High-throughput tetrad analysis

Ludlow, Catherine L.; Scott, Adrian C.; Cromie, Gareth A.; Jeffery, Eric; Sirr, Amy; Lin, Jake; Gilbert, Teresa; Hays, Michelle; Dudley, Andrew T.

doi:10.1038/nmeth.2479

Cited by 23 publications

(31 citation statements)

References 27 publications

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Section: Prospects Of Mapping Complex Traits In Model Organismsmentioning

confidence: 99%

“…SGRP-4X is useful in a variety of settings, as we have shown for the study of weaker alleles and of multiple contributors at a locus and following up allelic series and interactions between loci. With rapid advances in highly multiplexed whole-genome sequencing, an even larger number of new individuals derived from the SGRP4-X could be genotyped in the future to approach nearly full power to map trait loci (Bloom et al 2013;Ludlow et al 2013;Wilkening et al 2013).We have harnessed the awesome power of baker's yeast for quantitative genetics studies. Full-genome sequences containing a large fraction of natural variation in the species, high-resolution phenotypes, an extremely large number of individuals, and low linkage disequilibrium have produced a very powerful mapping population.…”

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confidence: 99%

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High-Resolution Mapping of Complex Traits with a Four-Parent Advanced Intercross Yeast Population

et al. 2013

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A large fraction of human complex trait heritability is due to a high number of variants with small marginal effects and their interactions with genotype and environment. Such alleles are more easily studied in model organisms, where environment, genetic makeup, and allele frequencies can be controlled. Here, we examine the effect of natural genetic variation on heritable traits in a very large pool of baker's yeast from a multiparent 12th generation intercross. We selected four representative founder strains to produce the Saccharomyces Genome Resequencing Project (SGRP)-4X mapping population and sequenced 192 segregants to generate an accurate genetic map. Using these individuals, we mapped 25 loci linked to growth traits under heat stress, arsenite, and paraquat, the majority of which were best explained by a diverging phenotype caused by a single allele in one condition. By sequencing pooled DNA from millions of segregants grown under heat stress, we further identified 34 and 39 regions selected in haploid and diploid pools, respectively, with most of the selection against a single allele. While the most parsimonious model for the majority of loci mapped using either approach was the effect of an allele private to one founder, we could validate examples of pleiotropic effects and complex allelic series at a locus. SGRP-4X is a deeply characterized resource that provides a framework for powerful and high-resolution genetic analysis of yeast phenotypes and serves as a test bed for testing avenues to attack human complex traits.T HE strong tendency for progeny to closely resemble their parents has turned out to be difficult to understand in detail. Nearly all traits, including lifetime risk for many common diseases, have a complex genetic basis that is determined by multiple quantitative trait loci (QTL) (Donnelly 2008;Manolio et al. 2009). The first step toward accurate models of trait variability, and a prerequisite for predicting and modulating them, is characterization of the underlying genetic factors in the context of rest of the genome and their external environment. Research in model systems has led the way in this effort and produced powerful experimental and computational approaches for genetic mapping (Nordborg and Weigel 2008;Flint and Mackay 2009;Mackay et al. 2009).A traditional, well-controlled approach for finding the QTL underlying natural phenotypic variation is to analyze a large number of progenies from two-parent crosses (Brem et al. 2002;Simon et al. 2008). Studies using this design have improved our understanding of complex traits and provided concrete evidence of natural segregating variants ), but have been limited in their scope with regard to the extent of genetic variation between the two parents. Mapping populations of popular model organisms ranging from fruit flies (King et al. 2012) (Churchill et al. 2004;Durrant et al. 2011) to plants (Kover et al. 2009;Gan et al. 2011;Huang et al. 2011) has recently expanded the genetic and phenotypic diversity available to study by ...

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Section: Prospects Of Mapping Complex Traits In Model Organismsmentioning

confidence: 99%

mentioning

confidence: 99%

High-Resolution Mapping of Complex Traits with a Four-Parent Advanced Intercross Yeast Population

et al. 2013

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“…Full-tetrad analysis in plants is contingent on the availability of mutants where tetrads fail to disassociate during pollen development (Copenhaver et al 2000). Pollen tetrads have been reported in hundreds of species to date (Copenhaver 2005); however, manual isolation of individual tetrads can be challenging (Copenhaver et al 2000;Ludlow et al 2013). Half-tetrad analysis may be more readily applicable: unreduced (2n) gametes have been reported in a wide range of plant and animal species (Ramsey and Schemske 1998).…”

