Ten novel microsatellite markers for the freshwater sleeper Micropercops swinhonis (Günther, 1873), with testing of cross‐species amplification in two other fishes from suborder Gobioidei

Ten novel polymorphic microsatellite loci were developed for the freshwater sleeper Micropercops swinhonis, a relatively abundant and broadly distributed lake‐dwelling fish in Asia, using next generation sequencing. These markers were tested in a population of 48 individuals, and characteristics suggest that they may be useful for population genetic studies. The average number of alleles per marker was 6.6 (range: 2–26), the average expected heterozygosity was 0.55 (range: 0.081–0.955), and the average polymorphic information content value per marker was 0.507 (range: 0.077–0.942). No marker showed significant deviation from Hardy‐Weinberg Equilibrium following Bonferroni correction of significance values, and no markers showed evidence of null alleles, large allele dropout, or scoring errors from stutter bands. Two markers had successful amplification in Odontobutis potamophila, and one marker had successful amplification in Rhinogobius giurinus. These novel markers may be useful for further research in population genetic studies involving M. swinhonis.     

利用产量位点标记分析中国水稻骨干恢复系与南亚及东南亚恢复系的遗传多样性

利用47个根据已报道的QTL位点或者已被精细定位或克隆的与水稻产量性状基因紧密连锁的产量位点标记,对来自中国、印度和越南等国的58份水稻恢复系进行遗传多样性分析。结果表明:(1)在中国恢复系中,47个产量位点标记中有36个具有多态性,共检测到90个等位基因,每个标记检测到等位基因2~4个,平均为2.500个;有效等位基因共62.905个,平均每个标记1.747个;36个有效标记的Shannon信息指数平均值为0.632,变幅为0.271~1.266。(2)在来自国外的材料中,47个产量位点标记均具有多态性,共检测到131个等位基因,每个标记检测到的等位基因数为2~6个,平均2.787个;有效等位基因数共82.686个,平均1.759个;47个标记的Shannon信息指数平均值为0.649,变幅为0.109~1.110。(3)聚类分析显示,在遗传相似系数为0.73水平上,参试资源聚为三大类群,中国资源多聚在第Ⅰ类群下的第1、2、3亚群,越南资源多聚在第Ⅰ-4亚群,孟加拉资源多聚在第Ⅲ-3亚群。由此表明,中国资源遗传基础较为狭窄,而其他国家的恢复系具有较远的亲缘关系。     

Development of Simple Sequence Repeat Markers from Functional Genes and Establishment of Molecular Identity for Tree Peony

Gene-derived simple sequence repeats (genic SSRs), also known as functional markers, are generally superior to random markers because they are located in genes and therefore may affect gene expression or function. However, extremely limited genic SSRs are available for tree peony. We used the functional gene sequences available from Paeonia to develop genic SSRs. A total of 132 SSR loci were identified from 35 cDNA sequences, of which trinucleotide (58, 43.9%) and hexanucleotide repeat (37, 28.0%) were dominant. Moreover, 121 primer pairs were successfully designed and synthesized, of which 49 primer pairs (40.5%) provided efficient and reliable amplification. By screening 16 tree peony varieties, we developed eight polymorphic genic SSRs with 37 alleles, ranging from 2 to 11 for each marker. Transferability analysis indicated that 100% of the genic SSRs could be amplified in eight other Paeonia samples. Based on eight polymorphic genic SSRs and 12 polymorphic EST-SSRs developed by predecessors, the molecular identity of 190 tree peony cultivars was constructed by capillary electrophoresis. The results showed that 146 alleles and 338 genotypes were detected, with 2–13 alleles and 3–36 genotypes for each marker. All cultivars were completely identified and exhibited unique DNA identity. In addition, the identification efficiency of different primers combinations was analyzed, and 190 germplasms were identified using 6 core primers. This study provides valuable genic SSR resources for marker-assisted selection breeding of the genus Paeonia. The DNA identity of cultivars is of great significance for the protection, utilization and management of tree peony resources.

     

Genetic diversity of elite sweet sorghum genotypes assessed by SSR markers

To determine genetic diversity among 47 elite sweet sorghum (Sorghum bicolor ssp. bicolor L.) genotypes, 46 simple sequence repeat (SSR) markers evenly distributed on all 10 chromosomes were selected. All SSR markers used were polymorphic among the genotypes studied. A total of 228 alleles were identified with an average of 4.96 alleles per marker. Furthermore, the genotypes studied showed medium genetic diversity. Clustering analysis grouped the 47 genotypes into 5 distinct clusters.     

