A new technology for genome editing may put the zinc finger nuclease franchise out of business, some believe. Not so fast, say the finger people. Laura DeFrancesco reports.
"Zinc finger nucleases have been used to generate DSBs and subsequently, for genome editing but with low efficiency and reproducibility. ... Here, we report the generation of a Hax3-based hybrid TALE nuclease with a user-selected DNA binding specificity. ... Transient expression assays in tobacco leaves suggest that the hybrid nuclease creates DSB (double strand breaks) in its target sequence, which is subsequently repaired by nonhomologous end-joining repair. ... great potential for site-specific genome modification in plants and eukaryotes in general"
It is this repeat domain that is responsible for specific DNA sequence recognition. Each repeat is almost identical with the exception of two variable amino acids termed the repeat-variable diresidues.
"Additionally, TALE-NT provides tools to design single TAL effectors to bind to a DNA sequence, and to identify binding sites in a DNA sequence for a given TAL effector. The TALE-NT website is maintained by the Bogdanove laboratory at Iowa State University."
Two papers published today, one in Nature Biotechnology and one in Nature Methods, show that although zinc-finger nucleases are highly specific, the methods previously used to predict where they might go wrong miss the mark.
A TAL effector of "Xanthomonas oryzae pv oryzae (Xoo) transcriptionally activates rice (Oryza sativa) susceptibility gene Xa13 to cause bacterial blight disease. Xa13 encodes an indispensable plasma membrane protein"..."We show that the XA13 protein cooperates with two other proteins"..."to promote removal of copper from xylem vessels, where Xoo multiplies and spreads to cause disease"
The TAL Effector Technology license was jointly signed with inventors Jens Boch, Ulla Bonas, Thomas Lahaye and Sebastian Schornack, and the Two Blades Foundation (2Blades), which exclusively licensed plant research tool ...
Another publication describing the use of type IIS restriction enzymes in two sequential cut-ligase reactions to build TAL DNA binding domain repeat arrays
"Recent elucidation of the basis for specificity in DNA binding by TALEs expedites further discovery and opens the door to biotechnological applications. This article reviews the most significant findings in TALE research, with a focus on recent advances, and discusses future prospects including pressing questions yet to be answered"
TAL effector - Description: TAL effectors (TALEs) are proteins secreted by Xanthomonas bacteria via their type III secretion system when they infect various plant species.
Transcription activator-like effector nucleases (TALENs) are a new technology for modifying the genome at specific loci of interest. Hockemeyer et al. now demonstrate the utility of TALENs for gene targeting in human pluripotent stem cells.
"We leverage codon degeneracy and type IIs restriction enzymes to generate orthogonal ligation linkers between individual repeat monomers, thus allowing full-length, customized, repeat domains to be constructed by hierarchical ligation. We" ... "demonstrate that they can specifically modulate transcription of endogenous genes (SOX2 and KLF4) in human cells"
One of two papers describing the original discovery of a linear code for DNA-binding site prediction Image: Adam Bogdanove and Harry Horner/Iowa State University; false color: Yael Kats/Science
A TAL effector is a protein of bacterial origin and is able to bind to DNA sequences. The bound sequence is defined by the structure of the TAL effector protein. Manipulate the effector protein and you can direct to any binding site you want. Powerful biotechnology, I say...
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