Engineering our Genome with TAL Effectors

Transcription activator-like effector (TALE) nucleases (TALENs) have recently emerged as a revolutionary genome engineering tool. TALE proteins are naturally used by a genus of pathogenic plant bacteria belonging to Xanthomonas spp. These proteins function as a virulence tool by binding to specific host DNA sequences and activating gene expression to support infection. DNA recognition by TALEs is mediated through a central repeat domain consisting of a variable number of amino acid repeats. Within each single repeat two adjacent amino acids known as ‘repeat variable di-residue’ (RVD) interact with a specific nucleotide base constituting a simple recognition code. The unique mechanism by which TALEs recognise specific DNA sequences offers the potential to tailor TALEs to recognise and target any DNA sequence.
Genetically engineered TALEs can be fixed to a DNA cleavage domain to produce TAL effector nucleases (TALENs). TALENs can be used to generate highly specific double stranded DNA breaks allowing the introduction of genetic modifications such as gene knockouts and insertions. Using a Golden Gate cloning strategy TALENs directed toward the SNAP25 endogenous loci have been constructed to create a SNAP 25 null mutation. The cleavage efficiency of the SNAP25 TALENS has been investigated by flow cytometry using fluorescent reporter plasmids. Future work involves the investigation of the genome-editing activity of the SNAP25 TALENS in rat cells.