated that the observed variation in P _N was not based on pigment content or other leaf traits.     

Polymorphism and phylogenetic relationships among species in the genus Oryza as determined by analysis of nuclear RFLPs

Summary Ninety-three accessions representing 21 species from the genus Oryza were examined for restriction fragment length polymorphism. The majority (78%) of the accessions, for which five individuals were tested, were found to be monomorphic. Most of the polymorphic accessions segregated for only one or two probes and appeared to be mixed pure lines. For most of the Oryza species tested, the majority of the genetic variation (83%) was found between accessions from different species with only 17% between accessions within species. Tetraploid species were found to have, on average, nearly 50% more alleles (unique fragments) per individual than diploid species reflecting the allopolyploid nature of their genomes.Classification of Oryza species based on RFLPs matches remarkably well previous classifications based on morphology, hybridization and isozymes. In the current study, four species complexes could be identified corresponding to those proposed by Vaughan (1989): the O. ridleyi complex, the O. meyeriana complex, the O. officinalis complex and the O. sativa complex. Within the O. sativa complex, accessions of O. rufipogon from Asia (including O. nivara) and perennial forms of O. rufipogon from Australia clustered together with accessions of cultivated rice O. sativa. Surprisingly, indica and japonica (the two major subspecies of cultivated rice) showed closer affinity with different accessions of wild O. Rufipogon than to each other, supporting a hypothesis of independent domestication events for these two types of rice. Australian annual wild rice O. meridionalis (previously classified as O. rufipogon) was clearly distinct from all other O. rufipogon accessions supporting its recent reclassification as O. meridionalis (Ng et al. 1981). Using genetic relatedness as a criterion, it was possible to identify the closest living diploid relatives of the currently known tetraploid rice species. Results from these analyses suggest that BBCC tetraploids (O. malampuzhaensis, O. punctata and O. minuta) are either of independent origins or have experienced introgression from sympatric C-genome diploid rice species. CCDD tetraploid species from America (O. latifolia, O. alta and O. grandiglumis) may be of ancient origin since they show a closer affinity to each other than to any known diploid species. Their closest living diploid relatives belong to C genome (O. eichingeri) and E genome (O. Australiensis) species. Comparisons among African, Australian and Asian rice species suggest that Oryza species in Africa and Australia are of polyphyletic origin and probably migrated to these regions at different times in the past.Finally, on a practical note, the majority of probes used in this study detected polymorphism between cultivated rice and its wild relatives. Hence, RFLP markers and maps based on such markers are likely to be very useful in monitoring and aiding introgression of genes from wild rice into modern cultivars.     

The Oryza Map Alignment Project: The Golden Path to Unlocking the Genetic Potential of Wild Rice Species

The wild species of the genus Oryza offer enormous potential to make a significant impact on agricultural productivity of the cultivated rice species Oryza sativa and Oryza glaberrima. To unlock the genetic potential of wild rice we have initiated a project entitled the ‘Oryza Map Alignment Project’ (OMAP) with the ultimate goal of constructing and aligning BAC/STC based physical maps of 11 wild and one cultivated rice species to the International Rice Genome Sequencing Project’s finished reference genome – O. sativa ssp. japonica c. v. Nipponbare. The 11 wild rice species comprise nine different genome types and include six diploid genomes (AA, BB, CC, EE, FF and GG) and four tetrapliod genomes (BBCC, CCDD, HHKK and HHJJ) with broad geographical distribution and ecological adaptation. In this paper we describe our strategy to construct robust physical maps of all 12 rice species with an emphasis on the AA diploid O. nivara – thought to be the progenitor of modern cultivated rice.     

Assessment of genome relationships in the genus Oryza L. based on seed-protein profile analysis

Summary Cultivated and wild Oryza species belonging to different genomic groups were studied with regard to their soluble seed-protein profiles. There is an essential uniformity in the banding patterns within various genomes and the basic patterns are not species-specific but genome-specific. O. meridionalis contains a subgenome similar to the A genome of O. rufipogon. Certain specific bands present among A genome species have been found to be useful in tracing the phylogenetic affinity between the cultivated species and their presumed wild progenitors.     

