Mining the global diversity of barley for Fusarium resistance using leaf and spike inoculations

Fusarium graminearum is a devastating fungal pathogen that causes significant yield and quality losses in cereals. We utilized a diversity set of barley (140 genotypes) to explore vital resistance alleles against this aggressive pathogen. The resistance assessment was carried out on spikes and leaves via artificial inoculations under control conditions. The phenotypic data was subjected to genome-wide association analysis using a genetic map based on DArT and SNP markers. This analysis revealed eleven and nine marker trait associations for leaf disease scoring (LDS) and spike disease scoring (SDS), respectively. The strongest QTL for LDS was found on chromosome 1H where a minor allele of wild origin decreased disease symptoms by 78%. The major QTL allele for SDS was linked with marker locus SCRI_RS174710 on chromosome 5H. In addition, four favorable epistatic interactions effects were found in decreasing disease symptoms. Overall, three QTL were common for LDS and SDS, which indicates a partial genetic relatedness of these resistances in barley. The QTL alleles for LDS and SDS will help to establish organ specific resistances in cultivated barley.     

Ribosomal DNA spacer-length polymorphisms in three samples of wild and cultivated barleys and their relevance to the origin of cultivated barley

This study was conducted to provide additional data for evaluating two important issues surrounding the origin of cultivated barley: (i) the level of genetic diversity of the two‐rowed wild barley from Tibet, and (ii) the distribution of rDNA allele 104 in wild and cultivated barleys in the Occidental region. A total of 198 accessions consisting of three distinct samples were used: 82 entries of two‐rowed wild barley from Tibet, 57 accessions of two‐rowed wild barley from 8 countries with a broad range of representation of two‐rowed wild barley in the world, and 59 landrace accessions from four countries representing a part of the barley‐growing areas in the Middle East. These were assayed for rDNA spacer‐length variants (slvs). In all, 27 rDNA space length pheno types were detected, from which 10 slvs were identified as alleles at the two rDNA loci. The two‐rowed wild barley samples from Tibet had the lowest level of genetic variation as evaluated by rDNA polymorphism. Together with results of previous studies, the two wild forms (two‐rowed and six‐rowed) from Tibet could not account for the large genetic diversity observed in the cultivated barley of this region, suggesting that Tibet is unlikely a centre of origin for cultivated barley. In samples from the Occidental region, allele 104 of Rm2 was very rare in wild barley, but occurred at the highest frequency in cultivated barley, while the reverse is the case for allele 107, which is consistent with previous results. The implications of such a contrasting distribution of these rDNA alleles between wild and cultivated barleys in the origin and evolution of cultivated barley were discussed.     

A preliminary genetic analysis of fibre traits and the use of new genomic SSRs for genetic diversity in jute

Jute is one of the most important fibre crops, which is second only to cotton in providing environment-friendly (biodegradable and renewable) ligno-cellulose fibre. In order to improve this largely neglected crop, we conducted a preliminary study involving the following: (i) analysis of nature and extent of the genetic variability for fibre yield and four other related traits in a set of 81 genotypes belonging to two commercially cultivated Corchorus species (45 genotypes of C. olitorius + 36 genotypes of C. capsularis), (ii) development and analysis of a set of simple sequence repeat (SSR) markers from C. olitorius, and (iii) use of a sub-set of SSRs for assessment of genetic diversity in the above set of 81 genotypes. The results suggested quantitative nature of fibre yield and other related traits, with a preponderance of dominance component in genetic variance. A sub-set of 45 SSRs derived from C. olitorius, when used for a study of DNA polymorphism and genetic diversity, showed high transferability of these C. olitorius SSRs to C. capsularis. The average number of alleles for individual SSRs was surprisingly low (3.04 for both species, 2.02 for C. capsularis and 2.51 for C. olitorius), and so was the average polymorphic information content (PIC; 0.23 and 0.24 in two species). In the dendrogram obtained using a similarity matrix, the 81 genotypes were grouped into three clusters, which largely corresponded to the two species, Cluster I belonging mainly to C. capsularis and the other two closely related clusters (clusters II and III) belonging to C. olitorius. It was also shown that a minimum of 15 SSRs could give the same information as 41 SSRs, thus making many SSRs redundant. The SSR markers developed during the present study and to be developed in future will prove useful not only for evaluation of genetic diversity, but also for molecular mapping/QTL analysis, and for comparative genome analysis of the two Corchorus species. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Comparison among available marker systems for cereal introgression breeding: A practical perspective

