Optimal Length for Left & Right Homology Arms?

DNA with homology to the sequences flanking a double-stranded break (DSB) can serve as template for homology directed-repair (HDR) of the DSB. The efficiency of HDR is determined by the concentration of donor DNA present at the time of repair, the length of the homology arms, the cell cycle, and the activity of the endogenous repair systems in the particular cell[1].

These factors contribute to the high variability of HDR efficiency observed across different cell lines, and particularly in immortalized cells [2].

While longer inserts are possible, the efficiency of recombination decreases as the insert size increases [3]. Finding successfully integrated inserts is likely to be challenging when inserts are greater than 3 kb in most mammalian cells.

Homology arm length varies based on insert length. IDT's HDR Design Tool will suggest the optimal homology arm length based on the insert size and our empirically supported design rules.

References:

1. Elliott B, Richardson C, Winderbaum J, Nickoloff JA, Jasin M. Gene conversion tracts from double-strand break repair in mammalian cells. Mol Cell Biol. 1998;18(1):93-101.
2. Lin S, Staahl BT, Alla RK, Doudna JA. Enhanced homology-directed human genome engineering by controlled timing of CRISPR/Cas9 delivery. Elife. 2014;3:e04766. Published 2014 Dec 15. 
3. Li K, Wang G, Andersen T, Zhou P, Pu WT. Optimization of genome engineering approaches with the CRISPR/Cas9 system. PLoS One. 2014;9(8):e105779. Published 2014 Aug 28.