Genome-Wide Analysis in Wild and Cultivated <Emphasis Type="Italic">Oryza</Emphasis> Species Reveals Abundance of NBS Genes in Progenitors of Cultivated Rice

NBS-encoding genes play a critical role in the plant defense system. Wild relatives of crop plants are rich reservoirs of plant defense genes. Here, we performed a stringent genome-wide identification of NBS-encoding genes in three cultivated and eight wild Oryza species, representing three different genomes (AA, BB, and FF) from four continents. A total of 2688 NBS-encoding genes were identified from 11 Oryza genomes. All the three progenitor species of cultivated rice, namely O. barthii, O. rufipogon, and O. nivara, were the richest reservoir of NBS-encoding genes (214, 313, and 307 respectively). Interestingly, the two Asian cultivated species showed a contrasting pattern in the number of NBS-encoding genes. While indica subspecies maintained nearly equal number of NBS genes as its progenitor (309 and 313), the japonica subspecies had retained only two third in the course of evolution (213 and 307). Other major sources for NBS-encoding genes could be (i) O. longistaminata since it had the highest proportion of NBS-encoding genes and (ii) O. glumaepatula as it clustered distinctly away from the rest of the AA genome species. The present study thus revealed that NBS-encoding genes can be exploited from the primary gene pool for disease resistance breeding in rice.     

Ribosomal gene spacer length variability in cultivated and wild rice species

Summary Restriction fragment length polymorphism of the rDNA spacer was studied in the genus Oryza using a cloned rice rDNA probe. One-hundred-five accessions, including 58 cultivated rice and 47 wild species with various genome types, were analysed. Seven size classes differing from one another by an increment of ca. 300 bp were observed amongst the Asiatic cultivated rice of the species O. sativa. A general tendency from a smaller spacer in the Japonica subtypes to longer ones in Indica is observed. Classification as Japonica or Indica on the basis of rDNA pattern generally agrees with classification based on isozyme patterns. In contrast, African rice of the species O. glaberrima does not display any rDNA size variation. When wild species are considered, extensive variation is observed, but the fragment sizes do not fall into regularly increasing size classes except for O. rufipogon and O. longistaminata. The variation is greater in these species than in the cultivated ones.     

Aminoacid profiles in the study of phylogenetic relationships in the genusOryza

Summary The amino acid profiles in seeds of thirteen different species ofOryza, including two cultivated rices,O. glaberrima andO. sativa and the two major geographical racesindica andjaponica were studied using an automatic amino acid analyser to assess differences in the profiles of cultivated species and their wild progenitors. The polygon graphic method was employed to envision the species relationship. Essential amino acid profiles in different species were also compared with those of the Food and Agriculture Organization (FAO) standards. The results suggest a wide range of variability amongOryza species for lysine (up to 4.4% as against 3.5% in cultivated rices) and other essential amino acids. This will be of considerable interest to rice breeders, when after overcoming genetic barriers, the possible utilization of these species in rice breeding becomes feasible.     

Phylogenetic Analysis of AA-genome Oryza Species (Poaceae) Based on Chloroplast, Mitochondrial, and Nuclear DNA Sequences

Species in the genus Oryza (Poaceae) contain 10 genomic types and are distributed in pan-tropics of the world. To explore phylogenetic relationships of Oryza species having the AA-genome, DNA sequences of the chloroplast trnL intron and trnL-trnF spacer, mitochondrial nad1 intron 2, and nuclear internal transcribed spacer were analyzed, based on materials from 6 cultivated (O. sativa and O. glaberrima) and 13 wild accessions, in addition to a CC-genome species (O. officinalis) that was used as an outgroup. Analyses of the combined sequence data set from different sources provide a much better resolution of the AA-genome species than the individual data set, indicating the limitation of a single gene in phylogenetic reconstruction. The phylogeny based on the combined data set demonstrated an apparent grouping of the AA-genome Oryza species that was well associated with their geographic origin, although the Australian O. meridionalis showed its affinity with the African species. The geographic pattern of the phylogenetic relationship was probably attributed to the frequent genetic exchange and introgression among the AA-genome species from the same continents. In addition, Asian cultivated rice O. sativa showed its close relation to O. rufipogon and O. nivara, whereas African cultivated rice O. glaberrima was closely linked to O. barthii and O. longistaminata, indicating the independent domestication of the two cultivated species in different geographic locations.     

Genetic relationship of Porteresia coarctata Tateoka using molecular markers

ABSTRACT

Twenty-one species belonging to Oryza, including wild rices, were compared with a tetraploid (2n=48) halophytic wild rice relative, Porteresia coarctata Tateoka (=Oryza coarctata) for the genetic relatedness using AFLP and RAPD markers. Diploid and tetraploid groups were clearly separated except in the case of a few species where the clustering was unique and different. The molecular analysis has helped in positioning Porteresia in the vicinity of other wild rice species, and to better understand the pattern of species differentiation in Oryza. From our study, O. australiensis seems to be related to P. coarctata; thus, O. australiensis may be an effective “bridge” species in transferring genetic traits from P. coarctata to O. sativa. The usefulness of molecular marker systems for studying polymorphism and classification, and in clarifying genetic relationships between wild species has been confirmed.     

