ArticleBMC genomics2008
Coupling amplified DNA from flow-sorted chromosomes to high-density SNP mapping in barley.
Article in BMC genomics, 2008. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 70 papers.
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70 citing papers in PubMed.
- A single NLR gene confers resistance to leaf and stripe rust in wheat.Nature communications · 2024Article
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- Chromosome genomics facilitates the marker development and selection of wheat-Aegilops biuncialis addition, substitution and translocation lines.Scientific reports · 2023Article
- The chromatin determinants and Ph1 gene effect at wheat sites with contrasting recombination frequency.Journal of advanced research · 2023Article
- The wheat stem rust resistance gene Sr43 encodes an unusual protein kinase.Nature genetics · 2023Article
- Flow Cytometric Analysis and Sorting of Plant Chromosomes.Methods in molecular biology (Clifton, N.J.) · 2023Article
- Isolation and Sequencing of Chromosome Arm 7RS of Rye,International journal of molecular sciences · 2022Article
- Aegilops sharonensis genome-assisted identification of stem rust resistance gene Sr62.Nature communications · 2022Article
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- Draft Sequencing Crested Wheatgrass Chromosomes Identified Evolutionary Structural Changes and Genes and Facilitated the Development of SSR Markers.International journal of molecular sciences · 2022Article
- Flow karyotyping of wheat-Frontiers in plant science · 2022Article
- Identification, High-Density Mapping, and Characterization of New Major Powdery Mildew Resistance Loci From the Emmer Wheat Landrace GZ1.Frontiers in plant science · 2022Article
- Chromosome analysis and sorting.Cytometry. Part A : the journal of the International Society for Analytical Cytology · 2021Review
- A membrane-bound ankyrin repeat protein confers race-specific leaf rust disease resistance in wheat.Nature communications · 2021Article
- Bridging the Gap between Vertebrate Cytogenetics and Genomics with Single-Chromosome Sequencing (ChromSeq).Genes · 2021Review
- Fundamentally different repetitive element composition of sex chromosomes in Rumex acetosa.Annals of botany · 2021Article
- Development of DNA Markers From Physically Mapped Loci inFrontiers in plant science · 2021Article
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- The improved assembly of 7DL chromosome provides insight into the structure and evolution of bread wheat.Plant biotechnology journal · 2020Article
- Comparative analysis of chromosome 2A molecular organization in diploid and hexaploid wheat.Molecular biology reports · 2020Article
10 more citing papers are in PubMed but not listed here.
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10 authors.
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No grant is acknowledged in the PubMed record.
Abstract
backgroundFlow cytometry facilitates sorting of single chromosomes and chromosome arms which can be used for targeted genome analysis. However, the recovery of microgram amounts of DNA needed for some assays requires sorting of millions of chromosomes which is laborious and time consuming. Yet, many genomic applications such as development of genetic maps or physical mapping do not require large DNA fragments. In such cases time-consuming de novo sorting can be minimized by utilizing whole-genome amplification.
resultsHere we report a protocol optimized in barley including amplification of DNA from only ten thousand chromosomes, which can be isolated in less than one hour. Flow-sorted chromosomes were treated with proteinase K and amplified using Phi29 multiple displacement amplification (MDA). Overnight amplification in a 20-microlitre reaction produced 3.7 - 5.7 micrograms DNA with a majority of products between 5 and 30 kb. To determine the purity of sorted fractions and potential amplification bias we used quantitative PCR for specific genes on each chromosome. To extend the analysis to a whole genome level we performed an oligonucleotide pool assay (OPA) for interrogation of 1524 loci, of which 1153 loci had known genetic map positions. Analysis of unamplified genomic DNA of barley cv. Akcent using this OPA resulted in 1426 markers with present calls. Comparison with three replicates of amplified genomic DNA revealed >99% concordance. DNA samples from amplified chromosome 1H and a fraction containing chromosomes 2H - 7H were examined. In addition to loci with known map positions, 349 loci with unknown map positions were included. Based on this analysis 40 new loci were mapped to 1H.
conclusionThe results indicate a significant potential of using this approach for physical mapping. Moreover, the study showed that multiple displacement amplification of flow-sorted chromosomes is highly efficient and representative which considerably expands the potential of chromosome flow sorting in plant genomics.
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