TALEN Gene Editing: 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. 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 across both arms (mismatched repeats 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.

A T G C TALE repeat (match)

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