Gas exchange,carbon balance and stomatal traits in wild and cultivated rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) genotypes

Carbon balancing within the plant species is an important feature for climatic adaptability. Photosynthesis and respiration traits are directly linked with carbon balance. These features were studied in 20 wild rice accessions Oryza spp., and cultivars. Wide variation was observed within the wild rice accessions for photosynthetic oxygen evolution or photosynthetic rate (A), dark (R _d), and light induced respiration (LIR) rates, as well as stomatal density and number. The mean rate of A varied from 10.49 μmol O₂ m^?2 s^?1 in cultivated species and 13.09 μmol O₂ m^?2 s^?1 in wild spp., The mean R _d is 2.09 μmol O₂ m^?2 s^?1 and 2.31 μmol O₂ m^?2 s^?1 in cultivated and wild spp., respectively. Light induced Respiration (LIR) was found to be almost twice in wild rice spp., (16.75 μmol O₂ m^?2 s^?1) compared to cultivated Oryza spp., Among the various parameters, this study reveals LIR and A as the key factors for positive carbon balance. Stomatal contribution towards carbon balance appears to be more dependent on abaxial surface where several number of stomata are situated. Correlation analysis indicates that R _d and LIR increase with the increase in A. In this study, O. nivara (CR 100100, CR 100097), O. rufipogon (IR 103404) and O. glumaepatula (IR104387) were identified as potential donors which could be used in rice breeding program. Co-ordination between gas exchange and patchiness in stomatal behaviour appears to be important for carbon balance and environmental adaptation of wild rice accessions, therefore, survival under harsh environment.     

Diversity in the Content of Some Nutritional Components in Husked Seeds of Three Wild Rice Species and Rice Varieties in Yunnan Province of China

In addition to rice (Oryza sativa L.) cultivars, there are three wild rice species, namely O.rufipogon Griff, O. officinalis Wall and O. granulata Baill, in Yunnan Province, China. Each species has different subtypes and ecological distributions. Yunnan wild rice species are excellent genetic resources for developing new rice cultivars. The nutritional components of the husked seeds of wild rice have not been investigated thus far. Herein, we report on the contents of total protein, starch, amylose, 17 amino acids, and five macro and five trace mineral elements in husked seeds from three wild rice species and six O. sativa cultivars. The mean (± SD) protein content in the husked rice of O. rufipogon, O. officinalis, and O. granulata was (14.5 ± 0.6)%, (16.3 ± 1. 1)%, and (15.3 ± 0.5)%, respectively. O. officinalis Ⅲ originating from Gengma had the highest protein content (19.3%). In contrast, the average protein content of six O. sativa cultivars was only 9.15%. The total content of 17 amino acids of three wild rice species was 30%-50% higher than that of the six cultivars. Tyrosine, lysine, and valine content in the three wild rice species was 34%-209% higher than that of the cultivars. However, the difference in total starch content among different O. sativa varieties or types of wild rice species was very small. The average amylose content of O. rufipogon, O. officinalis,and O. granulata was 12.0%, 9.7%, and 11.3%, respectively, much lower than that of the indica and japonica varieties (14.37%-17.17%) but much higher than that of the glutinous rice cultivars (3.89%). The sulfur, phosphorus, magnesium, zinc, and ferrite content in the three wild rice species was 30%-158% higher than that of the six cultivars. The considerable difference in some nutritional components among wild rice species and O. sativa cultivars represents a wide biodiversity of Yunnan Oryza species. Based on the results of the present study, it is predicted that some good genetic traits, especially high protein and ideal amylose content, of Yunnan wild rice species may be useful in improving the nutritional value of rice. This is the first report regarding the amino acid, mineral element, protein and amylose content of husked seeds of some Yunnan wild rice species that have important genetic characteristics for rice quality and nutritional value.     

