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.     

Genetic Diversity Analysis of Tibetan Wild Barley Using SSR Markers

One hundred and six accessions of wild barley collected from Tibet, China, including 50 entries of the two-rowed wild barley Hordeum vulgare ssp. spontaneum (HS), 29 entries of the six-rowed wild barley Hordeum vulgare ssp. agriocrithon (HA), and 27 entries of the six-rowed wild barley Hordeum vulgare ssp. agriocrithon var. lagunculiforme (HL), were analyzed using 30 SSR markers selected from the seven barley linkage groups for studying genetic diversity and evolutionary relationship of the three subspecies of Tibetan wild barley to cultivated barley in China. Over the 30 genetic loci that were studied, 229 alleles were identified among the 106 accessions, of which 70 were common alleles. H. vulgare ssp. spontaneum possesses about thrice more private alleles (2.83 alleles/locus) than HS (0.93 alleles/locus), whereas almost no private alleles were detected in HL. The genetic diversity among-subspecies is much higher than that within-subspecies. Generally, the genetic diversity among the three subspecies is of the order HS > HL > HA. Phylogenetic analysis of the 106 accessions showed that all the accessions of HS and HA was clustered in their own groups, whereas the 27 accessions of HL were separated into two groups (14 entries with group HS and the rest with group HA). This indicated that HL was an intermediate form between HS and HA. Based on this study and previous works, we suggested that Chinese cultivated barley might evolve from HS via HL to HA.     

基于SSR标记构建葡萄种质资源分子身份证

以国家葡萄品种资源圃内保存的80份葡萄种质为试材,对构建葡萄种质分子身份证的方法进行探讨研究。利用筛选后的SSR标记对供试品种进行区分,然后根据引物对不同品种扩增条带分子量的大小进行编码。从62对引物中筛选出来自葡萄19条染色体上的28对SSR引物,在供试种质间共检测出等位基因169个,每对引物平均检测到等位基因数为6.0个。将等位基因赋值后仅用9对引物构建了供试种质的分子身份证编码,平均每对引物区分种质达8.9份,达到了区分品种更加简洁明了,用最少的引物区分不同品种的目的,表明SSR标记技术可有效用于建立葡萄种质资源分子身份证。     

SNP discovery using Paired‐End RAD‐tag sequencing on pooled genomic DNA of Sisymbrium austriacum (Brassicaceae)

Single nucleotide polymorphisms SNPs are rapidly replacing anonymous markers in population genomic studies, but their use in non model organisms is hampered by the scarcity of cost‐effective approaches to uncover genome‐wide variation in a comprehensive subset of individuals. The screening of one or only a few individuals induces ascertainment bias. To discover SNPs for a population genomic study of the Pyrenean rocket (Sisymbrium austriacum subsp. chrysanthum), we undertook a pooled RAD‐PE (Restriction site Associated DNA Paired‐End sequencing) approach. RAD tags were generated from the PstI‐digested pooled genomic DNA of 12 individuals sampled across the species distribution range and paired‐end sequenced using Illumina technology to produce ~24.5 Mb of sequences, covering ~7% of the specie's genome. Sequences were assembled into ~76 000 contigs with a mean length of 323 bp (N₅₀ = 357 bp, sequencing depth = 24x). In all, >15 000 SNPs were called, of which 47% were annotated in putative genic regions based on homology with the Arabidopsis thaliana genome. Gene ontology (GO) slim categorization demonstrated that the identified SNPs covered extant genic variation well. The validation of 300 SNPs on a larger set of individuals using a KASPar assay underpinned the utility of pooled RAD‐PE as an inexpensive genome‐wide SNP discovery technique (success rate: 87%). In addition to SNPs, we discovered >600 putative SSR markers.     

Signatures of diversifying selection at EST‐SSR loci and association with climate in natural Eucalyptus populations

Understanding the environmental parameters that drive adaptation among populations is important in predicting how species may respond to global climatic changes and how gene pools might be managed to conserve adaptive genetic diversity. Here, we used Bayesian F_ST outlier tests and allele–climate association analyses to reveal two Eucalyptus EST‐SSR loci as strong candidates for diversifying selection in natural populations of a southwestern Australian forest tree, Eucalyptus gomphocephala (Myrtaceae). The Eucalyptus homolog of a CONSTANS‐like gene was an F_ST outlier, and allelic variation showed significant latitudinal clinal associations with annual and winter solar radiation, potential evaporation, summer precipitation and aridity. A second F_ST outlier locus, homologous to quinone oxidoreductase, was significantly associated with measures of temperature range, high summer temperature and summer solar radiation, with important implications for predicting the effect of temperature on natural populations in the context of climate change. We complemented these data with investigations into neutral population genetic structure and diversity throughout the species range. This study provides an investigation into selection signatures at gene‐homologous EST‐SSRs in natural Eucalyptus populations, and contributes to our understanding of the relationship between climate and adaptive genetic variation, informing the conservation of both putatively neutral and adaptive components of genetic diversity.     

