Figure 4
From: ZeBRα a universal, multi-fragment DNA-assembly-system with minimal hands-on time requirement
Three or four fragments can efficiently be assembled with PPY-cell extracts, while iVEC/“transformation-cloning” with three fragments is markedly less efficient. (a) The assembly of four fragments in a single reaction reduces the number of successful assemblies by a factor of two to ten as compared to the three-fragment-assembly. Cell-extracts were prepared from autoinduced PPY cells and the assemblies were purified prior transformation. (b) Assembly of three fragments by iVEC/“transformation-cloning” with NEB 5-alpha resulted in roughly 250 recombinant colonies/µg transformed DNA. A significant number of colonies harboring plasmids with defective inserts (grey bar) and PCR-template carry-over (dotted bar) were present on the plates.
