**** NEW: Relaxed PAM (TNTN) ttLbCas12a (pICSL90029) and multi-intron PAM-less Cas9 (pICSL90030) constructs have now been added to the Synbio repertoire. Also RUBY (He et al) non-invasive gene monitoring reporter (pICSL80094) construct added. ****
This page contains the recipe and reaction condition for assembling multiple overlapping fragments.
Read this article for an overview of Gibson Assembly and primer design.
5X ISO buffer. This recipe makes 6ml.
Gibson Assembly Enzyme Mix. This recipe makes 1.2ml
1. Thaw a 15 μl aliquot of Gibson Assembly Enzyme Mix on ice.
2. Add 5 μl of DNA (equimolar amounts of each fragment to a total of 50-200 ng)
3. Mix gently.
5. Incubate at 50 °C for 15 to 60 min.
6. Transform 1 – 5 μL.