Polymorphism of SSR alleles in pear cultivars grown in Belarus

Using SSR markers designed for Malus × domestica Borkh. genetic polymorphism of 43 pear accessions cultivated in Belarus was examined. A total of 217 alleles were identified with the mean number of 12.8 alleles per marker. The mean PIC value was 0.81; the mean number of informative alleles, 6.49. The heterozygosity level ranged from 0.30 to 0.84. Genetic diversity of SSR alleles in pear and apple genomes was compared. A method of identification of commercial pear cultivars using a set of six SSR markers was suggested.     

Microsatellite repeats in grapevine reveal DNA polymorphisms when analysed as sequence-tagged sites (STSs)

Microsatellite repeat sequences were investigated as sequenced-tagged site (STS) DNA markers to determine the potential for genetic analysis of the grapevine genome. The PCR-generated markers detect codominant alleles at a single locus or site in the genome. The marker type is very informative detecting high heterozygosity (69%–88%) within individual grapevine cultivars and high genetic variation between cultivars, making it a useful marker type for plant genome mapping and genome typing. For five loci a screening of 26 V. vinifera cultivars found 13, 12, 8, 5, and 4 different length alleles respectively with some alleles more common than others. The genomic DNA sequences surrounding microsatellite sequences were conserved within the genus permitting STS primers to amplify STSs from other Vitis species. These Vitis species were found to have some unique alleles not present in V. vinifera.     

Development and characterization of microsatellite markers for two dioecious Ficus species

Microsatellite markers for Ficus montana and Ficus septica were developed using genomic libraries enriched for di‐, tri‐ and tetranucleotide repeats. The subsets of five and three best scorable primer pairs were characterized on 24 F. montana and 36 F. septica individuals, respectively. For F. montana, loci showed five to 14 alleles per locus and expected heterozygosities ranged between 0.23 and 0.87. For F. septica, loci showed three to five alleles per locus and expected heterozygosities ranged between 0.36 and 0.49. Four primer pairs (two from each subset) cross‐amplified in the other species, indicating transportability of the markers within the genus Ficus.     

The use of microsatellite markers for detection of genetic diversity in barley populations

 A barley lambda-phage library was screened with (GA)n and (GT)n probes for developing microsatellite markers. The number of repeats ranged from 2 to 58 for GA and from 2 to 24 for GT. Fifteen selected microsatellite markers were highly polymorphic for barley. These microsatellite markers were used to estimate the genetic diversity among 163 barley genotypes chosen from the collection of the IPK Genebank, Germany. A total of 130 alleles were detected by 15 barley microsatellite markers. The number of alleles per microsatellite marker varied from 5 to 15. On average 8.6 alleles per locus were observed. Except for GMS004 all other barley microsatellite markers showed on average a high value of gene diversity ranging from 0.64 to 0.88. The mean value of gene diversity in the wild forms and landraces was 0.74, and even among the cultivars the gene diversity ranged from 0.30 to 0.86 with a mean of 0.72. No significant differences in polymorphism were detected by the GA and GT microsatellite markers. The estimated genetic distances revealed by the microsatellite markers were, on average , 0.75 for the wild forms, 0.72 for landraces and 0.70 among cultivars. The microsatellite markers were able to distinguish between different barley genotypes. The high degree of polymorphisms of microsatellite markers allows a rapid and efficient identification of barley genotypes. Received: 26 November 1997 / Accepted: 19 January 1998     

Isolation of 55 microsatellite markers for Jatropha curcas and its closely related species

Jatropha curcas L. (physic nut) is native to Central America and now naturalized widely in many tropical and subtropical areas. Microsatellite markers were isolated and characterized. Eleven out of 55 markers showed polymorphisms, and the allelic variation was investigated using 26 accessions of J. curcas collected from several provinces in Thailand. Each marker showed 2 to 5 alleles and the average polymorphic information content (PIC) was 0.49. Thirty four markers (62 %) were also successfully amplified in J. integerrima, J. gossypifolia and J. podagrica.     

Comparative study of the discriminating capacity of AFLP and ISTR markers for genetic analysis of<Emphasis Type="Italic">Agave fourcroydes</Emphasis>

Two DNA-based marker systems, amplified fragment legnth polymorphism (AFLP) and inverse sequence-tagged repeat (ISTR), were evaluated with respect to their discriminating capacity and efficiency in genetically analyzing henequen (Agave fourcroydes). Samples were taken from a mother plant, sucker-derived daughter plants, and bulbils. ISTR analysis produced more polymorphic markers than AFLP and had a better capacity for quantifying genetic diversity through an average number of alleles per locus, expected heterozygosis of polymorphic loci, expected heterozygosity, effective number of alleles per locus, and total number of effective alleles. ISTR also showed superior discriminatory capacity.     

