Genic microsatellite markers for discrimination of spinach cultivars

A set of simple sequence repeat (SSR or microsatellite) markers was used to discriminate a collection of 33 Spinacia oleracea hybrid cultivars from seven different breeding stations all over the world. All SSR markers were genic microsatellite markers located in coding or non-coding regions of genes of known function. Cluster analysis based on 13 of the SSR markers showed that the spinach hybrids grouped into three clusters. The first two clusters consisted of European spinach types, which were well discriminated according to their origin from different breeding stations. The third cluster was a mixture of Asian as well as European types of spinach. Subclusters in this group did not reflect differences in morphology, earliness or company origin. The data show that genic microsatellites are a powerful tool for discrimination of spinach cultivars.     

Isolation and characterization of microsatellite markers from guineagrass (Panicum maximum) for genetic diversity estimate and cross-species amplification

Guineagrass ( Panicum maximum Jacq.) is one of the major forage grasses in tropical and semitropical regions, largely apomictic and predominantly exist as tetraploid. Non-availability of polymorphic molecular markers has been a major limitation in its characterization and improvement. We report isolation and characterization of microsatellites in P. maximum and cross-species results with other five Panicum species. Based on microsatellite-motifs, 15 functional and polymorphic simple sequence repeat (SSR) primer-pairs were designed, validated and employed in estimating genetic relationship among 34 guineagrass accessions. Thirteen primer-pairs amplified single locus and remaining two generated more than two loci with an average of 3.57 bands per locus amounts to 63 bands with 34 guineagrass accessions. Average expected heterozygosity ( H _E) of 0.35 (maximum 0.97) and observed heterozygosity ( H _O) of 0.37 (maximum 0.91) established the efficiency of developed markers for discriminating guineagrass accessions. Dice's similarity coefficients-based unweighted pair group with arithmetic average method-clustering supported with high bootstrap values (≥40) indicated its significance and distinguished all accessions except IG97-93 and IG97-6. Utility of these new SSR loci in genetic diversity study of P. maximum and other cross–amplified species is discussed.     

Use of molecular marker diversity to increase forage yield in timothy (Phleum pratense L.)

Molecular marker technologies may provide a tool to overcome the forage yield plateau in timothy (Phleum pratense L.). Therefore, a study was designed to investigate the relationship between marker‐based genetic distance (GD) estimates and yield. Genetic distances were estimated using Dice coefficients framed by 28 simple sequence repeat markers. In Experiment 1, parents with high general combining ability (GCA) from two contrasting first‐generation synthetic strains (exhibiting high and low yields) were compared. Average GDs of each strain were 0.74 (high) and 0.68 (low). These differences indicated that GD may be partially the basis for contrasting yields. In Experiment 2, GDs among 40 parents of a polycross were used for calculating general genetic distance (GGD), which is indicative of their allelic complementation. Analyses revealed a significant (P < 0.01) moderate correlation (r) between GGD and GCA for yield (r = 0.45) and a significant (P < 0.01) residual mean square for the regression of yield on GGD, suggesting that considerable non‐additive effects were associated with GCA. The results are indicative of the potential use of GD estimation for yield improvement in timothy.     

Development,characterization and mapping of microsatellite markers for lentil (Lens culinaris Medik.)

Lentil is the sixth most important pulse crop terms of production in the world, but the number of available and mapped SSR markers are limited. To develop SSR markers in lentil, four genomic libraries for (CA)n, (GA)n, (AAC)n and (ATG)n repeats were constructed. A total of 360 SSR primers were designed and validated using 15 Turkish lentil cultivars and genotypes. The most polymorphic repeat motifs were GA and CT, with a mean number of alleles per locus of 7.80 and 6.55, respectively. Seventy‐eight SSR primers amplified a total of 400 polymorphic alleles, whereas 71 SSR primers produced markers within the expected size range. For 78 polymorphic SSR primers, the average number of alleles per locus was 5.1 and PIC value ranged from 0.07 to 0.89, with an average of 0.58. A linkage map was constructed using 92 individual F₂ plants derived from a cross between Karacada? × Silvan, with 47 SSR markers. The SSR markers developed in this study could be used for germplasm classification and identification and mapping of QTL in lentil.     

Development of genomic and EST-SSR markers in radish (Raphanus sativus L.)

Radish (Raphanus sativus L.) belongs to Brassicaceae family and is a close relative of Brassica. This species shows a wide morphological diversity, and is an important vegetable especially in Asia. However, molecular research of radish is behind compared to that of Brassica. For example, reports on SSR (simple sequence repeat) markers are limited. Here, we designed 417 radish SSR markers from SSR-enriched genomic libraries and the cDNA data. Of the 256 SSR markers succeeded in PCR, 130 showed clear polymorphisms between two radish lines; a rat-tail radish and a Japanese cultivar, ‘Harufuku’. As a test case for evaluation of the present SSRs, we conducted two studies. First, we selected 16 SSRs to calculate polymorphism information contents (PICs) using 16 radish cultivars and four other Brassicaceae species. These markers detected 3–15 alleles (average = 9.6). PIC values ranged from 0.54 to 0.92 (average = 0.78). Second, part of the present SSRs were tested for mapping using our previously-examined mapping population. The map spanned 672.7 cM with nine linkage groups (LGs). The 21 radish SSR markers were distributed throughout the LGs. The SSR markers developed here would be informative and useful for genetic analysis in radish and its related species.     

