plant genome
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Rescooped by hongmin_li from Publications from The Sainsbury Laboratory
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Nature Biotech: Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease (2013)

Nature Biotech: Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease (2013) | plant genome | Scoop.it

http://www.nature.com/nbt/journal/v31/n8/full/nbt.2655.html

 

Sustainable intensification of crop production is essential to ensure food demand is matched by supply as the human population continues to increase. This will require high-yielding crop varieties that can be grown sustainably with fewer inputs on less land. Both plant breeding and genetic modification (GM) methods make valuable contributions to varietal improvement, but targeted genome engineering promises to be critical to elevating future yields. Most such methods require targeting DNA breaks to defined locations followed by either nonhomologous end joining (NHEJ) or homologous recombination. Zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) can be engineered to create such breaks, but these systems require two different DNA binding proteins flanking a sequence of interest, each with a C-terminal FokI nuclease module. We report here that the bacterial clustered, regularly interspaced, short palindromic repeats (CRISPR) system, comprising a CRISPR-associated (Cas)9 protein and an engineered single guide RNA (sgRNA) that specifies a targeted nucleic acid sequence, is applicable to plants to induce mutations at defined loci.


Via Kamoun Lab @ TSL, The Sainsbury Lab
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Elizabeth Jones's curator insight, October 2, 2013 4:25 PM

CRISPR-Cas9 is hot right now, and it's being used in several different ways: for genome-editing in cells of various species (here they're showing it works in plants),  or to create heritable changes (i.e. new transgenic animal lines), and also as a transcription factor to regulate gene expression without altering the nuclear DNA. 

 

Constructs that use Cas9 and synthetic guide RNA (sgRNA) can be customized to target any gene of interest, and can produce insertions, deletions, or incorportation of variant sequences into genomic DNA. Customized constructs can be ordered from gene synthesis suppliers such as GenScript, and delivered much more quickly and cheaply than they can be produced in a typical lab using traditional molecular cloning techniques. 

Rescooped by hongmin_li from Publications from The Sainsbury Laboratory
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Resistance gene enrichment sequencing (RenSeq) enables re-annotation of the NB-LRR gene family from sequenced plant genomes and rapid mapping of resistance loci in segregating populations

Resistance gene enrichment sequencing (RenSeq) enables re-annotation of the NB-LRR gene family from sequenced plant genomes and rapid mapping of resistance loci in segregating populations | plant genome | Scoop.it

Summary

RenSeq is a NB-LRR gene-targeted, Resistance gene enrichment and sequencing method that enables discovery and annotation of pathogen resistance gene family members in plant genome sequences. We successfully applied RenSeq to the sequenced potatoSolanum tuberosum clone DM, and increased the number of identified NB-LRRs from 438 to 755. The majority of these identified R gene loci reside in poorly- or previous un-annotated regions of the genome. Sequence and positional details on the twelve chromosomes have been established for 704 NB-LRRs and can be accessed through a genome browser that we provide. We compared these NB-LRR genes and the corresponding oligonucleotide baits with the highest sequence similarity and demonstrated that ~80% sequence identity is sufficient for enrichment. Analysis of the sequenced tomato S. lycopersicum extended the NB-LRR complement to 394 loci. We further describe a methodology that applies RenSeq to rapidly identify molecular markers that co-segregate with a trait of interest. In two independent segregating populations involving the wild Solanum species S. berthaultii (Rpi-ber2) and S. ruiz-ceballosii (Rpi-rzc1), we were able to apply RenSeq to successfully identify markers that co-segregate with resistance towards the late blight pathogenPhytophthora infestans. These SNP identification workflows were designed as easy-to-adapt Galaxy pipelines.


Via The Sainsbury Lab
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