Tolerance to soil water stress by <Emphasis Type="Italic">Oryza sativa</Emphasis> cv. IR20 was improved by expression of <Emphasis Type="Italic">Wsi18</Emphasis> gene locus from <Emphasis Type="Italic">Oryza nivara</Emphasis>

Wild rice genotypes are rich in genetic diversity. This has potential to improve agronomic rice by allele mining for superior traits. Late embryogenesis abundant (LEA) proteins are often associated with desiccation tolerance and stress signalling. In the present study, a group 3 LEA gene, Wsi18 from the wild rice Oryza nivara was expressed under its own inducible promoter element in stress susceptible cultivated indica rice (cv. IR20). The resulting transgenic plants cultivated in a greenhouse showed enhanced tolerance to soil water deficit. Transgenic plants had higher grain yield, plant survival rate, and shoot relative water content compared to wild type (WT) IR20. Cell membrane stability index, proline and soluble sugar content were also greater in transgenic than WT plants under water stress. These results demonstrate the potential for improving SWS tolerance in agronomically important rice cultivar by incorporating Wsi18 gene from a wild rice O. nivara.     

New insights into <Emphasis Type="Italic">Oryza</Emphasis> genome evolution: high gene colinearity and differential retrotransposon amplification

An ∼247-kb genomic region from FF genome of wild rice Oryza brachyantha, possessing the smallest Oryza genome, was compared to the orthologous ∼450-kb region from AA genome, O. sativa L. ssp. japonica. 37 of 38 genes in the orthologous regions are shared between japonica and O. brachyantha. Analyses of nucleotide substitution in coding regions suggest the two genomes diverged ∼10 million years ago. Comparisons of transposable elements (TEs) reveal that the density of DNA TEs in O. brachyantha is comparable to O. sativa; however, the density of RNA TEs is dramatically lower. The genomic fraction of RNA TEs in japonica is two times greater than in O. brachyantha. Differences, particularly in RNA TEs, in this region and in BAC end sequences from five wild and two cultivated Oryza species explain major genome size differences between sativa and brachyantha. Gene expression analyses of three ObDREB1 genes in the sequenced region indicate orthologous genes retain similar expression patterns following cold stress. Our results demonstrate that size and number of RNA TEs play a major role in genomic differentiation and evolution in Oryza. Additionally, distantly related O. brachyantha shares colinearity with O. sativa, offering opportunities to use comparative genomics to explore the genetic diversity of wild species to improve cultivated rice. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Data deposition: Sequence data from this article were deposited with GenBank Library under accession number DQ810282. Shibo Zhang and Yong Qiang Gu contributed equally to the work     

Occurrence and immunological relationships of lectins in gramineous species

A survey of the occurrence of lectins in seeds from more than 100 grass species showed that all species belonging to the Triticeae tribe and the genera Brachypodium and Oryza contain lectins. All these lectins have the same sugar-binding specificity and are related to wheat-germ agglutinin, but to different degrees. Lectins from Triticeae species are immunologically indistinguishable from wheat lectin, whereas Brachypodium and rice lectins are only immunologically related to the wheat lectin. Attempts to detect lectin-deficient lines or varieties in wild and cultivated species of the three lectin-containing groups were unsuccessful. The possible use of lectins as a chemotaxonomic tool is discussed.     

Evolutionary trajectory of phytoalexin biosynthetic gene clusters in rice

Plants frequently possess operon‐like gene clusters for specialized metabolism. Cultivated rice, Oryza sativa, produces antimicrobial diterpene phytoalexins represented by phytocassanes and momilactones, and the majority of their biosynthetic genes are clustered on chromosomes 2 and 4, respectively. These labdane‐related diterpene phytoalexins are biosynthesized from geranylgeranyl diphosphate via ent‐copalyl diphosphate or syn‐copalyl diphosphate. The two gene clusters consist of genes encoding diterpene synthases and chemical‐modification enzymes including P450s. In contrast, genes for the biosynthesis of gibberellins, which are labdane‐related phytohormones, are scattered throughout the rice genome similar to other plant genomes. The mechanism of operon‐like gene cluster formation remains undefined despite previous studies in other plant species. Here we show an evolutionary insight into the rice gene clusters by a comparison with wild Oryza species. Comparative genomics and biochemical studies using wild rice species from the AA genome lineage, including Oryza barthii, Oryza glumaepatula, Oryza meridionalis and the progenitor of Asian cultivated rice Oryza rufipogon indicate that gene clustering for biosynthesis of momilactones and phytocassanes had already been accomplished before the domestication of rice. Similar studies using the species Oryza punctata from the BB genome lineage, the distant FF genome lineage species Oryza brachyantha and an outgroup species Leersia perrieri suggest that the phytocassane biosynthetic gene cluster was present in the common ancestor of the Oryza species despite the different locations, directions and numbers of their member genes. However, the momilactone biosynthetic gene cluster evolved within Oryza before the divergence of the BB genome via assembly of ancestral genes.     

