Lab Notebook

14th July

Lab work: Sriram & Orr

Prepared 1 litre of LB agar medium and sent to Kluyver lab's kitchen.

Using 650 µl of competent cells (E.coli DH5alpha) and the transformation protocol, we inserted 13 biobricks [R0010, I14018, I12006, R0040, J23008, J23031, E0040, E1010, C0051, B0034, B0015, S03335, S03473, K081013 and C0040] for delay into competent cells and grew them in a solid media to increase the amount of biobricks we will have.

Diluted the 13 biobricks with 15 µl RNase free water and stored in -20°C freezer.

Standard transformation procedure:

Remove competent cells from -80°, let thaw for 10 min on ice and aliquot in 50 µl amounts.
add 2-5 µl of vector, to 50 µl cells, no mixing by pipet due to shear induction.
keep on ice for 20 minutes (vector spreading through volume).
heat shock (42°C) for 45 seconds.
keep on ice for 2 minutes.
add 200 µl SOC, put on 37°C for 1 hour or longer with agitation (160 rpm).
plate out 250 µl on appropriate antibiotics.

Weenink

Brought beaker of eppendorf tubes to autoclaving

Calin

Looked into the details of lambda red knockout protocol. Started designing the required primers.