Linkage Analysis with Multiplexed Short Tandem Repeat Polymorphisms Using Infrared Fluorescence and M13 Tailed Primers

Oetting, William S.; Lee, H. K.; Flanders, Dean; Wiesner, Georgia L.; Sellers, Thomas A.; King, Rodney A.

doi:10.1006/geno.1995.1264

Cited by 337 publications

(262 citation statements)

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“…The primers were selected from all 21 chromosomes of hexaploid wheat. The PCR was performed in a volume of 10 mL containing 2 mL of DNA (20 ng/mL) and 8 mL of the master mix that consisted of 3.2 mL of ddH 2 0, 1 mL of 10 Â PCR buffer, 1.1 mL of 25 mM MgCl 2 , 0.5 mL of 5 mM dNTPs, 0.5 mL of each forward and reverse primer (1 p.m./mL), 1 mL of tailed fluorescence-labelled (IRD-700 or IRD-800) M13 primer (Oetting et al 1995), and 0.2 mL of Taq polymerase (5 U/mL). The PCR was performed by using a touchdown universal program consisting of 5 cycles of denaturing at 958C for 45 s, annealing at 688C for 5 min decreasing by 28C in each subsequent cycle, and extension at 728C for 1 min.…”

Section: Ssr Analysismentioning

confidence: 99%

Genetic diversity in the U.S. hard red winter wheat cultivars as revealed by microsatellite markers

Prasad¹,

Babar

et al. 2009

Crop Pasture Sci.

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Abstract. Knowledge of the genetic diversity existing in previously released hard red winter wheat (HRWW, Triticum aestivum L.) cultivars in the Great Plains region, United States, is essential for effective utilisation of these genetic resources in the various HRWW breeding programs. To ascertain a measure of the genetic diversity of the existing US HRWW, 60 cultivars were analysed with 62 microsatellite markers distributed throughout the wheat genome. Marker data were subjected to distance-based analysis and analysis of molecular variances. In total, 341 polymorphic alleles were scored with a range of 2-12 alleles per locus. Genetic diversity gradually increased in cultivars released after the 1970s. Cultivars released in the 1990s had the highest allelic richness (4.79), gene diversity (0.60), and polymorphic information content (0.56). Levels of genetic diversity were similar between the major HRWW breeding programs. Cluster analysis resulted in eight clusters. Cluster grouping gave close matches with pedigrees and with regional distribution of the cultivars. Using decadal information, cultivars released from 1900-1969 were grouped into one cluster, cultivars from 1990-2005 were grouped into a separate cluster, whereas cultivars from the 1980s did not group with any other decades. Analysis of molecular variance revealed a significant variation among the clusters, signifying that a true genetic variation existed among the clusters. The higher proportion of genetic variation explained by cultivars within clusters compared with among clusters indicates greater genetic diversity among cultivars within clusters. Our results indicate that genetic diversity of Great Plains HRWW cultivars has increased in the past century, and the trend is continuing.

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Section: Ssr Analysismentioning

confidence: 99%

Genetic diversity in the U.S. hard red winter wheat cultivars as revealed by microsatellite markers

Prasad¹,

Babar

et al. 2009

Crop Pasture Sci.

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“…From these sequences, primer pairs were designed by Primer 3.0 (Rozen and Skaletsky 2000), http://frodo.wi.mit.edu/ cgi-bin/primer3/primer3_www.cgi), adopting default parameter settings. A tailing primer strategy was used, as described by Oetting et al (1995). The newly developed SSR markers were identified by a number, prefixed by CELMS (Cynara Enriched Library MicroSatellite) ( Table 2).…”

Section: Methodsmentioning

confidence: 99%

Genetic mapping and annotation of genomic microsatellites isolated from globe artichoke

et al. 2009

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Cynara cardunculus includes three taxa, the globe artichoke (subsp. scolymus L. Hegi), the cultivated cardoon (var. altilis) and their progenitor, the wild cardoon (var. sylvestris). Globe artichoke is an important component of the Mediterranean rural economy, but its improvement through breeding has been rather limited and its genome organization remains largely unexplored. Here, we report the isolation of 61 new microsatellite loci which amplified a total of 208 alleles in a panel of 22 C. cardunculus genotypes. Of these, 51 were informative for linkage analysis and 39 were used to increase marker density in the available globe artichoke genetic maps. Sequence analysis of the 22 loci associated with genes showed that 9 are located within coding sequence, with the repetitive domain probably being involved in DNA binding or in protein-protein interactions. The expression of the genes associated with 9 of the 22 microsatellite loci was demonstrated by RT-PCR.

