In order to make the new biobricks, the 4 new parts that were digested with the enzymes XbaI and SpeI must be ligated to the biofusion vector, that was also already digested with the same enzymes. So today we prepared the ligation reactions (according to Protocol 11) that will be kept at 4ºC O/N.
Taís
ColiGuard
Starting the biobricks of kill mechanism with colicins
We made the culture for miniprep of the transformed cells with the biobrick BBa_J131009 (operon colicin-E2). We want to isolate the CeaB (colicin protein) and the CeiB (immunity protein) in order to put them separately in the conjugative plasmid and in the genome of E. coli respectively and according to the killing mechanism (explained in the project description). To fulfill this aim, we designed 3 “primers” for isolating. For isolating the CeaB, we used like forward primer, VF2 (general primer forward for backbone plasmid) and like reverse primer, our oligonucleotide designed for us, ColR. We also designed 2 primers forward and reverse for isolating the CeiB: Anticol F and Anticol R.
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