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confidence: 99%

Centromere Locations inBrassicaA and C Genomes Revealed Through Half-Tetrad Analysis

et al. 2015

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Locating centromeres on genome sequences can be challenging. The high density of repetitive elements in these regions makes sequence assembly problematic, especially when using short-read sequencing technologies. It can also be difficult to distinguish between active and recently extinct centromeres through sequence analysis. An effective solution is to identify genetically active centromeres (functional in meiosis) by half-tetrad analysis. This genetic approach involves detecting heterozygosity along chromosomes in segregating populations derived from gametes (half-tetrads). Unreduced gametes produced by first division restitution mechanisms comprise complete sets of nonsister chromatids. Along these chromatids, heterozygosity is maximal at the centromeres, and homologous recombination events result in homozygosity toward the telomeres. We genotyped populations of half-tetrad-derived individuals (from Brassica interspecific hybrids) using a high-density array of physically anchored SNP markers (Illumina Brassica 60K Infinium array). Mapping the distribution of heterozygosity in these half-tetrad individuals allowed the genetic mapping of all 19 centromeres of the Brassica A and C genomes to the reference Brassica napus genome. Gene and transposable element density across the B. napus genome were also assessed and corresponded well to previously reported genetic map positions. Known centromerespecific sequences were located in the reference genome, but mostly matched unanchored sequences, suggesting that the core centromeric regions may not yet be assembled into the pseudochromosomes of the reference genome. The increasing availability of genetic markers physically anchored to reference genomes greatly simplifies the genetic and physical mapping of centromeres using half-tetrad analysis. We discuss possible applications of this approach, including in species where half-tetrads are currently difficult to isolate.

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“…Small‐scale cell disruption of yeasts has often been achieved using bead lysis methods . However, limitations of bead‐mediated lysis include the requirement of additional process bottlenecks demanding the removal of beads from lysate, poor temperature control, lower process reproducibility due to a requirement of bead size uniformity, loss of product due to residual volume after sample processing and high surface‐to‐volume ratios which could lead to adhesion of cellular material …”

Section: Introductionmentioning

confidence: 99%

“…22 However, limitations of bead-mediated lysis include the requirement of additional process bottlenecks demanding the removal of beads from lysate, poor temperature control, lower process reproducibility due to a requirement of bead size uniformity, loss of product due to residual volume after sample processing and high surface-to-volume ratios which could lead to adhesion of cellular material. 23 Adaptive focused acoustics (AFA) has shown to be a suitable miniaturized platform for the disruption of the yeast, Saccharomyces cerevisiae. 24 An AFA device generates acoustic shock waves in the kilohertz (KHz) region.…”

Section: Introductionmentioning

confidence: 99%

Development of a high‐throughput microscale cell disruption platform for Pichia pastoris in rapid bioprocess design

Blaha

Morris

Ogonah

et al. 2017

Biotechnology Progress

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The time and cost benefits of miniaturized fermentation platforms can only be gained by employing complementary techniques facilitating high-throughput at small sample volumes. Microbial cell disruption is a major bottleneck in experimental throughput and is often restricted to large processing volumes. Moreover, for rigid yeast species, such as Pichia pastoris, no effective high-throughput disruption methods exist. The development of an automated, miniaturized, high-throughput, noncontact, scalable platform based on adaptive focused acoustics (AFA) to disrupt P. pastoris and recover intracellular heterologous protein is described. Augmented modes of AFA were established by investigating vessel designs and a novel enzymatic pretreatment step. Three different modes of AFA were studied and compared to the performance high-pressure homogenization. For each of these modes of cell disruption, response models were developed to account for five different performance criteria. Using multiple responses not only demonstrated that different operating parameters are required for different response optima, with highest product purity requiring suboptimal values for other criteria, but also allowed for AFA-based methods to mimic large-scale homogenization processes. These results demonstrate that AFA-mediated cell disruption can be used for a wide range of applications including buffer development, strain selection, fermentation process development, and whole bioprocess integration. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 34:130-140, 2018.

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High-throughput tetrad analysis

Cited by 23 publications

References 27 publications

High-Resolution Mapping of Complex Traits with a Four-Parent Advanced Intercross Yeast Population

High-Resolution Mapping of Complex Traits with a Four-Parent Advanced Intercross Yeast Population

Centromere Locations inBrassicaA and C Genomes Revealed Through Half-Tetrad Analysis

Development of a high‐throughput microscale cell disruption platform for Pichia pastoris in rapid bioprocess design

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