TALEN Gene Editing 2D: RVD Code & FokI Dimerization

TALE Array Design

Repeat array length (per arm) 15

Spacer length (bp) 14

RVD mismatches introduced 0

Live Readouts

RVD match score

100%

Spacer window

Matched bp (both arms)

Expected off-targets

–

Left arm RVDs: —

How it works

A TALEN monomer is an engineered TALE (Transcription Activator-Like Effector) DNA-binding domain fused to a FokI nuclease domain. Unlike zinc fingers (3 bp per finger), each TALE repeat recognizes exactly one base pair through two hypervariable residues — the repeat-variable diresidue (RVD):

NI → A     HD → C
NG → T     NN → G (or A)

Two TALEN monomers are designed against two DNA half-sites flanking a central spacer. FokI only cuts as a dimer, so cleavage requires both monomers to bind with their FokI domains close enough to associate — in practice spacers of about 12–21 bp place the two FokI domains in register:

cut occurs  ⇔  12 ≤ spacer ≤ 21  AND  mismatches ≤ 2

Specificity comes from the combined length of both half-sites that actually land on the 60 bp target. Treating each matched repeat as an independent 1-in-4 filter, the expected number of look-alike sites in a genome of length L is approximately:

E[off-target sites] ≈ L / 4^n
n = matched repeats that actually fall on the target (mismatches, and any
    arm positions that would run off the end of the 60 bp window, don't count)

Longer, more accurate arrays push this number toward zero (used here against a 3.2 Gbp human-genome-sized L); a handful of mismatches — or a spacer outside the FokI dimerization window — collapses cleavage even though each monomer may still bind. Drag to pan the linear map, scroll or pinch to zoom.

A T G C TALE repeat (match)

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