Two new genomes in the Oryza complex identified on the basis of molecular divergence analysis using total genomic DNA hybridization

The genus Oryza to which cultivated rice belongs has 24 species (2n?=?24 or 48), representing seven genomes (AA, BB, CC, EE, FF, BBCC and CCDD). The genomic constitution of five of these species is unknown. These five species have been grouped into two species complexes, the tetraploid ridleyi complex (O. ridleyi, O.?longiglumis) and the diploid meyeriana complex (O.?granulata, O. meyeriana, O. indandamanica). To evaluate the genomic structure of these species in terms of divergence at the molecular level vis-à-vis other known genomes of Oryza, we used the total genomic DNA hybridization approach. Total genomic DNA (after restriction digestion) of 79 accessions of 23 Oryza species, 6 related genera, 5 outgroup taxa (2 monocots, 3 dicots) and 6 F₁s and BC₁s derived from crosses of O.?sativa with wild species were hybridized individually with ³²P-labeled total genomic DNA from 12 Oryza species: O. ridleyi, O.?longiglumis, O. granulata, O.?meyeriana, O. brachyantha, O. punctata, O. officinalis, O. eichingeri, O. alta, O. latifolia, O. australiensis, and O.?sativa. The labeled genomic DNAs representing the ridleyi and meyeriana complexes cross-hybridized best to all the accessions of their respective species, less to those representing other genomes of Oryza and related genera, and least to outgroup taxa. In general, the hybridization differential measured in terms of signal intensities was >50-fold under conditions that permit detection of 70–75% homologous sequences, both in the presence and in the absence of O. sativa DNA as competitor. In contrast, when total DNAs representing other Oryza genomes were used as probes, species of the O.?ridleyi and O.?meyeriana complexes did not show any significant cross-hybridization (<5%). These results demonstrate that the genome(s) of both of these complexes are highly diverged and distinct from all other known genomes of Oryza. We, therefore, propose new genomic designations for these two species complexes: GG for the diploid O. meyeriana complex and HHJJ for the allotetraploid O. ridleyi complex. The results also suggest that the uniqueness of these genomes is not restricted to species-specific highly repetitive DNA sequences, but also applies to dispersed sequences present in single or low to moderate copy numbers. Furthermore these appear to share relatively more genome-specific repeat sequences between themselves than with other genomes of rice. The study also demonstrates the potential of total genomic DNA hybridization as a simple but powerful tool, complementary to existing approaches, for ascertaining the genomic makeup of an organism.     

Evidences of introgression from cultivated rice to Oryza rufipogon (Poaceae) populations based on SSR fingerprinting: implications for wild rice differentiation and conservation

Crop-to-wild introgression may play an important role in evolution of wild species. Asian cultivated rice (Oryza sativa L.) is of a particular concern because of its cross-compatibility with the wild ancestor, O. rufipogon Griff. The distribution of cultivated rice and O. rufipogon populations is extensively sympatric, particularly in Asia where many wild populations are surrounded by rice fields. Consequently, gene flow from cultivated rice may have a potential to alter genetic composition of wild rice populations in close proximity. In this study, we estimated introgression of cultivated rice with O. rufipogon based on analyses of 139 rice varieties (86 indica and 53 japonica ecotypes) and 336 wild individuals from 11 O. rufipogon populations in China. DNA fingerprinting based on 17 selected rice simple sequence repeat (SSR) primer pairs was adopted to measure allelic frequencies in rice varieties and O. rufipogon samples, and to estimate genetic associations between wild and cultivated rice through cluster analysis. We detected consanguinity of cultivated rice in O. rufipogon populations according to the admixture model of the STRUCTURE program. The analyses showedz that four wild rice populations, DX-P1, DX-P2, GZ-P2, and HL-P, contained some rare alleles that were commonly found in the rice varieties examined. In addition, the four wild rice populations that scattered among the rice varieties in the cluster analysis showed a closer affinity to the cultivars than the other wild populations. This finding supports the contention of substantial gene flow from crop to wild species when these species occur close to each other. The introgressive populations had slightly higher genetic diversity than those that were isolated from rice. Crop-to-wild introgression may have accumulative impacts on genetic variations in wild populations, leading to significant differentiation in wild species. Therefore, effective measure should be taken to avoid considerable introgression from cultivated rice, which may influence the effective in-situ conservation of wild rice species.