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PubMed2002

High-throughput targeted SSR marker development in peach (Prunus persica).

Simple sequence repeats (SSRs) have proven to be highly polymorphic, easily reproducible, codominant markers. However, developing an SSR map is very time consuming and expensive, and most SSRs are not specifically linked to gene loci of immediate interest. The ideal situation would be to combine a high-throughput, relatively inexpensive mapping technique with rapid identification of SSR loci in mapped regions of interest. For this reason, we coupled the high-throughput technique of AFLP mapping with subsequent direct targeting of SSRs identified in AFLP-marked regions of interest. This approach relied on the availability of peach bacterial artificial chromosome (BAC) library resources. We present examples of using this strategy to rapidly identify SSR loci tightly linked to two important, simply inherited traits in peach (Prunus persica (L.) Batsch): root-knot nematode resistance and control of the evergrowing trait. SSRs developed in this study were also tested for their transportability in other Prunus species and in apricots.

Alleles↗

PubMed2002

An expanded genetic linkage map of Prunus based on an interspecific cross between almond and peach.

The genetic linkage map of Prunus constructed earlier and based on an interspecific F2 population resulting from a cross between almond (Prunus dulcis D.A. Webb) and peach (Prunus persica L. Batsch) was extended to include 8 isozyme loci, 102 peach mesocarp cDNAs, 11 plum genomic clones, 19 almond genomic clones, 7 resistance gene analogs (RGAs), 1 RGA-related sequence marker, 4 morphological trait loci, 3 genes with known function, 4 simple sequence repeat (SSR) loci, 1 RAPD, and 1 cleaved amplified polymorphic sequence (CAP) marker. This map contains 161 markers placed in eight linkage groups that correspond to the basic chromosome number of the genus (x = n = 8) with a map distance of 1144 centimorgans (cM) and an average marker density of 6.8 cM. Four more trait loci (Y, Pcp, D, and SK) and one isozyme locus (Mdh1) were assigned to linkage groups based on known associations with linked markers. The linkage group identification numbers correspond to those for maps published by the Arús group in Spain and the Dirlewanger group in France. Forty-five percent of the loci showed segregation distortion most likely owing to the interspecific nature of the cross and mating system differences between almond (obligate outcrosser) and peach (selfer). The Cat1 locus, known to be linked to the D locus controlling fruit acidity, was mapped to linkage group 5. A gene or genes controlling polycarpel fruit development was placed on linkage group 3, and control of senesced leaf color (in late fall season) (LFCLR) was mapped to linkage group 1 at a putative location similar to where the Y locus has also been placed.

Calcium-Binding Proteins↗

PubMed2002

Molecular linkage maps of the Populus genome.

We report molecular genetic linkage maps for an interspecific hybrid population of Populus, a model system in forest-tree biology. The hybrids were produced by crosses between P. deltoides (mother) and P. euramericana (father), which is a natural hybrid of P. deltoides (grandmother) and P. nigra (grandfather). Linkage analysis from 93 of the 450 backcross progeny grown in the field for 15 years was performed using random amplified polymorphic DNAs (RAPDs), amplified fragment length polymorphisms (AFLPs), and inter-simple sequence repeats (ISSRs). Of a total of 839 polymorphic markers identified, 560 (67%) were testcross markers heterozygous in one parent but null in the other (segregating 1:1), 206 (25%) were intercross dominant markers heterozygous in both parents (segregating 3:1), and the remaining 73 (9%) were 19 non-parental RAPD markers (segregating 1:1) and 54 codominant AFLP markers (segregating 1:1:1:1). A mixed set of the testcross markers, non-parental RAPD markers, and codominant AFLP markers was used to construct two linkage maps, one based on the P. deltoides (D) genome and the other based on P. euramericana (E). The two maps showed nearly complete coverage of the genome, spanning 3801 and 3452 cM, respectively. The availability of non-parental RAPD and codominant AFLP markers as orthologous genes allowed for a direct comparison of the rate of meiotic recombination between the two different parental species. Generally, the rate of meiotic recombination was greater for males than females in our interspecific poplar hybrids. The confounded effect of sexes and species causes the mean recombination distance of orthologous markers to be 11% longer for the father (P. euramericana; interspecific hybrid) than for the mother (P. deltoides; pure species). The linkage maps constructed and the interspecific poplar hybrid population in which clonal replicates for individual genotypes are available present a comprehensive foundation for future genomic studies and quantitative trait locus (QTL) identification.

