TAILIEUCHUNG - Targeted genome modifications in soybean with CRISPR/Cas9

The ability to selectively alter genomic DNA sequences in vivo is a powerful tool for basic and applied research. The CRISPR/Cas9 system precisely mutates DNA sequences in a number of organisms. Here, the CRISPR/ Cas9 system is shown to be effective in soybean by knocking-out a green fluorescent protein (GFP) transgene and modifying nine endogenous loci. | Jacobs et al. BMC Biotechnology 2015 15 16 DOI S12896-015-0131-2 BMC Biotechnology RESEARCH ARTICLE Open Access Targeted genome modifications in soybean with CRISPR Cas9 Thomas B Jacobs1 3 5 Peter R LaFayette2 3 Robert J Schmitz4 and Wayne A Parrott1 2 3 Abstract Background The ability to selectively alter genomic DNA sequences in vivo is a powerful tool for basic and applied research. The CRISPR Cas9 system precisely mutates DNA sequences in a number of organisms. Here the CRISPR Cas9 system is shown to be effective in soybean by knocking-out a green fluorescent protein GFP transgene and modifying nine endogenous loci. Results Targeted DNA mutations were detected in 95 of 88 hairy-root transgenic events analyzed. Bi-allelic mutations were detected in events transformed with eight of the nine targeting vectors. Small deletions were the most common type of mutation produced although SNPs and short insertions were also observed. Homoeologous genes were successfully targeted singly and together demonstrating that CRISPR Cas9 can both selectively and generally target members of gene families. Somatic embryo cultures were also modified to enable the production of plants with heritable mutations with the frequency of DNA modifications increasing with culture time. A novel cloning strategy and vector system based on In-Fusion cloning was developed to simplify the production of CRISPR Cas9 targeting vectors which should be applicable for targeting any gene in any organism. Conclusions The CRISPR Cas9 is a simple efficient and highly specific genome editing tool in soybean. Although some vectors are more efficient than others it is possible to edit duplicated genes relatively easily. The vectors and methods developed here will be useful for the application of CRISPR Cas9 to soybean and other plant species. Keywords CRISPR Cas9 Plant transformation Soybean Genomic engineering Gene targeting Hairy roots Background Methods to specifically target and modify DNA .

• Sinh học
• BMC Biotechnology
• Plant transformation
• Genomic engineering
• Gene targeting
• Hairy roots
• Transgenic animals
• Recombinant proteins
• Animal biotechnology
• Functional cytokines
• ABC transporter
• Taxonomic diversity
• ATP binding cassette
• Transporter gene superfamily
• Community engagement
• Agricultural biotechnology
• Tissue culture
• Global health
• Global development
• Stakeholder engagement
• BMC Chemistry
• Citric acid
• White biotechnology
• Organic acid
• Emerging bioeconomy
• MicroRNAs
• cDNA synthesis
• synthetic templates
• Design of PCR primers
• Quantitative PCR
• Error prone PCR
• Site directed mutation
• Site saturation mutagenesis
• Catalytic efficiency
• Tobacco stalk
• Nicotine degradation
• Phanerochaete chrysosporium
• Lignocellulolytic enzymes
• Chaetomium globosum
• Synergistic action
• Lignocellulosic materials
• Phenolic alcohols
• β xylosidase
• Highly thermostable GH39
• Geobacillus sp
• Hydrolyzing xylo oligosaccharides
• Heterologous expression
• Kluyveromyces lactis
• Baking industry
• Plectosphaerella cucumerina
• Gelatin scaffold
• Human articular chondrocytes
• Extracellular matrix
• Cartilage tissue engineering
• Protein design
• Protein expression
• Protein interaction
• Protein engineering
• Pseudomonas putida strains
• Raw glycerol
• PHA depolymerase
• Metabolic engineering
• Protein cross linking
• Protein stability
• Baculovirus expression vector system
• Human influenza strain
• Arteriovenous loop
• PHD inhibitor
• Dimethyloxallyl Glycine
• HIF α level
• Substitution pattern
• Hydroxypropyl methylcellulose
• Application of carbohydrate binding modules
• Hydroxyethyl cellulose
• Rhizomucor pusillus
• α amylase
• Co expression
• Pichia pastoris
• Alkaline phosphatase
• Escherichia coli
• Psychrophilic alkaline phosphatase
• Hydrolytic cleavage
• Leuconostoc citreum
• Inulosucrase IslA
• Multidomain fructosyltransferase
• Multidomain enzyme
• Hydrolytic activity
• CD spectroscopy
• Homology modelling
• Peroxidase activity
• Superoxide dismutase
• Trichoderma reesei
• Endoglucanase I
• Cell free synthesis
• Saccharomyces cerevisiae
• Neurospora crassa
• Glutaminyl cylcase
• Tobacco mosaic virus
• Protein purification
• N benthamiana
• Active pharmaceutical ingredient
• Adenoviral vector
• Inducible vector
• Gene therapy
• Optimizing therapeutic outcomes
• Co transformation
• Gaussia princeps luciferase gene
• Genetic engineering
• Green algae
• Inducible promoter
• Nitrate reductase
• Spider silk
• Synthetic spidroin
• Atomic force microscopy
• Outstanding mechanical