Summary There is an increasing amount of public sequence information for the main cultivated cereals, such as wheat and barley. It is not foreseeable that comparable efforts or resources could be devoted to related wild species. However, wild species are interesting sources of genetic variation through introgression breeding. Comparative genomics can be a helpful approach to make use of the available genomic resources. In this context, the potential of the wild barley species Hordeum chilense has been explored in recent years. It exhibits great levels of polymorphism and high crossability with different cereal genera. In addition, interesting biotic and abiotic stress resistance genes, and important quality traits like carotene content and seed storage protein variability shown in the species are also expressed in wheat backgrounds, and are the basis of a breeding program. Different approaches have been undertaken for tagging H. chilense genomic regions in a wheat background. The search for the most suitable DNA marker system started with the development of RAPD and SCAR markers due to a lack of sequence information from the wild species. Transferability of markers from wheat and barley (like STSs or SSRs) have also been useful approximations. More recently, SNP development is being accomplished for the species. In this work, the situation and prospects with the available molecular tools are considered from a practical point of view.     

Characterization of novel sugarcane expressed sequence tag microsatellites and their comparison with genomic SSRs

Microsatellites or simple sequence repeats (SSRs) are one of the most suitable markers for genome analysis as they have great potential to aid breeders to develop new improved sugarcane varieties. The development of SSR derived from expressed sequence tags (EST) opens new opportunities for genetic investigations at a functional level. In the present work, the polymorphism obtained with a subset of 51 EST–SSRs derived from sucest was compared with those generated by 50 genomic SSRs (gSSR) in terms of number of alleles, polymorphism information content, discrimination power and their ability to establish genetic relationships among 18 sugarcane clones including three Saccharum species (S. officinarum, S. barberi, S. sinense). The majority of EST–SSRs loci had four to six alleles in contrast to the seven to nine observed for the gSSRs loci. Approximately, 35% of the gSSRs had PIC values around 0.90 in contrast to 15% of the EST–SSRs. However, the mean discrimination power of the two types of SSR did not differ significantly as much as the average genetic similarity (GS) based on Dice coefficient. The correlation between GS of the two types of SSRs was high (r = 0.71/P = 0.99) and significant. Although differences were observed between dendrograms obtained with each SSR type, both were in good agreement with pedigree information. The S. officinarum clone IJ76‐314 was grouped apart from the other clones evaluated. The results here demonstrate that EST–SSRs can be successfully used for genetic relationship analysis, extending the knowledge of genetic diversity of sugarcane to a functional level.     

Identification of sugarcane microsatellites associated to sugar content in sugarcane and transferability to other cereal genomes

Microsatellites or simple sequence repeats (SSRs) markers are very informative for various applications in genetics and breeding. Information obtained with these markers has contributed to a better understanding of evolution and the complexity of the sugarcane genome. With the objective of identifying a large set of polymorphic microsatellite markers designated as Unigene derived Sugarcane Microsatellite (UGSM) and Sugarcane Enriched Genomic Microsatellite (SEGMS), 351 UGSM and 36 SEGMS were tested to find out informative SSRs marker for sugar content. These markers were screened and validated for their use in genetic diversity, cross transferability and comparative linkage potential in high and low sugar bulk of two segregating progenies and twenty each, cultivated high and low sugar cultivars. 158 (40.83%) of the microsatellite markers (144-UGSM: 14-SEGMS) were found to be highly robust and polymorphic. Cross amplification was estimated among nineteen accessions of six sugarcane cultivars, one inter specific hybrids, five related species, four related genera, and three divergent genera by using 27 UGSM primers. Analysis of 388 alleles, amplified by these markers, indicated the high number of observed allele ranged from 2 to 26, with an average of 14.37 alleles detected per locus. High level of polymorphism detected by these markers among sugarcane species, genera and cultivars was 96.3%, while cross-transferability rate was 98.0% within Saccharum complex and 88.27% to cereals. Wide range of genetic diversity (0.33–0.79 with an average of 0.56) assayed with UGSM markers suggested their importance in various genotypic applications in sugarcane.     