Chi-Square Distribution↗

PubMed2002

Molecular and physical mapping of a barley gene on chromosome arm 1 HL that causes sterility in hybrids with wheat.

Addition of the long arm of barley chromosome 1H (1HL) to wheat causes severe meiotic abnormalities and complete sterility of the plants. To map the barley gene responsible for the 1H-induced sterility of wheat, a series of addition lines of translocated 1H chromosomes were developed from the crosses between the wheat 'Shinchunaga' and five reciprocal translocation lines derived from the barley line St.13559. Examination of the seed fertility of the addition lines revealed that the sterility gene is located in the interstitial 25% region of the 1HL arm. The genetic location of the sterility gene was also estimated by physically mapping sequence-tagged site (STS) markers and simple-sequence repeat (SSR) markers with known map locations. The sterility gene is designated Shw (sterility in hybrids with wheat). Comparison of the present physical map of 1HL with two previously published genetic maps revealed a paucity of markers in the proximal 30% region and non-random distribution of SSR markers. Two inconsistencies in marker order were found between the present physical map and the consensus genetic map of group 1 chromosomes of Triticeae. On the basis of the effects on meiosis and chromosomal location, the relationship of the present sterility gene with other fertility-related genes of Triticeae is discussed.

Genetic Markers↗

PubMed2002

Identification of putative genes in bean (Phaseolus vulgaris) genomic (Bng) RFLP clones and their conversion to STSs.

A set of 79 previously mapped bean (Phaseolus vulgaris) genomic (Bng) clones were partially sequenced. BLAST database searches detected homologies between 59 of these clones and genes from a variety of plants, especially Arabidopsis thaliana. Some matches in the database to the Bng clones included a putative P-glycoprotein-ABC transporter from Arabidopsis, an early nodulin-binding protein (ENBPI) from Medicago truncatula, a lon-protease protein from spinach, a branched-chain amino-acid aminotransferase from Arabidopsis, and a vacuolar sorting receptor (BP-80) from Pisum sativum. Additional matches were found for genes involved in isoprenoid biosynthesis, sulfur metabolism, proline biosynthesis, and floral development. Sequence tagged site (STSs) were produced for 16 of the clones, 2 of which contain simple sequence repeats (SSRs). Polymorphisms were detected for six of the STSs.

Base Sequence↗

PubMed2003

Characterization of microsatellites in wild and sweet cherry (Prunus avium L.)--markers for individual identification and reproductive processes.

Nuclear microsatellites were characterized in Prunus avium and validated as markers for individual and cultivar identification, as well as for studies of pollen- and seed-mediated gene flow. We used 20 primer pairs from a simple sequence repeat (SSR) library of Prunus persica and identified 7 loci harboring polymorphic microsatellite sequences in P. avium. In a natural population of 75 wild cherry trees, the number of alleles per locus ranged from 4 to 9 and expected heterozygosity from 0.39 to 0.77. The variability of the SSR markers allowed an unambiguous identification of individual trees and potential root suckers. Additionally, we analyzed 13 sweet cherry cultivars and differentiated 12 of them. An exclusion probability of 0.984 was calculated, which indicates that the seven loci are suitable markers for paternity analysis. The woody endocarp was successfully used for resolution of all microsatellite loci and exhibited the same multilocus genotype as the mother tree, as shown in a single seed progeny. Hence, SSR fingerprinting of the purely maternal endocarp was also successful in this Prunus species, allowing the identification of the mother tree of the dispersed seeds. The linkage of microsatellite loci with PCR-amplified alleles of the self-incompatibility locus was tested in two full-sib families of sweet cherry cultivars. From low recombination frequencies, we inferred that two loci are linked with the S locus. The present study provides markers that will significantly facilitate studies of spatial genetic variation and gene flow in wild cherry, as well as breeding programs in sweet cherry.

Genetic Linkage↗

PubMed2003

Genetic identification of a female partial-sterile mutant in soybean.

