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      High-Resolution Genotyping via Whole Genome Hybridizations to Microarrays Containing Long Oligonucleotide Probes

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          To date, microarray-based genotyping of large, complex plant genomes has been complicated by the need to perform genome complexity reduction to obtain sufficiently strong hybridization signals. Genome complexity reduction techniques are, however, tedious and can introduce unwanted variables into genotyping assays. Here, we report a microarray-based genotyping technology for complex genomes (such as the 2.3 GB maize genome) that does not require genome complexity reduction prior to hybridization. Approximately 200,000 long oligonucleotide probes were identified as being polymorphic between the inbred parents of a mapping population and used to genotype two recombinant inbred lines. While multiple hybridization replicates provided ∼97% accuracy, even a single replicate provided ∼95% accuracy. Genotyping accuracy was further increased to >99% by utilizing information from adjacent probes. This microarray-based method provides a simple, high-density genotyping approach for large, complex genomes.

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          Most cited references 28

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          Statistical significance for genomewide studies.

          With the increase in genomewide experiments and the sequencing of multiple genomes, the analysis of large data sets has become commonplace in biology. It is often the case that thousands of features in a genomewide data set are tested against some null hypothesis, where a number of features are expected to be significant. Here we propose an approach to measuring statistical significance in these genomewide studies based on the concept of the false discovery rate. This approach offers a sensible balance between the number of true and false positives that is automatically calibrated and easily interpreted. In doing so, a measure of statistical significance called the q value is associated with each tested feature. The q value is similar to the well known p value, except it is a measure of significance in terms of the false discovery rate rather than the false positive rate. Our approach avoids a flood of false positive results, while offering a more liberal criterion than what has been used in genome scans for linkage.
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            The B73 maize genome: complexity, diversity, and dynamics.

            We report an improved draft nucleotide sequence of the 2.3-gigabase genome of maize, an important crop plant and model for biological research. Over 32,000 genes were predicted, of which 99.8% were placed on reference chromosomes. Nearly 85% of the genome is composed of hundreds of families of transposable elements, dispersed nonuniformly across the genome. These were responsible for the capture and amplification of numerous gene fragments and affect the composition, sizes, and positions of centromeres. We also report on the correlation of methylation-poor regions with Mu transposon insertions and recombination, and copy number variants with insertions and/or deletions, as well as how uneven gene losses between duplicated regions were involved in returning an ancient allotetraploid to a genetically diploid state. These analyses inform and set the stage for further investigations to improve our understanding of the domestication and agricultural improvements of maize.
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              Rapid SNP Discovery and Genetic Mapping Using Sequenced RAD Markers

              Single nucleotide polymorphism (SNP) discovery and genotyping are essential to genetic mapping. There remains a need for a simple, inexpensive platform that allows high-density SNP discovery and genotyping in large populations. Here we describe the sequencing of restriction-site associated DNA (RAD) tags, which identified more than 13,000 SNPs, and mapped three traits in two model organisms, using less than half the capacity of one Illumina sequencing run. We demonstrated that different marker densities can be attained by choice of restriction enzyme. Furthermore, we developed a barcoding system for sample multiplexing and fine mapped the genetic basis of lateral plate armor loss in threespine stickleback by identifying recombinant breakpoints in F2 individuals. Barcoding also facilitated mapping of a second trait, a reduction of pelvic structure, by in silico re-sorting of individuals. To further demonstrate the ease of the RAD sequencing approach we identified polymorphic markers and mapped an induced mutation in Neurospora crassa. Sequencing of RAD markers is an integrated platform for SNP discovery and genotyping. This approach should be widely applicable to genetic mapping in a variety of organisms.

                Author and article information

                Role: Editor
                PLoS One
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                2 December 2010
                : 5
                : 12
                [1 ]Department of Agronomy, Iowa State University, Ames, Iowa, United States of America
                [2 ]Department of Plant Biology, University of Minnesota, St Paul, Minnesota, United States of America
                [3 ]Interdepartmental Genetics Graduate Program, Iowa State University, Ames, Iowa, United States of America
                [4 ]Department of Genetics, Development and Cell Biology, Iowa State University, Ames, Iowa, United States of America
                [5 ]Center for Plant Genomics, Iowa State University, Ames, Iowa, United States of America
                [6 ]Roche NimbleGen, Inc., Madison, Wisconsin, United States of America
                [7 ]Department of Biology and the Genetics Institute, University of Florida, Gainesville, Florida, United States of America
                [8 ]Department of Statistics, Iowa State University, Ames, Iowa, United States of America
                United States Department of Agriculture, Agricultural Research Service, United States of America
                Author notes

                Conceived and designed the experiments: YF NMS JAJ PSS. Performed the experiments: NMS ALI TR WW. Analyzed the data: YF NMS KY CTY WBB DN PSS. Contributed reagents/materials/analysis tools: JAJ PSS. Wrote the paper: YF NMS JAJ PSS.


                Current address: Monsanto Company, St. Louis, Missouri, United States of America,

                Fu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                Page count
                Pages: 8
                Research Article
                Genetics and Genomics
                Genetics and Genomics/Comparative Genomics
                Genetics and Genomics/Plant Genetics and Gene Expression



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