花生栽培种EST-SSRs分布特征及应用研究

利用自行开发的20 160条花生栽培种荚果EST, 通过序列拼接, 获得8 289条无冗余EST。经搜索, 共检测出740个SSR位点, 分布于651条EST中, 发生频率为7.8%, 平均每6.8 kb EST序列含一个SSR位点。功能注释结果表明具生物过程、分子功能和细胞组分的EST分别为73、111和56条。在花生荚果EST-SSR中, 三核苷酸重复类型出现频率最高, 占总SSR的62.8%, 其次是二核苷酸重复类型, 占总SSR的33.6%。在出现的26类重复基序中, AG/TC重复基序出现频率最高, AAG/TTC次之。利用Primer premier 5从651条含有SSR的EST中共设计引物233对, 从中随机选取100对引物检测EST-SSR在花生栽培种中的多态性及在野生种中的可转移性。结果表明, 有86对引物在供试的22个花生栽培品种中得到有效扩增, 其中10对在栽培种中具有多态性, 每对引物检测出的等位基因数2~3个, 平均2.2个。可扩增引物在野生种中的可转移率为12.5%~100%,平均96%。在野生种间检测出多态性的引物76对,每对引物检测出等位基因2~9个, 平均4.06个。     

Genetic variation in South Korean natural populations of wild soybean (Glycine soja)

Summary Wild relatives are valuable genetic resources for crop improvement. Evaluating genetic variation in these species is not only important for their use in breeding programs, but will also provide information about evolution of crops. Seeds representing six natural populations were used to study the level of variation in the South Korean wild soybean. Electrophoretic assays of the seeds on horizontal slab gels were conducted to determine the genotypes of each natural plant at 35 loci in 17 isozymes and one protein. The results indicated a surprisingly high variation. The number of alleles at each locus was as high as four. Seventy two of the 94 reported alleles for the 35 loci were present in these populations. The average number of alleles per locus, 99% polymorphism and the expected heterozygosity in the total population were 2.1, 77.1% and 0.215, respectively. This amount of variation was not only higher than that reported for 857 soybean cultivars and wild soybean populations from other geographic regions, but also higher than the average for 123 self-fertilized plant species and 473 plant species of all mating systems. The high variation in the South Korean wild soybean as well as cultivated soybean indicated in this and other population genetic studies prompts us to propose that South Korea is one of the major soybean gene centers.     

Development, characterization and utilization of microsatellite markers in pigeonpea

Pigeonpea is a major legume of the semi‐arid tropics that has been neglected in terms of molecular breeding. The objectives of this study were to develop microsatellite markers and evaluate their potential for use in pigeonpea genetics and breeding. Two hundred and eight microsatellite loci were isolated by screening a non‐enriched partial genomic library. Primers were designed for 39 microsatellite loci, 20 of which amplified polymerase chain reaction products of the expected size. Nineteen of the primer pairs were polymorphic amongst 15 cultivated and nine wild pigeonpea accessions providing evidence for cross‐species transferability within the genus Cajanus. A total of 98 alleles were detected at the 19 polymorphic loci with an average of 4.9 alleles per locus. The observed heterozygosity ranged from 0.17 to 0.80 with a mean of 0.42 per locus. Less allelic variation (31 alleles) was observed within the cultivated species than across the wild species (92 alleles). The diversity analysis readily distinguished all wild relatives from each other and from the cultivated germplasm. Development of more microsatellites is recommended for future genomic studies in pigeonpea.     

Genetic diversity in groundnut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.) genotypes and detection of marker trait associations for plant habit and seed size using genomic and genic SSRs

Groundnut (Arachis hypogaea L.) an important oilseed crop in India is known to have narrow genetic base. Therefore, the assessment of genetic diversity and detection of marker-trait association are important objectives for the genetic improvement of groundnut. The present study involved the development of 192 SSR markers from Arachis genomic survey sequences. From these, seven polymorphic SSRs along with 15 other genomic SSRs, 19 genic SSRs, and three STS markers were used to detect genetic diversity among 44 groundnut genotypes. These polymorphic SSR markers amplified 155 bands (76 genomic and 79 genic), of these 128 bands (67 genomic and 61 genic) were polymorphic. The genomic SSR exhibited 88.1% and genic SSRs displayed 77.2% allelic polymorphism. The polymorphic information content (PIC) of the markers ranged from 0.04 to 0.95. The pair-wise genetic similarity ranged from 24.2 to 90.7% for genomic SSR and 32.9 to 97.9% for genic SSR markers. Cluster analysis based on the pooled data from both genomic and genic SSRs revealed a dendrogram which could distinguish all the genotypes. Further, the AMOVA analysis detected 16.7% genetic variation due to differences in seed size and 13.0% due to plant habit. Based on locus-by-locus AMOVA and Kruskal-Wallis ANOVA and further confirmation by discriminant analysis and general linear model, six markers were found to be associated with plant habit and four markers with seed size.     

