Replacing UV with Blue Light during DNA Purification Increases the Efficiency of Ligation-Independent Cloning
Trends in Peptide and Protein Sciences,
Vol. 3 (2018),
1 June 2018
Page 1-8 (e1)
Ligation-independent cloning is a simple method that provides several advantages over conventional cloning. However, the efficiency of ligation-independent cloning is considerably lower than that of conventional methods. Several studies have shown that competent cells used for ligation-independent cloning should preferably have a transformation efficiency of 106-107 cfu/μg DNA. Although such levels can be easily achieved using standard protocols with most Escherichia coli strains, some strains attain mush lower values. When such strains have to be used for ligation-independent cloning, certain measures need to be taken to avoid any situation that may further decrease the efficiency of the process. These measures, however, are usually time-consuming. This problem is exacerbated by the fact that some strains such as BL21 (DE3) appear to be intrinsically unsuited for ligation-independent cloning. Here we suggest that by avoiding DNA damage during purification simply by replacing UV transilluminators with blue light systems BL21 (DE3) cells with a transformation efficiency of 105 cfu/μg DNA can satisfactorily be used for ligation-independent cloning without any additional steps.
•Using blue light instead of UV light increases the efficiency of LIC.
•A simple LED blue light projector combined with a filter can be used for this purpose.
•This approach is not recommended for applications that require higher sensitivity.