We report here the genetic identification of a female partial-sterile mutant derived from soybean mutant L67-3483. L67-3483, which originated from the cultivar Clark after X-ray irradiation, is male and female fertile. All F1 plants in reciprocal pollinations of L67-3483 with 'Clark', 'Minsoy', or 'BSR 101' were female partial sterile. Partial sterility is expressed in the heterozygous condition at a single locus and upon self-pollination this locus exhibits a 1:1 segregation pattern. This locus is located on the terminus of the soybean molecular linkage group D1b+W, between simple sequence repeat (SSR) markers Satt157 and Satt266, and is linked to each by 5.3 and 1.2 cM, respectively. This gene is transmitted through both female and male gametes and there was no segregation distortion of SSR markers linked to this gene. We concluded that this female partial-sterile gene is a new mutation class, and differs from the previously reported mutation classes in soybean, i.e., sporophytic mutation, gametophytic female-specific mutation, and general gametophytic mutation. Restriction of recombination around the mutant gene suggested that this gene is located near or within (a) small inversion(s) or adjacent to (a) chromosomal deletion(s).

Chromosome Mapping↗

PubMed2003

Mapping quantitative trait loci for a common bean (Phaseolus vulgaris L.) ideotype.

Breeding a model plant that encompasses individual traits thought to enhance yield potential, known as ideotype breeding, has traditionally focused on phenotypic selection of plants with desirable morphological traits. Broadening this breeding method to the molecular level through the use of molecular markers would avoid the environmental interactions associated with phenotypic selection. A population of 110 F5 recombinant inbred lines (RILs), derived from the cross between WO3391 and 'OAC Speedvale', was used to develop a genetic linkage map consisting of 105 random amplified polymorphic DNA (RAPD), simple sequence repeat (SSR), and sequence-tagged site (STS) markers. The map has a total length of 641 cM distributed across 8 linkage groups (LGs). Five of them were aligned on the core linkage map of bean. Twenty-one quantitative trait loci (QTLs) were identified over three environments for eight agronomic and architectural traits previously defined for a bean (Phaseolus vulgaris L.) ideotype. The QTLs were mapped to seven LGs with several regions containing QTLs for multiple traits. At least one QTL was located for each trait and a maximum of four were associated with lodging. Total explained phenotypic variance ranged from 10.6% for hypocotyl diameter to 45.4% for maturity. Some of the QTLs identified will be useful for early generation selection of tall, upright, high-yielding lines in a breeding program.

Analysis of Variance↗

PubMed2003

Linking porcine microsatellite markers to known genome regions by identifying their human orthologs.

Microsatellites, or tandem simple sequence repeats (SSRs), have become one of the most popular molecular markers in genome mapping because of their abundance across genomes and because of their high levels of polymorphism. However, information on which genes surround or flank them has remained very limited for most SSRs, especially in livestock species. In this study, an in silico comparative mapping approach was developed to link porcine SSRs to known genome regions by identifying their human orthologs. From a total of 1321 porcine microsatellites used in this study, 228 were found to have blocks in alignment with human genomic sequences. These 228 SSRs span about 1459 cM of the porcine genome, but with uneven distributions, ranging from 2 on SSC12 to 24 on SSC14. Linking these porcine SSRs to the known genome regions in the human genome also revealed 16 new putative synteny groups between these two species. Fifteen SSRs on SSC3 with identified human orthologs were typed on a pig-hamster radiation hybrid (RH) panel and used in a joint analysis with 80 known gene markers previously mapped on SSC3 using the same panel. The analysis revealed that they were all highly linked to either one or both adjacent markers. These results indicated that assigning the porcine SSRs to known genome regions by identifying their human orthologs is a reliable approach. The process will provide a foundation for positional cloning of causative genes for economically important traits.

Animals↗

PubMed2003

Development and characterization of recombinant chromosome substitution lines (RCSLs) using Hordeum vulgare subsp. spontaneum as a source of donor alleles in a Hordeum vulgare subsp. vulgare background.

