Abstract: Methods and constructs for the multiplex expression of highly active CRISPR guide RNAs (gRNAs) from RNA Polymerase II and III promoters, optionally in mammalian cells. The present invention is based, at least in part, on the discovery that Csy4, an endoribonuclease that recognizes a short RNA hairpin sequence, can be used to cleave out multiple functional gRNAs encoded on a single longer RNA transcript (produced from an RNA pol II or III promoter) in which the individual gRNAs are separated by Csy4 cleavage sites.

Abstract: Engineered CRISPR-Cas9 nucleases with altered and improved PAM specificities and their use in genomic engineering, epigenomic engineering, and genome targeting.

Abstract: Engineered CRISPR-Cas9 nucleases with altered and improved PAM specificities and their use in genomic engineering, epigenomic engineering, and genome targeting.

Abstract: Unbiased, genomewide and highly sensitive methods for detecting mutations, e.g., off-target mutations, induced by engineered nucleases.

Abstract: Many studies have shown that CRISPR-Cas nucleases can tolerate up to five mismatches and still cleave; it is hard to predict the effects of any given single or combination of mismatches on activity. Taken together, these nucleases can show significant off-target effects but it can be challenging to predict these sites. Described herein are methods for increasing the specificity of genome editing using the CRISPR/Cas system, e.g., using RNA-guided Foki Nucleases (RFNs), e.g., Fokl-Cas9 or Foki-dCas9-based fusion proteins.

Abstract: CRISPR-Cas genome editing uses a guide RNA, which includes both a complementarity region, which binds the target DNA by base-pairing, and a Cas9-binding region, to direct a Cas9 nuclease to a target DNA. Further disclosed are methods for increasing specificity of RNA-guided genome editing using CRISPR/Cas9 systems by using truncated guide RNAs (tru-gRNAs).

Abstract: Methods for increasing specificity of RNA-guided genome editing, e.g., editing using CRISPR/Cas9 systems.

Abstract: Methods and constructs for RNA-guided targeting of heterologous functional domains such as transcriptional activators to specific genomic loci.

Abstract: Computer programs, algorithms, and methods for identifying TALE-activator binding sites, and methods for generation and use of TALE-activators that bind to said sites.

Abstract: Methods and constructs for RNA-guided targeting of transcriptional activators to specific genomic loci.

Abstract: Methods for increasing specificity of RNA-guided genome editing, e.g., editing using CRISPR/Cas9 systems, using truncated guide RNAs (tru-gRNAs).

Abstract: Methods for increasing specificity of RNA-guided genome editing, e.g., editing using CRISPR/Cas9 systems.

Abstract: The disclosure describes methods that include providing a first nucleic acid having a sequence encoding a first set comprising one or more transcription activator-like effector (TALE) repeat domains and/or one or more portions of one or more TALE repeat domains; contacting the first nucleic acid with a first enzyme, wherein the first enzyme creates a first ligatable end; providing a second nucleic acid having a sequence encoding a second set comprising one or more TALE repeat domains and/or one or more portions of one or more TALE repeat domains; contacting the second nucleic acid with a second enzyme, wherein the second enzyme creates a second ligatable end, and wherein the first and second ligatable ends are compatible; and ligating the first and second nucleic acids through the first and second ligatable ends to produce a first ligated nucleic acid, wherein the first ligated nucleic acid is linked to a solid support, and wherein the first ligated nucleic acid encodes a polypeptide comprising said first and

Abstract: The invention relates to organisms and compositions comprising one or more stem cells or one or more embryos, wherein the one or more stem cells or one or more embryos comprise one or more of the following mutations: (i) a deletion mutation; (ii) a knockout mutation; and/or (iii) an addition of a heterologous nucleic acid sequence; wherein the one or more mutations of (i), (ii), and/or (iii) are site-specific mutations caused by a Xanthomonas TAL nuclease (XTN). The invention also relates to method of mutating an embryo, iPS cell, stem cell, or more particularly a spermatogonial stem cell by exposing the nucleic acid sequence contained within such embryos or cell with a Xanthomonas TAL nuclease.

Abstract: Fusion proteins comprising a DNA binding domain, e.g., a TAL effector repeat array or zinc finger, and a catalytic domain comprising a sequence that catalyzes hydroxylation of methylated cytosines in DNA, and methods of use thereof.

Abstract: A method of designing a multi-zinc-finger polypeptide predicted to bind to a sequence of interest that has at least three subsites includes the steps of: a) providing a nucleotide sequence of interest having first, second, and third consecutive subsites, wherein each of the first and third subsites are adjacent to the second subsite; b) identifying first and second adjacent zinc finger polypeptide sequences previously shown to bind to the first and second subsites in the context of a multi-zinc finger polypeptide; c) identifying a third zinc finger polypeptide previously shown to bind to a third subsite adjacent to the second subsite when present in the context of a multi-zinc finger polypeptide adjacent to the second zinc finger polypeptide; and d) combining the first, second, and third zinc finger polypeptide sequences in linear order, thereby designing a multi-zinc finger polypeptide predicted to bind to the sequence of interest.

Abstract: A combined yeast/bacterial two-hybrid system is disclosed.

Abstract: The present invention provides methods and compositions for interaction trap assays for detecting protein-protein, protein-DNA, or protein-RNA interactions. The methods and compositions of the invention may also be used to identify agents which may agonize or antagonize a protein-protein, protein-DNA, or protein-RNA interaction. In certain embodiments, the interaction trap system of the invention is useful for screening libraries with greater than 107 members. In other embodiments, the interaction trap system of the invention is used in conjunction with flow cytometry. The invention further provides a means for simultaneously screening a target protein or nucleic acid sequence for the ability to interact with two or more test proteins or nucleic acids.

Abstract: A combined yeast/bacterial two-hybrid system is disclosed.

Abstract: The present invention provides methods and compositions for interaction trap assays for detecting protein-protein, protein-DNA, or protein-RNA interactions. The methods and compositions of the invention may also be used to identify agents which may agonize or antagonize a protein-protein, protein-DNA, or protein-RNA interactions. In certain embodiments, the interaction trap system of the invention is useful for screening libraries with greater than 107 members. In other embodiments, the interaction trap system of the invention is used in conjunction with flow cytometry. The invention further provides a means for simultaneously screening a target protein or nucleic acid sequence for the ability to interact with two or more test proteins or nucleic acids.