Genetic diversity and differentiation of Chinese wild soybean germplasm (G. soja Sieb. & Zucc.) in geographical scale revealed by SSR markers

Wild soybean (Glycine soja), as the progenitor of soybeans (G. max), is widely distributed in China and has been collected as a supplementary germplasm pool of soybeans. In this study, 375 wild soybean accessions from a set of genebank core collection were analysed for genetic diversity by using 42 simple sequence repeat primer pairs. The mean allele number per locus was 19.62. Ten‐percent unique alleles involving 35 or 83.33% loci differentiated among the geographical regions. The mean gene diversity (h) per locus was 0.89. A very low mean coefficient of gene differentiation (G_ST = 0.08) for geographical regions and a high mean within‐region gene diversity (H_S = 0.81) were observed, indicating that most genetic diversity existed within the regions. There was an obvious relationship between genetic distance and geographical distance. The results showed multiple centers of genetic diversity for Chinese wild soybean in North China, the Huanghe River Valley, and Central China as well as the Changjiang River Valley, implicating multiple site origins of soybeans within China.     

Evaluation of genetic diversity in jute (Corchorus species) using STMS, ISSR and RAPD markers

Jute is an important fibre crop that has dominated the packaging sector for over one and a half centuries in India. For sustenance of the trade in the face of tough competition from synthetics, there is an urgent need to redesign the ongoing breeding strategy to improve both the yield and quality of jute fibre. It is therefore, essential to understand the pattern of diversity in this important commercial crop species. In the present study, genetic diversity analysis of 20 exotic germplasm lines and 20 commercial varieties of the two cultivated species (Corchorus olitorius and C. capsularis) and two wild relatives of jute (C. aestuans and C. trilocularis) was carried out using sequence tagged microsatellite site (STMS), inter simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD) markers. The first set of six STMS markers developed from the genomic sequence of C. olitorius was not fully transferable to the related species C. capsularis. The level of intraspecific polymorphism revealed by these markers was very low. The four ISSR and 22 RAPD primers employed in the study revealed 98.44% and 100% polymorphism, respectively, across all the species, while the level of polymorphism was significantly low within a species. The commercial varieties, particularly those of C. capsularis, had an extremely narrow genetic base that demands immediate effort for diversification. The germplasm accessions in both the cultivated species showed considerably higher levels of diversity and thus should be used in broadening the base of the varieties. All the accessions of C. olitorius together with the wild species C. aestuans clustered separately from those of C. capsularis and C. trilocularis, suggesting a polyphyletic origin of the two cultivated species.     

Development of EST‐SSR markers for diversity and breeding studies in opium poppy

All publicly available opium poppy expressed sequence tag (EST) sequences, totalling 20 885, were assembled into unigenes and examined for simple sequence repeats (SSRs). Nearly 19% of the 14 957 unigenes contained SSRs with 4% harbouring more than one SSR. Average density of the SSRs was 1 SSR per 3.6 kb of non‐redundant EST sequence. Trinucleotide SSRs were most frequently identified (39%), and many of the most prevalent motifs were AT‐rich. Flanking primers were designed for 86% of the SSRs and 67 primer pairs were tested on 37 opium poppy accessions and seven related species. All markers were transferable to the related species. Polymorphism information content (PIC) values for the markers were intermediate for comparisons within opium poppy (average of 0.27) and slightly higher for comparisons across species (average of 0.29). The markers were found to be useful for diversity analysis as they successfully distinguished among Turkish opium poppy accessions and land races.     

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.     

表型与分子标记相结合分析130份糖用甜菜种质的遗传多样性

旨在对甜菜种质进行比较系统的遗传多样性分析与遗传距离分析。将表型与分子标记相结合,从3 个角度分析了130 份甜菜种质的遗传多样性,并对种质进行了遗传距离的分析。130 份甜菜种质表现出比较丰富的遗传多样性,其中22 项描述性性状的多样性指数在0.0451~1.1867 之间,平均值为0.8559;23 项数值型性状的变异系数在2.94%~594.84%之间,平均值为59.10%;34 个SSR标记的多态性信息含量在0.1289~0.7847 之间,平均值为0.4887。联合使用形态标记与分子标记计算遗传距离,将130份种质分成了5个近缘组,相邻组间产糖量、块根锤度、根腐病病情指数3项性状均有显著差异。结果表明被分析的甜菜种质遗传多样性比较丰富,遗传距离分析结果有生物学意义,可为种质创新工作提供参考。     

基于ISSR标记的龙眼种质资源遗传多样性及亲缘关系分析

为明确收集的龙眼种质资源遗传多样性水平和亲缘关系,以20份龙眼种质资源为研究对象,采用ISSR分子标记进行遗传多样性及亲缘关系分析。11条引物共扩增出113条DNA片段,其中多态性片段90条,占总扩增片段数的79.65%。供试龙眼材料的遗传相似系数介于0.5487～0.8673之间,平均遗传相似系数0.7758。20份龙眼资源材料的ISSR聚类结果表明,供试龙眼材料间的亲缘关系较近,聚类结果受亲本关系影响,一定程度上反映了品种的地理分布情况,与传统分类方法较为一致。研究结果为龙眼种质资源的分类、遗传进化研究及杂交组合选配提供一定的理论和技术支持。