Lab Notebook

3 August 2009

Tim Weenink

I stole p(tetR) promotor and double terminator from the delay team to redo the !B and !C assemblies. The ligation products were run on Calin's gel (see below). The transformants were plated on Cam agar plates (unfortunatele only LB agar was left that seemed infected.) and stored in the 37 degrees C stove.

Sriram

All the transformations done on friday gave growth. However comparing to the chemically competent cells prepared by TMF buffer (RbCl) the number of colonies from TSS buffer competent cells was less. However the concentration of DNA added plays a major role in the efficiency of transformation. The λp-R-GFP-T DNA isolated by miniprep had a concentration of 116.5 ng/µl which gave huge number of colonies compared to the diluted pSB1K3 with mRFP biobrick. The electro-transformation of λp-rep biobrick was successful and gave good colonies. hence we have decided to work with electro-transformation hereafter.

Thus we can very well start the assemblies today. We did restriction for the DNA samples needed for 8 assemblies. We might have some problem with backbones but only in future assemblies.

Calin

Tim pointed out that the plasmids should be linearized before doing the gel. Here are possible digestions on some of the parts:

Growth on all plates. The pSB4C5 and R751 plates were parafilmed and placed in fridge.

Did linearization of oriT-R, promoter, pSB1AK3, and pSB4C5 using EcoRI.

Did digests of oriT-R (9uL) and pSB4C5 (4.5uL).

Did ligation CA and CB:

Ran a 2% gel to check various parts and digests.

Orr

Made some LB Agar medium and some agarose gel with pre-made TBE at concentration 1x.