Genetic variability in melon based on microsatellite variation

A set of 18 simple‐sequence repeat (SSR or microsatellite) markers was used to study genetic diversity in a collection of 27 melon (Cucumis melo L.) accessions, representing a broad range of wild and cultivated melons. The materials studied were highly polymorphic for SSRs and a total of 114 alleles were detected (average of 6.3 alleles per locus). Cluster analysis suggests the division of these accessions into two major groups, largely corresponding to the division of C. melo in the two subspecies agrestis and melo. The assignment of the accession to the subspecies was generally in agreement with published reports, except for those corresponding to the ‘dudaim’ and ‘chito’ cultivar groups, which, according to the observed SSR variability, should be included in subspecies agrestis. Based on cluster analysis, five groups of accessions were defined. The two most divergent groups include mainly accessions from the Mediterranean which form one group, and accessions from China, Japan, Korea and India forming the other. Both groups shared a low level of intra‐accession variation compared with the other groups, which suggests an erosion of their genetic variability because of drift and/or inbreeding. The remaining accessions, mainly from Central Africa and India, were more variable and may be an important source of genetic variation for melon breeding.     

Homologous analysis of SSR-ESTs and transferability of wheat SSR-EST markers across barley, rice and maize

Summary Comparative genetic analysis has shown that different plant species often share orthologous genes for very similar functions, especially for cereal crops. In this study, we focused on analyzing the utility of EST-SSR markers among monocots in silico and by experiment. Based on 423,611 wheat ESTs, 101,299 were found to express commonly in rice, maize and barley ESTs, which accounted for 23.94% of wheat EST data. 1707 SSR-containing ESTs (SSR-ESTs) were mined from the 101,299 homologous ESTs, which accounted for 8.8% of all the 19,434 wheat EST-SSRs. It showed that the number of homologous SSR-ESTs much less than the homologous ESTs among grasses. The number of conserved ESTs and SSR-ESTs were inversely proportional to the evolutionary distance among species. Although these SSR-ESTs were in high similarity, the number of conserved SSRs between wheat and the other three crops decreased with the increased aligning regions, and only 13 types of SSR motifs in 21 ESTs were obtained by aligning sequences over a stretch of more than 40 bp. The variation at a microsatellite came from not only the differences in the numbers of repeat units and its location shift, but also base mutations flanking microsatellite repeats. From the 428 uni-SSR-ESTs universally in the four cereals, 243 primers were designed and tested in two genotypes of each species. 154 (63.4%) primers produced clear amplicons across the four species, which showed a high efficient transferability of wheat EST-SSR markers to the other three cereal crops.     

Isozyme variation in Helianthus petiolaris and sunflower,H. annuus

Summary Helianthus petiolaris is a wild species used in genetic improvement of sunflower, as a donor of cytoplasmic male sterility and of genetic resistance to diseases. Isozyme variation for ADH, ACP, EST, GDH, LAP, PGI, PGD and SKDH in this species was studied using starch gel electrophoresis. The patterns thus obtained were compared with zymograms of inbred lines, hybrids and open pollinated varieties of H. annuus. The same alleles for EST and SKDH isozymes were found in both species, while ACP showed an allele that has not been found in sunflower. The rest of the isozyme systems showed both common alleles and characteristic ones for each species. ACP, GDH and PGD were monomorphic in H. petiolaris, while ADH and LAP were monomorphic in H. annuus. The isozyme markers obtained here could be useful in breeding programs involving interspecific crosses, and studies on introgression and on genetic variation in other populations of this wild species.     

EST‐SSR analysis provides insights about genetic relatedness,population structure and gene flow in grass pea (Lathyrus sativus)

Grass pea (Lathyrus sativus) is an important food‐legume crop for resource‐poor farmers in the developing world. However, given its cultivation in the most underprivileged regions, the crop has not received appropriate scientific attention particularly from the genomic perspective, thereby giving it a status of genomic orphan. Nevertheless, some recent studies have attempted to develop modern molecular tools to strengthen the genetic and genomic research. In the present investigation, a comprehensive collection comprising 176 accessions was analysed using EST‐simple sequence repeats (SSRs). The SSR analysis revealed existence of a total of 51 alleles with an average polymorphic information content value of 0.35. A moderate level of gene diversity was noticed that ranged from 0.04 to 0.73 with an average of 0.43. Noticeably, two distinct subpopulations were recovered using cluster analysis. In addition, the presence of admixtures in population reflected the strong possibilities of gene flow between the accessions across the geographical boundary. In summary, we provide additional insights about the informativeness of available EST‐SSR markers along with an extended understanding of relatedness, genetic structure and gene flow in an under‐researched legume crop.     

