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Riaz Ahmad, Louise Ferguson, and Stephen M. Southwick

A genomic DNA library enriched for dinucleotide (CT)n and (CA)n and trinucleotide (CTT)n microsatellite motifs has been developed from `Kerman' pistachio (Pistacia vera L.). The enrichment method based on magnetic or biotin capture of repetitive sequences from restricted genomic DNA revealed an abundance of simple sequence repeats (SSRs) in the pistachio genome which were used for marker development. After an enrichment protocol, about 64% of the clones contained (CT)n repeats while 59% contained (CA)n for CT and CA enriched libraries, respectively. In the (CT)n enriched library, compound sequences were 45% while for (CA)n it was 13.5%. In both dinucleotide enriched libraries, about 80% of the clones having microsatellites have a repeat length in the range of 10 to 30 units. A library enriched for trinucleotide (CTT)n contained <19% of the clones with (CTT)n repeats. Of the clones that contained microsatellites, 62% had sufficient flanking sequence for primer design. An initial set of 25 pairs of primers was designed, out of which 14 pairs amplified cleanly and produced an easily interpretable PCR product in the commercially important American, Iranian, Turkish, and Syrian pistachio cultivars. The efficient DNA extraction method developed for pistachio kernels and shells (roasted and nonroasted) yielded DNA of sufficient quality to use PCR to create DNA fingerprints. In total, 46 alleles were identified by 14 primer pairs and a dendrogram was constructed on the basis of that information. The SSR markers distinguished most of the tested cultivars from their unique DNA fingerprint. An UPGMA cluster analysis placed most of the Iranian samples in one group while the Syrian samples were the most diverse and did not constitute a single distinct group. The maximum number of cultivar specific markers were found in `Kerman'(4), the current industry standard in the United States, and the Syrian cultivar Jalab (5). The technique of using extracted DNA from pistachio kernal or shell coupled with the appropriate marker system developed here, can be used for analyses and measurement of trueness to type.

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Phillip A. Wadl, Robert N. Trigiano, Dennis J. Werner, Margaret R. Pooler, and Timothy A. Rinehart

markers that are randomly dispersed throughout the nuclear genome have the required distribution and frequency to provide information at the population and single plant levels. Simple sequence repeat (SSR) markers are widely used because of their high

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Jaeho Yoon, Dongcheng Liu, Wonseob Song, Weisheng Liu, Aimin Zhang, and Shaohua Li

The genetic relationships among 96 peach and nectarine [Prunus persica (L.) Batsch.] genotypes and botanical varieties originating from different ecogeographical regions of China, Japan, North America, and South Korea were evaluated with 33 SSR markers screened from 108 published SSR markers developed for peach or sweet cherry (P. avium L.). The 33 SSRs detected polymorphisms among 96 genotypes and revealed a total of 283 alleles with an average of 8.6 alleles per locus. The polymorphism information content (PIC) value ranged from 0.40 (BPPCT041) to 0.98 (BPPCT009) with an average of 0.80. Unweighted pair group method average (UPGMA) cluster analysis based on Nei's genetic distances classified genotypes into six groups, corresponding to their ecogeographical origin. Group I consisted of northern Chinese and northwestern Chinese local cultivars, and was divided into two subgroups, white and yellow peaches. Group II contained mainly southern Chinese local, Japanese, and North American cultivars and can be divided into four subgroups: Japanese white, Chinese flat, North American yellow, and some Chinese local ornamental peach cultivars. Groups III, IV, and V were comprised of Chinese local ancient cultivars, and contained `Xinjiangdatianren' and `Renmiantao', Chinese dwarf cultivars, and `Fenshouxing', respectively. Group VI had only `Baishanbitao', a Chinese ornamental cultivar. Northern and northwestern Chinese local cultivars clustered together with a greater diversity than southern Chinese local cultivars, indicating that the northern and northwestern Chinese local cultivars are similar ecotypes, and southern Chinese local cultivars are a subset of the northern Chinese group. Moreover, the Japanese and North American genotypes had a close phylogenetic relationship with southern Chinese local cultivars. The taxonomic placement of P. ferganensis (Kost. et Kiab) Kov. et Kost. and the phylogenetic relationship of `Baishanbitao' with peaches are discussed.

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K.S. Lewers, S.M.N. Styan, S.C. Hokanson, and N.V. Bassil

Although simple sequence repeat (SSR) markers have been developed for species in the closely related genera Fragaria L. (strawberry) and Rubus L. (raspberry and blackberry), the number of SSRs available is insufficient for genetic mapping. Our objective was to use and compare multiple approaches for developing additional SSRs for Fragaria and Rubus. The approaches included: the development of SSRs from GenBank sequences from species of varied relatedness to Fragaria and Rubus and identified with two different data-mining methods (BLAST and SSRIT); the evaluation of some previously published SSRs designed from related species; and the development of SSRs from a genomic library made from F. ×ananassa Duschene ex Rozier `Earliglow'. When an SSR was developed from a known gene sequence, the location of the repeat in the gene was determined to evaluate the effect on amplification and polymorphism detection. Cross-generic amplification between closely related Fragaria and Rubus as well as transference from species of varied relatedness to Fragaria and Rubus also was evaluated and indicated limited transference within the subfamily Rosoideae. However, development of SSRs for Fragaria and Rubus from Rosa L. (rose) and Rosaceae genera outside Rosoideae was not efficient enough to be practical for new map development. SSRIT was superior to BLAST for identifying GenBank sequences containing repeats. SSRs developed from repeats found in either the 5′UTR (80% polymorphic) or 3′UTR (85% polymorphic) were most likely to detect polymorphisms, compared with those developed from coding regions (30%). SSRs developed from the genomic library were only slightly superior to GenBank-derived SSRs in their ability to detect polymorphisms.