Development and Application of Microsatellites in Candidate Genes Related to Wood Properties in the Chinese White Poplar (Populus tomentosa Carr.)

Gene-derived simple sequence repeats (genic SSRs), also known as functional markers, are often preferred over random genomic markers because they represent variation in gene coding and/or regulatory regions. We characterized 544 genic SSR loci derived from 138 candidate genes involved in wood formation, distributed throughout the genome of Populus tomentosa, a key ecological and cultivated wood production species. Of these SSRs, three-quarters were located in the promoter or intron regions, and dinucleotide (59.7%) and trinucleotide repeat motifs (26.5%) predominated. By screening 15 wild P. tomentosa ecotypes, we identified 188 polymorphic genic SSRs with 861 alleles, 2–7 alleles for each marker. Transferability analysis of 30 random genic SSRs, testing whether these SSRs work in 26 genotypes of five genus Populus sections (outgroup, Salix matsudana), showed that 72% of the SSRs could be amplified in Turanga and 100% could be amplified in Leuce. Based on genotyping of these 26 genotypes, a neighbour-joining analysis showed the expected six phylogenetic groupings. In silico analysis of SSR variation in 220 sequences that are homologous between P. tomentosa and Populus trichocarpa suggested that genic SSR variations between relatives were predominantly affected by repeat motif variations or flanking sequence mutations. Inheritance tests and single-marker associations demonstrated the power of genic SSRs in family-based linkage mapping and candidate gene-based association studies, as well as marker-assisted selection and comparative genomic studies of P. tomentosa and related species.     

甘薯与牵牛EST资源的SSR信息分析

为挖掘番薯（Ipomoea）属EST-SSR资源,从NCBI数据库下载23406条甘薯（Ipomoea batatas (L.) Lam.）EST和62282条牵牛（Ipomoea nil (L.) Roth）EST,利用生物信息学软件预处理、去冗余、拼接处理后得到12812条无冗余的甘薯EST（6.70 Mb）和28422条牵牛唯一序列（17.19 Mb）。对这些序列进行SSR搜索,在甘薯上获得328个SSR位点,发生频率为2.56%;牵牛上筛选到962个SSR位点,出现频率为3.38%。甘薯和牵牛EST-SSR具有多个共同特征：在SSR位点中,主要是二核苷酸重复类型,其次是三核苷酸重复;在二核苷酸重复中,出现最多的重复基序为AG/CT,其次是AT/AT;在三核苷酸重复中,主要基序是AAG/CCT;SSR位点的长度主要集中在20~22 bp。结果表明,这些搜索出的EST-SSR重复基序类型丰富、多态性潜能高,具有较高的开发和利用价值。     

38份晾晒烟种质资源遗传关系的SSR分析

利用SSR标记技术对38份晾晒烟种质资源的遗传关系进行了分析。从自己开发的近3000对烟草SSR引物中随机选出30对引物,在38份供试材料中共检测出173个等位基因,每对SSR引物可检测的等位基因数为2~11个,平均为5.77个。38份材料间遗传相似系数（GS）的变化范围为0.165-0.928,平均GS 为0.546。表明38份晾晒烟的遗传多样性丰富,遗传差异较大,亲缘关系较远。聚类分析表明,在L1（GS-0.165）处可将38个品种分为2大类,即晾晒烟类群和美国从烟草起源地收集的烟草（TI：Tobacco Instruction）类群;晾晒烟类群又可进一步分为4组,其聚类结果与所期望的结果基本一致。表明SSR是一种有效、稳定和可靠的分子标记,能较好地从分子水平上揭示烟草(尤其是晾晒烟)种质资源的遗传背景和亲缘关系。     

基于SSR标记的宁夏水稻遗传多样性分析

选择自上世纪80年代以来的59份宁夏水稻种质,利用分布于12条染色体的82对SSR引物进行遗传多样性和遗传相似性分析。共检测到339个等位基因,品种间不同位点等位基因数目2~19个,平均4.13个。Nei基因多样性指数变幅为0.0333~0.9164,平均为0.4394。按种质释放或审定年代,55份水稻分为3组分析,随着年代的增加,等位基因数和遗传多样性指数均呈增大趋势,且3群体间等位基因数和多样性指数差异均达极显著水平（P<0.001）。UPGMA聚类分析表明,参试59份水稻种质在遗传相似系数0.78水平上聚为5大类,其中第Ⅰ类为1份香稻种质;第Ⅱ、Ⅳ类均为3份种质;第Ⅲ类有5份种质,与吉林水稻相似;绝大多数品种被聚为第Ⅴ类,占参试材料数的79.7％。对比宁夏水稻选育品种系谱,大多数种质具有宁夏骨干亲本红旗12、京引39、东方红2号、京引59等的血缘,种质间亲缘关系较近。虽然近年来宁夏水稻的遗传多样性有增加的趋势,但遗传基础狭窄,参试种质的相似系数在0.75以上,最高达0.97。应继续加大力度引进和创新亲本材料,拓宽宁夏水稻的遗传基础。