Team:Cambridge/Diary/Week 7

Monday

Like many days, today was a good day for shopping. No shoes and salads for us though - we had chemicals to order. NAD, Iron (III) chloride, LB base and potassium acetate, all for the greater glory of our project.

Oli re-ran his vector fragment B PCR and, despite some slightly weak bands, extracted the DNA needed. A similar retry of the magnesium sensitive promoter failed though, and a quick look at the registry revealed that the reliability of the part (both in sequence and content) was somewhat suspect. Irritating, but it will look good when we make our own really good magnesium sensor. Which we will. Don't worry.

We've run out of competent e.coli cells, and we need some more. Unfortunately, the TOP10 cells that were grown up over the weekend were too old to use, so Paul is using our final vial of TOP10 to make some more that should be in the log phase by tomorrow morning. Then we can get on with the protocol to make them competent.

Tuesday

A very practical day today, as various people (Paul and Andreas in particular) undertook to perform the rather long electro-competent cell production protocol. We also recieved our delivery of reagents we ordered yesterday. Using some quite stinky NAD+, the isothermal reaction buffer for the Gibson assembly reaction was finished off and lots of aliquots of Gibson master mix stored.

Having made all this, we did some Gibson for the riboswitch and fluorescent constructs, which *should* work this time. Certainly, we're pretty confident we have all the ingredients in the mix this time. We'll transform some cells with the resultant DNA cocktail as soon as we have some competent cells. Hopefully tomorrow then, though such efficient hopes have had a way of failing in the last couple of weeks.

Tom struggled through his PCR again, retrying the split luciferase primers. No successes, alas...

Wednesday

Thursday

Friday