Microsatellite analysis of Aegilops tauschii germplasm

The highly polymorphic diploid grass Aegilops tauschii isthe D-genome donor to hexaploid wheat and represents a potential source for bread wheat improvement. In the present study microsatellite markers were used for germplasm analysis and estimation of the genetic relationship between 113 accessions of Ae. tauschii from the gene bank collection at IPK, Gatersleben. Eighteen microsatellite markers, developed from Triticum aestivum and Ae. tauschii sequences, were selected for the analysis. All microsatellite markers showed a high level of polymorphism. The number of alleles per microsatellite marker varied from 11 to 25 and a total of 338 alleles were detected. The number of alleles per locus in cultivated bread wheat germplasm had previously been found to be significantly lower. The highest levels of genetic diversity for microsatellite markers were found in accessions from the Caucasian countries (Georgia, Armenia and the Daghestan region of Russia) and the lowest in accessions from the Central Asian countries (Uzbekistan and Turkmenistan). Genetic dissimilarity values between accessions were used to produce a dendrogram of the relationships among the accessions. The result showed that all of the accessions could be distinguished and clustered into two large groups in accordance with their subspecies taxonomic classification. The pattern of clustering of the Ae. tauschii accessions is according to their geographic distribution. The data suggest that a relatively small number of microsatellites can be used to estimate genetic diversity in the germplasm of Ae. tauschii and confirm the good suitability of microsatellite markers for the analysis of germplasm collections. Received: 8 September 1999 / Accepted: 7 October 1999     

Genetic variation at pig plasma protein locus PLP1 and its assignment to chromosome 5

A new genetic polymorphism of an unidentified plasma protein (PLP1) in pigs was described by using a method of two-dimensional gel electrophoresis and protein staining. Two codominant alleles, with frequencies of 0.83 and 0.17, were found in the Swedish Yorkshire breed. The PLP1 marker was typed in a three-generation pedigree and tested for linkage against a set of 128 markers. The PLP1 locus showed significant LOD score values with three different microsatellite markers (S0092, DAGK and S005), previously assigned to chromosome 5.     

Features of SNP and SSR diversity in a set of ICARDA barley germplasm collection

Detection and utilization of genetic variation available in the germplasm collection for crop improvement have been the prime activities of breeders. Here a set of ICARDA barley germplasm collection comprising of 185 cultivated (Hordeum vulgare L.) and 38 wild (H. spontaneum L.) genotypes originated from 30 countries of four continents was genotyped with 68 single nucleotide polymorphism (SNP) and 45 microsatellite or simple sequence repeat (SSR) markers derived from genes (expressed sequence tags, ESTs). As two SNP markers provided 2 and 3 datapoints, a total of 71 SNPs were surveyed that yielded a total of 143 alleles. The number of SSR alleles per locus ranged from 3 to 22 with an average of 7.9 per marker. Average PIC (polymorphism information content) value for SSR and SNP markers were recorded as 0.63 and 0.38, respectively. Heterogeneity was recorded at both SNP and SSR loci in an average of 5.72 and 12.42% accessions, respectively. Genetic similarity matrices for SSR and SNP allelic data were highly correlated (r = 0.75, P < 0.005) and therefore allelic data for both markers were combined and analyzed for understanding the genetic relationships among the germplasm surveyed. Majority of clusters/subclusters were found to contain genotypes from the same geographic origins. While comparing the genetic diversity, the accessions coming from Middle East Asia and North East Asia showed more diversity as compared to that of other geographic regions. Majority of countries representing Africa, Middle East Asia, North East Asia and Arabian Peninsula included the genotypes that contained rare alleles. As expected, spontaneum accessions, as compared to vulgare accessions, showed a higher number of total alleles, higher number of alleles per locus, higher effective number of alleles and higher allelic richness and a higher number of rare alleles were observed. In summary, the examined ICARDA germplasm set showed ample natural genetic variation that can be harnessed for future breeding of barley as climate change and sustainability have become important throughout all growing areas of the world, drought/heat tolerance being the most important ones.     

PCR-based molecular markers for the fragrance gene in rice (Oryza sativa. L.)

The genomic DNA clone RG28, linked to the major fragrance gene of rice (fgr), was assessed for polymorphism in order to produce a PCR-based marker for fragrance. A small mono-nucleotide repeat, that was polymorphic between a pair of fragrant and non-fragrant cultivars, was identified and developed into a co-dominant PCR-based marker. The polymorphism-information-content determinations for three microsatellite markers, that have been genetically mapped near RG28, are also presented. These PCR-based markers will be highly useful in distinguishing fragrance-producing alleles from non-fragrance-producing alleles at the fgr locus. Received: 19 October 1999 / Accepted: 16 December 1999     

Novel microsatellite markers for Dalechampia scandens (Euphorbiaceae) and closely related taxa: application to studying a species complex