The ancestor of barley (Hordeum vulgare subsp. spontaneum) may be a source of novel alleles for crop improvement. We developed a set of recombinant chromosome substitution lines (RCSLs) using an accession of H. vulgare subsp. spontaneum (Caesarea 26-24, from Israel) as the donor and Hordeum vulgare subsp. vulgare 'Harrington' (the North American malting quality standard) as the recurrent parent via two backcrosses to the recurrent parent, followed by six generations of selfing. Here we report (i) the genomic architecture of the RCSLs, as inferred by simple sequence repeat (SSR) markers, and (ii) the effects of H. vulgare subsp. spontaneum genome segment introgressions in terms of three classes of phenotypes: inflorescence yield components, malting quality traits, and domestication traits. Significant differences among the RCSLs were detected for all phenotypes measured. The phenotypic effects of the introgressions were assessed using association analysis, and these were referenced to quantitative trait loci (QTL) reported in the literature. Hordeum vulgare subsp. spontaneum, despite its overall inferior phenotype, contributed some favorable alleles for agronomic and malting quality traits. In most cases, the introgression of the ancestral genome resulted in a loss of desirable phenotypes in the cultivated parent. Although disappointing from a plant breeding perspective, this finding may prove to be a useful tool for gene discovery.

Alleles↗

PubMed2004

Duplicate chlorophyll-deficient loci in soybean.

Three lethal-yellow mutants have been identified in soybean (Glycine max (L.) Merr.), and assigned genetic type collection numbers T218H, T225H, and T362H. Previous genetic evaluation of T362H indicated allelism with T218H and T225H and duplicate-factor inheritance. Our objectives were to confirm the inheritance and allelism of T218H and T225H and to molecularly map the locus and (or) loci conditioning the lethal-yellow phenotype. The inheritance of T218H and T225H was 3 green : 1 lethal yellow in their original parental source germplasm of Glycine max 'Illini' and Glycine max 'Lincoln', respectively. In crosses to unrelated germplasm, a 15 green : 1 lethal yellow was observed. Allelism tests indicated that T218H and T225H were allelic. The molecular mapping population was Glycine max 'Minsoy' x T225H and simple sequence repeat (SSR) markers were used. The first locus, designated y18-1, was located on soybean molecular linkage group B2, between SSR markers Satt474 and Satt534, and linked to each by 4.4 and 13.4 cM, respectively. The second locus, designated y18-2, was located on soybean molecular linkage group D2, between SSR markers Satt543 and Sat-001, and linked to each by 2.2 and 4.4 cM, respectively.

Chlorophyll↗

PubMed2004

Analysis of phylogenetic relationship among five mulberry (Morus) species using molecular markers.

Species identification in mulberry (Morus) continues to be a point of great debate among scientists despite the number of criteria such as floral characters, wood, and leaf anatomical and biochemical characters used to identify the species within this genus. However, no consensus system of classification has emerged. Hence, an investigation was undertaken with inter-simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD) markers to find out the possibility of using these DNA markers to confirm the identity of genotypes in a particular species. Fifteen ISSR and 15 RAPD primers generated 86% and 78% polymorphism, respectively, among 19 mulberry genotypes. The polymorphism among the species varied from 50% to 57% in ISSR markers and 31% to 53% in RAPD markers. Similarity coefficients were higher among the genotypes of M. latifolia, M. bombycis and M. alba. Cluster analyses separated genotypes of M. laevigata and M. indica from those of the other species. Population structure analysis of these species further showed high genetic differentiation coefficients (GST), high heterozygosity between two species (DST), and total heterozygosity among populations (Ht) coupled with considerably low gene flow (Nm) when M. laevigata was paired with other species. Based on these parameters and the result of cluster analysis it is concluded that M. laevigata can be considered as a separate species of mulberry, whereas the other four species may be grouped together and treated as subspecies.

Genetic Markers↗

PubMed2004

Development of SSR markers for the phylogenetic analysis of almond trees from China and the Mediterranean region.

