Life Technologies

27 Life Technologies ™ | Genome editing Figure 8. Designing target sites for customized TAL effectors for maximal binding. (A) GeneArt ® Precision TALs encode a DNA-binding protein specific to a customer-submitted sequence, fused to a FokI nuclease domain for genome editing. The sequence targeted by our first-generation TAL effectors must have a T at its 5´ end, and spacing between forward and reverse TALs must be 13–18 bp for proper pairing of the FokI nucleases and creation of a double-stranded break. (B) GeneArt ® PerfectMatch TALs eliminate the 5´ T constraint of GeneArt ® Precision TALs. GeneArt ® PerfectMatch TALs allow targeting of any sequences across the genome; 15–16 bp spacing between the two TAL effector targets is optimal for GeneArt ® PerfectMatch TALs. DNA binding domain Functional domain FokI FokI TNNNNNNNNNNNNNNNNNN NNNNNNNNNNNNNNNNNNT 13–18 bp DNA binding domain Functional domain FokI FokI NNNNNNNNNNNNNNNNNNN NNNNNNNNNNNNNNNNNNN 15–16 bp A B DNA binding domain Functional domain Fok1 Fok1 An MCS version of the GeneArt ® Precision TAL vector allows you to customize the resulting TAL fusion protein with an effector domain of your choice. Precision TALs fused to a Fok1 nuclease allow sequence- specific, double-stranded DNA breaks to be introduced. Precision TALs fused to a vp16/KRAB functional domain allow specific activation/repression of gene expression. Gene targeting (Fok1 nuclease pair) • Silencing • Incorporation of exogenous DNA Activation of transcription (Activator vp16 or vp64) • Increasing the expression level of endogenous gene isoforms Epigenetic repression of transcription (KRAB) • Heritable knockdown of gene expression Steric repression and custom design (MCS vector) • Transient knockdown of gene expression • Target any locus in the genome with the effector domain of your choice (multiple cloning site vector) Figure 7. Available effector domains

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