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“…Oetting et al (1995) were the first group to use the fluorescent method for genotyping purposes and used five primer pairs to perform different multiplex amplification reactions. The technique was based on a fluorescent-labeled universal M13 primer, which allowed automated DNA sequencer analysis and subsequent genotyping with high precision of allele size.…”

Section: Adaptação Da Técnica De Fluorescência Para Fins De Genotipagmentioning

confidence: 99%

“…In fact, beans are a good model to work with due to their diploid nature (2n = 22) and small genome (Broughton et al, 2003;Schlueter et al, 2008). Microsatellite markers have been developed for beans from published sequences and from microsatellite-enriched libraries (Gepts et al, 2008).…”

Section: Adaptação Da Técnica De Fluorescência Para Fins De Genotipagmentioning

confidence: 99%

Adaptation of fluorescent technique for genotyping with new microsatellite markers in common bean

Oblessuc

Campos

Cardoso

et al. 2009

Pesq. agropec. bras.

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-The objectives of this work were to adapt the fluorescent labeling polymerase chain reaction (PCR) technique using M13 universal primer for genotyping purposes, and to present a new set of microsatellite markers for common bean (Phaseolus vulgaris L.). A large population (380 common bean lines) was used for microsatellite genotyping. PCR fluorescent labeling method showed to be very efficient for multiplex analysis, providing lower costs and saving time, thus increasing the quality of genotyping analysis. A new set of 50 microsatellites developed from an enriched library derived from cultivar IAC-UNA was presented. This study provides better tools for assisting common bean breeding programs.Index terms: Phaseolus vulgaris, fluorescence, molecular marker, multiplex analysis. Adaptação da técnica de fluorescência para fins de genotipagem com novos marcadores microssatélite em feijoeiroResumo -Os objetivos deste trabalho foram adaptar a técnica de marcação fluorescente de produtos da reação em cadeia da polimerase (PCR) com uso do iniciador universal M13, para aplicação em genotipagem, e apresentar novos marcadores microssatélite para o feijoeiro (Phaseolus vulgaris L.). Uma população de grande tamanho amostral (380 linhagens) foi utilizada para genotipagem dos microssatélites. O método de PCR marcado por fluorescência demonstrou ser muito eficiente para a análise "multiplex" e proporcionou a redução de custos e ganho de tempo, aumentando a qualidade de análise da genotipagem. Foram apresentados 50 novos locos de microssatélites, desenvolvidos a partir de biblioteca enriquecida a partir da cultivar IAC-UNA. Este estudo fornece ferramentas melhores para assistir aos programas de melhoramento do feijoeiro.Termos para indexação: Phaseolus vulgaris, fluorescência, marcador molecular, análise multiplex.Microsatellite genotyping is widely used nowadays for different goals, such as diversity studies and genetic mapping. Microsatellites (Simple Sequence RepeatsSSRs) are molecular markers characterized as small DNA sequences of one to six base pairs tandemly repeated, spread all over the genome of plants and animals (Li et al., 2002;Varshney et al., 2005). They are multiallelic, codominant and have Mendelian inheritance. Moreover, they are easy to assay and are reproducible by polymerase chain reaction (PCR). Currently, the most used technique for SSR genotyping is the 6% polyacrylamide gel electrophoresis stained with silver nitrate (Creste et al., 2001). However, this method is time consuming and costly, especially when working with a large amount of individuals.Due to this fact, microsatellite fluorescencebased detection has been used in many crops in order to reduce general labor and costs. Oetting et al. (1995) were the first group to use the fluorescent method for genotyping purposes and used five primer pairs to perform different multiplex amplification reactions. The technique was based on a fluorescent-labeled universal M13 primer, which allowed automated DNA sequencer analysis and subsequent genotyping w...

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Linkage Analysis with Multiplexed Short Tandem Repeat Polymorphisms Using Infrared Fluorescence and M13 Tailed Primers

Cited by 337 publications

References 0 publications

Genetic diversity in the U.S. hard red winter wheat cultivars as revealed by microsatellite markers

Genetic diversity in the U.S. hard red winter wheat cultivars as revealed by microsatellite markers

Genetic mapping and annotation of genomic microsatellites isolated from globe artichoke

Adaptation of fluorescent technique for genotyping with new microsatellite markers in common bean

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