Expressed sequence tag (EST) derived simple sequence repeats (SSRs, microsatellites) were screened and identified from 3863 almond and 10 185 peach EST sequences, and the spectra of SSRs in the non-redundant EST sequences were investigated after sequence assembly. One hundred seventy-eight (12.07%) almond SSRs and 497 (9.97%) peach SSRs were detected. The EST-SSR occurs every 4.97 kb in almond ESTs and 6.57 kb in peach, and SSRs with di- and trinucleotide repeat motifs are the most abundant in both almond and peach ESTs. Twenty one EST-SSRs were thereafter, developed and used together with 7 genomic SSRs, to study the genetic relationship among 36 almond (P. communis Fritsch.) cultivars from China and the Mediterranean area, as well as 8 accessions of other related species from the genus Prunus. Both EST-derived and genomic SSR markers showed high cross-species transferability in the genus. Out of the 112 polymorphic alleles detected in the 36 cultivated almonds, 28 are specific to Chinese cultivars and 25 to the others. The 44 accessions were clustered into 4 groups in the phylogenetic tree and the 36 almond cultivars formed two distinct subgroups, one containing only Chinese cultivars and one of unknown origin and the other only those originating from the Mediterranean area, indicating that Chinese almond cultivars have a distinct evolutionary history from the Mediterranean almond. Our preliminary results indicated that common almond was more closely related to peach (P. persica (L.) Batsch.) than to the four wild species of almond, (P. mongolica Maxim., P. ledebouriana Schleche, P. tangutica Batal., and P. triloba Lindl.). The implications of these SSR markers for evolutionary analysis and molecular mapping of Prunus species are discussed.

Alleles↗

PubMed2004

High level of genetic diversity among spelt germplasm revealed by microsatellite markers.

The genetic diversity of spelt (Triticum aestivum (L.) Thell. subsp. spelta (L.) Thell.) cultivated presently is very narrow. Although the germplasm collections of spelt are extensive, the related genetic knowledge is often lacking and makes their use for genetic improvement difficult. The genetic diversity and structure of the spelt gene pool held in gene banks was determined using 19 simple sequence repeat (SSR) markers applied to 170 spelt accessions collected from 27 countries and 4 continents. The genetic distances (1 - proportion of shared alleles) were calculated and an unweighted pair-group method with arithmetic averaging (UPGMA)-based dendrogram was generated. The genetic diversity was high: 259 alleles were found and the mean interaccession genetic distance was 0.782 +/- 0.141. The dendrogram demonstrated the much higher genetic diversity of spelt held in germplasm collections than in the currently used genotypes. Accessions with the same geographical origin often tended to cluster together. Those from the Middle East were isolated first. All but one of the Spanish accessions were found in a unique subcluster. Most accessions from eastern Europe clustered together, while those from northwestern Europe were divided into two subclusters. The accessions from Africa and North America were not separated from the European ones. This analysis demonstrates the extent of genetic diversity of spelts held in germplasm collections and should help to widen the genetic basis of cultivated spelt in future breeding programs.

Alleles↗

PubMed2005

Mapping of genes expressed in Fusarium graminearum-infected heads of wheat cultivar 'Frontana'.

The isolation, physical, and genetic mapping of a group of wheat genes expressed in infected heads of Triticum aestivum 'Frontana' resistant to Fusarium head blight is reported. A cDNA library was built from heads of 'Frontana' through suppressive subtractive hybridization, to enrich for sequences induced by the pathogen Fusarium graminearum during infection. A group of 1794 clones was screened by dot blot hybridization for differential gene expression following infection. Twenty of these clones showed a strong difference in intensity of hybridization between infected and mock-inoculated wheat head samples, suggesting that they corresponded to genes induced during infection. The 20 clones were sequenced and used for mapping analysis. We determined a precise chromosomal location for 14 selected clones by using series of chromosome deletion stocks. It was shown that the 14 clones detected 90 fragments with the use of the restriction enzyme EcoRI; 52 bands were assigned to chromosome bins, whereas 38 fragments could not be assigned. The selected clones were also screened for polymorphisms on a 'Wuhan' x 'Maringa' wheat doubled haploid mapping population. One clone, Ta01_02b03, was related to a quantitative trait locus for type II resistance located on chromosome 2AL, as determined with simple sequence repeat markers on another mapping population, but did not map in the same location on our population. Another clone, Ta01_06f04, was identified by BLAST (basic local alignment search tool) search in public databases to code for a novel beta-1,3-glucanase, homologous to a major pathogenesis-related protein. This clone mapped to chromosomal regions on chromosome 3, including 3BL and 3DL, where B glucanase gene clusters are known to exist. Seven other clones, including 1 coding for an ethylene-response element binding protein and 3 for ribosomal proteins, and 4 clones corresponding to proteins with unknown function, were also mapped.