We developed novel microsatellite markers for D alechampia scandens L. (Euphorbiaceae). The target plants belong to a distinct, but undescribed, species in the D . scandens species complex, characterized by small resin‐producing glands. In total, 110 alleles over 36 novel markers were identified across 39 individuals from three populations. The number of alleles varied from one to seven, with an average of 3.06 ± 0.26 alleles per locus. The developed markers, along with previously developed ones for a large‐glanded D . scandens species, were tested for amplification in 11 additional species of the genus D alechampia. Four markers did not produce any detectable allele in 37 individuals from two populations of the large‐glanded species. Average expected heterozygosity across all small‐ and large‐glanded specific loci was 0.36 and 0.15, for the small and large glanded populations, respectively. Cross‐species amplification showed that 89% of all markers were successfully amplified in at least one of the 11 other D alechampia species. These microsatellite markers may be useful for detecting undescribed species in the D . scandens species complex, and can be used for comparative analyses of genetic structure, mating system and phylogeography of other D alechampia species.     

Isolation and characterization of new microsatellite markers in mango (Mangifera indica)

Six microsatellite loci that were isolated from a microsatellite‐enriched genomic library of mango (Mangifera indica) along with their specific primer sets were each characterized by using 36 cultivars collected mainly in Thailand. The observed and expected heterozygosities ranged from 0 to 0.83 and from 0.29 to 0.73, respectively. The number of putative alleles are two to six. Three of the six alleles have frequencies of over 75%. The high frequency may be attributed to the bias in the origin of cultivars. Among 36 mango cultivars tested, 29 cultivars showed a unique pattern by six primer sets, whereas seven cultivars cannot be identified because of genotype similarities. This suggests the potentials for identification of mango cultivars by microsatellite markers.     

Genetic characterization of four strains of Nile tilapia (Oreochromis niloticus L.) using microsatellite markers

Four domesticated strains of Nile tilapia (Oreochromis niloticus L.) were genetically characterized using 14 microsatellite markers and 64 animals per strain. Two strains, Chitralada (AIT) and International Development Research Centers (IDRC) were obtained from the AIT institute, Bangkok, Thailand. The GIFT strain (5th generation) came from NAGRI, Thailand, and the GÖTT strain was supplied by the University of Göttingen, Germany. The average numbers of alleles per marker were 5.0 (GÖTT), 5.4 (AIT), 5.6 (IDRC) and 7.5 (GIFT). Private alleles were found at all markers with the exception of two. No fixation of alleles was found at any marker. Population differentiation, F_ST, was 0.178 (great genetic differentiation) and confirmed grouping of the animals in strains. The expected level of heterozygosity ranged from 0.624 to 0.711, but the observed level of heterozygosity significantly deviated from the expected level in three strains. This was probably because of small population size. Moderate to great genetic differentiation was found between strains. A phylogenetic tree reflected the strains known histories. Application of the Weitzman approach showed that all strains have added value for the total genetic diversity and thus should be retained.     

Introgression of the Vf source of scab resistance and distribution of linked marker alleles within the Malus gene pool

A chromosomal region originating from Malus floribunda 821 confers Vf scab resistance to many isolates of Venturia inaequalis. Twelve DNA markers located in this region were used to scan the equivalent of 31 cM in 98 Malus accessions. This allowed a molecular diagnosis of a source of resistance in apple germplasm with the aid of pedigree information, and in the context of a limited marker survey representing other chromosomes. At least five marker alleles were present in all scab-resistant breeding selections or varieties arising from M. floribunda. The validity of findings based on RAPD markers was confirmed with SCAR assays and Southern-hybridisation experiments. The order of markers determined in previous mapping studies was confirmed and sets of recombinants identified that establish reliable fine-mapping orders within 0.7 cM of the resistance locus. None of the marker alleles were present in the accessions that are either susceptible or possess weak polygenic resistance to scab. The presence of some alleles corresponding to those present at least 5.3 cM from Vf in M. floribunda was detected in some accessions. Other major sources of scab resistance do not appear to possess alleles in common with the Vf region, which will simplify future allelism tests. The results are discussed in the context of the introgression of resistance loci together with marker-assisted selection. The use of breeding pedigrees to assist in fine-scale mapping and map-based cloning is discussed. Received: 16 February 1999 / Accepted: 11 March 1999     

Development of novel microsatellite markers for the grassland species Lolium multiflorum,Lolium perenne and Festuca pratensis

Twelve microsatellite markers were isolated from Lolium multiflorum. Allelic variability and cross‐species amplification were assessed on 16 individuals of each of the three grassland species L. multiflorum, Lolium perenne and Festuca pratensis. Cross‐species amplification success was 100% for L. perenne and 83% for F. pratensis. The number of alleles detected ranged from one to 14 with an average of 3.4. While three microsatellite loci were polymorphic in all three species, one marker produced species‐specific alleles in all three species. These microsatellite markers provide a valuable tool for population genetic studies within and among species of the Festuca–Lolium complex.