野生花生遗传物质渗入到栽培种的分子证据

花生属野生种具有高抗严重影响花生产量的主要病虫害的优良基因,花生区组二倍体野生种A.correntina对锈病、斑驳病毒病、PStV、蓟马、蚜虫、叶蝉、螨虫、玉米螟等多种病虫害也具有抗性.19份由珍珠豆型农家品种贺粤1号与A.correntina经可育性杂交获得三倍体F1代,F1代再经过人工染色体加倍、回交和多代自交选择,形成的能稳定遗传且性状优良的四倍体新品系,4份栽培品种和野生亲本A.correntina共24份材料用于SSR分析,40对SSR引物中有3对引物PM36、PM50、PM305,能在部分杂种后代（PM36：T60;PM50：J17、J20、J22;PM350：J7、S11、T62）中稳定地扩增出野生亲本的特异谱带,表明这些材料整合了野生亲本的遗传物质.SSR特异谱带可以作为这几份材料中野生亲本A.correntina的特异遗传标记.此外,有两对引物PM36、PM106能在一些杂种后代中扩增出父母本没有的DNA条带,使SSR分子标记表现出非共显性.     

A method for testing drought tolerance in <Emphasis Type="Italic">Fragaria</Emphasis> based on fast screening for water deficit response and use of associated AFLP and EST candidate gene markers

Drought stress is one of the most important environmental factors that limit plant growth and development, thus reducing yield. The objective of the present research was to correlate the genetic structure of different Fragaria genotypes, as assessed by Expressed Sequence Tag (EST) and Amplified Fragment Length Polymorphism (AFLP) markers, and plant responses to drought stress. Firstly, physiological parameters related to the plant response to drought stress such as leaf relative water content (RWC) and water losing rate (WLR) were measured. WLR and RWC were compared for 20 cultivars of the octaploid Fragaria × ananassa, two ecotypes of the diploid species F. vesca and one octaploid species F. chiloensis. These parameters could discriminate genotypes showing a contrasting response to water stress. Secondly, AFLP and ESTs were compared in terms of their information content and efficiency in the study of genetic diversity and relationships among these 23 Fragaria genotypes. To evaluate the genetic basis for the observed variation in the measured physiological parameter, the effect of specific AFLP/EST loci on WLR and RWC for the different Fragaria genotypes was quantified by Kruskal–Wallis analysis. By Mantel testing, the hierarchical clustering of the Fragaria genotypes based on associated EST or AFLP markers was compared to the observed eco-physiological relevant grouping. A better discriminating capacity for associated markers was noted, enabling a functional marker selection approach to screen the strawberry gene pool for drought tolerance. Correlation of EST markers to leaf RWC and WLR enforces them as potential candidate genes in control of plant responses to drought stress in Fragaria sp.     

Genetic diversity and relationships among Lablab purpureus genotypes evaluated using RAPD as markers

Summary Genetic variation in 40 accessions of Lablab purpureus was evaluated using random amplified polymorphic DNA as markers. A high level of genetic variation in this species was detected but this was mainly restricted to the difference between cultivated and wild forms. Of the cultivated genotypes, genetic variation among Asian collections was significantly higher than that among African collections. The three most divergent cultivated genotypes were all from Asia. Four of the five wild accessions, two from Zimbabwe and the other two from Zambia, were closely related. The other one, CPI 31113 collected from Uganda, was highly divergent. The two commercial forage varieties used in Australia, Rongai from Kenya and Highworth from India, were not very different.     

Simple sequence repeats for genetic analysis in pear

The development of highly informative DNA markers, such as simple sequence repeats (SSRs), is essential for breeding to select agronomically important traits and for genetic studies in pear. We developed SSR markers by using two approaches, RAHM (random amplified hybridization microsatellites) and 5' anchored PCR methods. Segregation analysis of the SSRs revealed that amplified fragments were derived from the same loci, using 3 sets of progenies from crosses between pear varieties. Genetic diversity was characterized using 32 varieties, including 10 from Japanese pear (Pyrus pyrifolia), 9 from Chinese pear (P. bretschneideri, P. ussuriensis), 10 from European pear (P. communis) as well as 3 wild relatives (P. calleryana). Diversity of SSR genotypes was observed among species as well as within species and 65 putative alleles were detected. The use of seven SSR markers was sufficient to differentiate between all of the 32 varieties. This revised version was published online in August 2006 with corrections to the Cover Date.