Chromosome Mapping↗

PubMed2005

Genetic analysis and molecular mapping of a pale flower allele at the W4 locus in soybean.

In soybean (Glycine max (L.) Merr.), the w4-mutable line that harbors the w4-m allele was identified in 1983. It was proposed that this line contained an autonomous transposable element at the W4 locus, which is a major locus controlling the biosynthesis of anthocyanin. The w4-m allele can revert to the W4 allele that produces the wild-type phenotype, or sometimes to other alleles that produce intermediate phenotypes. Mutant plants that produce pale flowers were identified among the progeny of a single germinal revertant event from the w4-mutable line. Through genetic analysis, we established that the pale-flower mutation was conditioned by a new allele (w4-p) at the W4 locus. The w4-p allele is dominant to the w4 allele but recessive to the W4 allele, and the w1 allele has an epistatic effect on the w4-p allele. The pale-mutant line (w4-pw4-p) was designated as Genetic Type Collection number T369. An F2 mapping population derived from the cross of Minsoy (W4W4) x T369 (w4-pw4-p) was used to map the W4/w4-p locus, using simple sequence repeat (SSR) markers. The W4 locus was located at one end of molecular linkage group D2, 2.3 cM from the SSR marker Satt386 and close to the nearby telomere.

Alleles↗

PubMed2005

Development and characterization of microsatellite markers in Cynara cardunculus L.

Cynara cardunculus L. is a species native to the Mediterranean basin that comprises 2 crops, globe artichoke (var. scolymus L.) and cultivated cardoon (var. altilis DC), as well as wild cardoon (var. sylvestris (Lamk) Fiori). Globe artichoke represents an important component of the South European agricultural economy but is also cultivated in North Africa, the Near East, South America, the United States, and China. Breeding activities and molecular marker studies have been, to date, extremely limited. Better knowledge of the genome of the species might be gained by developing a range of molecular markers. Here, we report on the development of 14 microsatellites (simple sequence repeats (SSRs)) through a novel approach that we have defined as the microsatellite amplified library (MAL). The approach represents a combination of amplified fragment length polymorphism and a primer extension based enriched library, is rapid, and requires no hybridization enrichment steps. The technique provided a approximately 40-fold increase in the efficiency of SSR identification compared with conventional library procedures. The developed SSRs were applied for genotyping 36 accessions of C. cardunculus, including a core of 27 varietal types of globe artichoke, 3 accessions of cultivated cardoon, and 6 Sicilian accessions of wild cardoon. Principal coordinates analysis made it possible to differentiate both cultivated and wild forms from each other.

Cynara↗

PubMed2005

Application of ISSR, RAPD, and cytological markers to the certification of Picea mariana, P. glauca, and P. engelmannii trees, and their putative hybrids.

Picea glauca (white spruce) and P. engelmannii (Engelmann spruce) are so similar and integrated that it is impossible to distinguish between them and their hybrids using morphological characteristics. Although natural hybrids between P. glauca and P. mariana (black spruce) do not generally occur, even though the 2 species are sympatric in North America, a first-generation hybrid, called the Rosendahl spruce, has been reported in the literature. In this study, several inter-simple sequence repeat (ISSR) markers were developed, as were randomly amplified polymorphic DNA (RAPD) markers, to certify spruce trees and their hybrids. ISSR fingerprinting was more efficient than RAPD assay; it detected 70% polymorphic DNA markers among the spruce species analyzed, whereas RAPD fingerprinting detected only 53%. Species-diagnostic ISSR and RAPD markers differentiating P. glauca from P. engelmannii and P. mariana were cloned and sequenced. Molecular certification of the spruce samples analyzed confirmed that all the seeds from interior spruce populations were true hybrids of P. glauca and P. engelmannii. But the analysis of seeds derived from the putative Rosendahl spruce indicated that this tree is likely a pure P. glauca genotype, rather than a hybrid of P. glauca and P. mariana. These data were confirmed by cytological analyses. Further analysis, using a more sensitive DNA amplification method with designed primers flanking the species-diagnostic ISSR and RAPD markers, revealed that such sequences are not generally species-specific because they are present in other spruce species.

Base Sequence↗