Week 2: First Success

New week, New try!

On Tuesday I set up a new reaction for my SDMs...and again disappointment on Wednesday morning...no colonies at all!

One thing I have definitely learnt by now is not to take it to heart and to keep trying! 

So I precipitated the rest of my PCR product again and did another transformation of E. coli DH5 alpha...finally, a pleasant look into the incubator on Thursday morning: one colony for each mutant!!

Finally, the transformation with my mutant plasmids worked

I guess one has to treasure those moments of joy when experiments actually do work and remember them when things aren't working quite as desired (again)!

To verify the mutation we sent off the SDM plasmids for sequencing ... let's hope they each have the mutation we wanted!

I simultaneously started preparing the over-expression of my WT InhA by transforming E.coli BL-21 with my initial plasmid containing the wild type gene inhA which worked well first time!

Transformed E. coli BL-21

Another worthwhile skill in the lab: multitasking! Making use of incubation times by e.g. making more agar and broths, cleaning up common areas, thinking ahead and planing your days and week...or just having a good laugh with people around you!

Lots of LB broth to over-express lots of InhA

My task for the next two weeks will be to make as much InhA as possible for Paul's post-doc and Phd to take down to the Rutherford Appleton Laboratory (RAL) for 2D-IR Spectroscopy which will give more detailed and specific data with a greater time resolution due to the advanced laser.