Getting ready for punctual mutation
- Today, I inoculated E. coli strains containing a plasmid with a constitutive promoter so that, after 6-7 hrs of incubation at 37°C, I can extract the plasmid and ligate the luciferase and double terminator with it.
- I also did a restriction with Xba I and Pst I for a total of 30μL as follows:
-H2O -------> 18μl
-Buffer 2 --> 4μl
-BSA ------> 1μl
-DNA -------> 5μl
-XBA I -----> 1μl
-PST I ------> 1μl
- Restrictions were incubated at 37°C for 4 hr and labeled as: lucif + TT c/rbs + 1 purif restr. XBA y PST Mar and the date in the top of the eppendorf.
- I also ran a gel at 90V for 50min to quantify the ligation PCR product with the mutated RBS to know how to prepare the mix for the PCR with Pfx.