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Kirk W. Pomper, Jeremiah D. Lowe, Li Lu, Sheri B. Crabtree, Shandeep Dutta, Kyle Schneider, and James Tidwell

geographic range, insect-pollinated outcrossing breeding systems, secondary asexual reproduction, and animal-dispersed seed. Microsatellites, or simple sequence repeats (SSRs), are a marker of choice for genetic diversity estimates, genetic mapping, and DNA

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Xiaoxu Yang, Yinshan Guo, Junchi Zhu, Zaozhu Niu, Guangli Shi, Zhendong Liu, Kun Li, and Xiuwu Guo

many peer-reviewed research articles ( Chacón et al., 2012 ; Chang et al., 2014 ; Ghaste et al., 2015 ; Maoz et al., 2016 ). Grape genetics, genomics studies, and simple sequence repeat (SSR) analysis have provided an efficient means of assessing

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Ying Wang, Ming Kang, and Hongwen Huang

critical, and a set of molecular markers is also required. Microsatellites or simple sequence repeats (SSRs) are DNA segments containing short sequence repeats (1–6 nucleotides) and have been widely recognized as powerful and informative markers due to

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A. Levi and L.J. Rowland

Fifteen highbush (or highbush hybrid) blueberry cultivars (Vaccinium corymbosum Linnaeus), two rabbiteye blueberry cultivars (V. ashei Reade), and one southern lowbush (V. darrowi Camp) selection from the wild were examined using seventeen 10-base RAPD and seven 15- to 18-base SSR-anchored primers (primers comprised of SSR motifs) in polymerase chain reactions (PCRs). Fifteen RAPD and three SSR markers resulting from these reactions were chosen to construct a DNA fingerprinting table to distinguish among the genotypes included in this study. Similarity values were calculated based on 132 RAPD and 51 SSR bands, and a dendrogram was constructed based on the similarity matrix. The V. ashei cultivars and V. darrowi selection grouped out separately from the V. corymbosum cultivars as expected. However, estimates of relative genetic similarity between genotypes within the V. corymbosum group did not agree well with known pedigree data and, thus, indicated that RAPD and SSR data did not accurately assess the genetic relationships of cultivars within this species.

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Thomas M. Davis, Laura M. DiMeglio, Ronghui Yang, Sarah M.N. Styan, and Kim S. Lewers

The cultivated strawberry, Fragaria ×ananassa Duchesne ex Rozier, originated via hybridization between octoploids F. chiloensis (L.) Mill. and F. virginiana Mill. These three octoploid species are thought to share a putative genome composition of AAA`A'BBB`B'. Diploid F. vesca L., is considered to have donated the A genome. Current attention to the development of a diploid model system for strawberry genomics warrants the assessment of simple sequence repeat (SSR) marker transferability between the octoploid and diploid species in Fragaria L. In the present study, 23 SSR primer pairs derived from F. ×ananassa `Earliglow' by genomic library screening were evaluated for their utility in six diploid Fragaria species, including eight representatives of F. vesca, four of F. viridis Weston, and one each of F. nubicola (Hook. f.) Lindl. ex Lacaita, F. mandshurica Staudt, F. iinumae Makino, and F. nilgerrensis Schltdl. ex J. Gay. SSR primer pair functionality, as measured by amplification success rate (= 100% - failure rate) in each species, was ranked (from highest to lowest) as follows: F. vesca (98.4%) > F. iinumae (93.8%) = F. nubicola (93.8%) > F. mandshurica (87.5%) > F. nilgerrensis (75%) > F. viridis (73.4%). The extent to which these octoploid-derived SSR primer pairs generated markers that could be added to the F. vesca linkage map also was assessed. Of the 13 F. ×ananassa SSR markers that segregated codominantly in the F. vesca mapping population, 11 were assigned to linkage groups based upon close linkages to previously mapped loci. These markers were distributed over six of the seven F. vesca linkage groups, and can serve as anchor loci defining these six groups for purposes of comparative mapping between F. vesca and F. ×ananassa.

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Innocenzo Muzzalupo, Francesca Stefanizzi, and Enzo Perri

abundance, enormous extent of allelic diversity, and the ease of assessing simple sequence repeat (SSR) size variation by PCR with pairs of flanking primers ( Agarwal et al., 2008 ; Muzzalupo et al., 2006 , 2008 ; Rekik et al., 2008 ). Recently, many