What is the smallest DNA fragment compatible with In-Fusion Cloning?
What is the smallest DNA fragment compatible with In-Fusion Cloning?
The smallest insert successfully cloned with In-Fusion Cloning was a 50-bp synthetic oligonucleotide (including two 15-nt homologous overlaps with the vector termini). For In?Fusion Cloning of short synthetic oligos (between 50 and 150 bp), the suggested oligo-to-vector molar ratio is 5–15:1. Depending on oligo length, the optimal ratio must be determined empirically. Note: Non-phosphorylated oligonucleotides are compatible with In?Fusion Cloning. However, 3' exonuclease activity in the In?Fusion enzyme mix requires terminal 3' OH groups.