Targeted Capture and Sequencing of Gene-Sized DNA Molecules

Giolai, Michael; Paajanen, Pirita; Verweij, Walter; Percival‐Alwyn, Lawrence; Baker, David; Witek, Kamil; Jupe, Florian; Bryan, Glenn; Hein, Ingo; Jones, Jonathan D. G.; Clark, Matt

doi:10.2144/000114484

Cited by 35 publications

(30 citation statements)

References 21 publications

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“…The methodology takes advantage of the focused re-sequencing of NLRs through RenSeq (Jupe et al, 2013). RenSeq has previously been used for improving genome annotations and genetic mapping of plant NLRs (Chen et al, 2018;Jupe et al, 2013), the prioritisation of novel NLRs in wild diploid species (Jiang et al, 2018;Van Weymers et al, 2016), and identification of candidate NLRs when combined with long-read sequencing technology (Giolai et al, 2016;Witek et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

Tracking disease resistance deployment in potato breeding by enrichment sequencing

Armstrong

Vossen

Lim

et al. 2018

Plant Biotechnology Journal

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Summary Following the molecular characterisation of functional disease resistance genes in recent years, methods to track and verify the integrity of multiple genes in varieties are needed for crop improvement through resistance stacking. Diagnostic resistance gene enrichment sequencing ( dR enSeq) enables the high‐confidence identification and complete sequence validation of known functional resistance genes in crops. As demonstrated for tetraploid potato varieties, the methodology is more robust and cost‐effective in monitoring resistances than whole‐genome sequencing and can be used to appraise (trans) gene integrity efficiently. All currently known NB ‐ LRR s effective against viruses, nematodes and the late blight pathogen Phytophthora infestans can be tracked with dR enSeq in potato and hitherto unknown polymorphisms have been identified. The methodology provides a means to improve the speed and efficiency of future disease resistance breeding in crops by directing parental and progeny selection towards effective combinations of resistance genes.

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Section: Introductionmentioning

confidence: 99%

Tracking disease resistance deployment in potato breeding by enrichment sequencing

Armstrong

Vossen

Lim

et al. 2018

Plant Biotechnology Journal

Self Cite

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“…Such approaches, for example, have been successfully applied to investigating R-gene variation in crop plants (Jupe et al, 2012(Jupe et al, , 2013Andolfo et al, 2014;Giolai et al, 2016;Russell et al, 2016;Van Weymers et al, 2016). Whole genome resequencing approaches could be useful for setting the genomic context and fate of duplications, but there are still substantial challenges to resolve in distinguishing loss of copies from lack of coverage or lack of assembly to the reference due to high sequence divergence.…”

Section: Conclusion and Recommendationsmentioning

confidence: 99%

“…For example, the "big bang" theory of the emergence of the adaptive immune systems in vertebrates invokes multiple rounds of WGD as the major source of this potential (Flajnik and Kasahara, 2010). Similarly, investigation of the genomic repertoire of pathogen-associated genes (R genes) in several crop plants through targeted sequence capture (Jupe et al, 2012(Jupe et al, , 2013Giolai et al, 2016;Van Weymers et al, 2016) has revealed much more extensive gene families than was previously predicted based on whole genome resequencing studies. R genes have also been demonstrated to show signatures of adaptive introgression between closely related species of Arabidopsis, with extensive trans-specific sharing of alleles across species (Bechsgaard et al, 2017).…”

Section: Introduction Background and Aimsmentioning

confidence: 99%

Adding Complexity to Complexity: Gene Family Evolution in Polyploids

et al. 2018

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Comparative genomics of non-model organisms has resurrected whole genome duplication (WGD) from being viewed as a somewhat obscure process that happens in plants to a primary driver of eukaryotic diversification. The shadow of past ploidy increases has left a strong signature of duplicated genes organized into gene families, even in small genomes that have undergone effectively complete rediploidization. Nevertheless, despite continually advancing technologies and bioinformatics pipelines, resolving the fate of duplicate genes remains a substantial challenge. For example, many important recognition processes are driven not only by allelic expansion through retention of duplicates but also by diversification and copy number variation. This creates technical difficulties with assembly to reference genomes and accurate interpretation of homology. Thus, relatively little is known about the impacts of recent polyploidization and hybridization on the evolution of gene families under selective forces that maintain diversity, such as balancing selection. Here we use a complex of species and ploidy levels in the genus Arabidopsis (A. lyrata and A. arenosa) as a model to investigate the evolutionary dynamics of a large and complicated gene family known to be under strong balancing selection: the receptor-like kinases, which include the female component of genetically controlled self-incompatibility. Specifically, we question: (1) How does diversity of S-receptor kinase (SRK) alleles in tetraploids compare to that in their close diploid relatives? (2) Is there increased trans-specific polymorphism (i.e., sharing of alleles that transcend speciation, characteristic of balancing selection) in tetraploids compared to diploids due to the higher number of copies they carry? (3) Do these highly variable loci show evidence of introgression among extant species/ploidy levels within or outside known zones of hybridization? (4) Is there evidence for copy number variation among paralogs? We use this example to highlight specific issues to consider when interpreting gene family evolution, particularly in relation to polyploids but also more generally in diploids. We conclude with recommendations for strategies to address the challenges of resolving such complex loci in the future, using advances in deep sequencing approaches.

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“…These approaches were initially developed for targeted sequencing of protein-coding regions of genomic DNA (whole exome sequencing) 13 and RNA fragments from short-read RNA-seq experiments (e.g., CaptureSeq) [14][15][16][17][18] . Such approaches have been adapted for targeted sequencing of long genomic fragments (>2kb) or full-length cDNA molecules [19][20][21][22][23][24] . In two notable studies described recently, complex pools of biotinylated oligos were used to enrich for and sequence thousands of protein-coding and lncRNA targets, leading to considerable gains in isoform detection and new insights about the nature of transcriptomic complexity 25,26 .…”

Section: Introductionmentioning

confidence: 99%

ORF Capture-Seq: a versatile method for targeted identification of full-length isoforms

Sheynkman

Tuttle

Tseng

et al. 2019

Preprint

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AbstractMost human protein-coding genes are expressed as multiple isoforms. This in turn greatly expands the functional repertoire of the encoded proteome. While at least one reliable open reading frame (ORF) model has been assigned for every gene, the majority of alternative isoforms remains uncharacterized experimentally. This is primarily due to: i) vast differences of overall levels between different isoforms expressed from common genes, and ii) the difficulty of obtaining contiguous full-length ORF sequences. Here, we present ORF Capture-Seq (OCS), a flexible and cost-effective method that addresses both challenges for targeted full-length isoform sequencing applications using collections of cloned ORFs as probes. As proof-of-concept, we show that an OCS pipeline focused on genes coding for transcription factors increases isoform detection by an order of magnitude, compared to unenriched sample. In short, OCS enables rapid discovery of isoforms from custom-selected genes and will allow mapping of the full set of human isoforms at reasonable cost.

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Targeted Capture and Sequencing of Gene-Sized DNA Molecules

Cited by 35 publications

References 21 publications

Tracking disease resistance deployment in potato breeding by enrichment sequencing

Tracking disease resistance deployment in potato breeding by enrichment sequencing

Adding Complexity to Complexity: Gene Family Evolution in Polyploids

ORF Capture-Seq: a versatile method for targeted identification of full-length isoforms

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