Valencia/October
From 2007.igem.org
Work Progress
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← September
1/10/07
we leave some overnight PCRs reactions, using VF2 and VR, in order to properly analyze our clones...
at the same time, we are inoculating all those clones in order to make some minipreps.
2/10/07
we have done the gel of the PCR reactions, but it looks like the DNA has not amplified our samples, but the reaction has worked fine for the controls.
we repeat the PCRs and make some other controls with different DNA quantities. the gels shows some bands, but their weights do not correspond to anything we wanted... could be some other lab dwarf??
we have miniprepped the inoculations that have grown overnight. this way, we will have more DNA to sequence
3/10/07
we have digested the plasmid that were amplified with the PCRs with EcoRI and PstI, because we want to be sure if it is, or if it is not, what we want... at the gel we see two bands on one clone and three (??) on the other... and the weights are not the correct ones... we must have had some false positives clones...
They ARE BioBricks, as they get amplified with VR and VF2 and their miniprep yields a plasmid that corresponds to the ones we want. But, for the construction pLac-TetR-CFP, when we have digested it with EcoRI and PstI we see three bands: 2,2 kb, 2 kb and 400 bp. For the construction pTet-LacI-YFP, we see two bands: one at 3kb and the other at 2,8 kb. sadly, this does not correpond to any simple, straigh forward hipothesys we may come up with...
we doing more sequencing reactions of pTet-LacI. We are 100% sure that our pLac-TetR is what is what meant to be, but with the consensus sequence of pTet-LacI still has some gaps and missed parts. We want to be sure to have the correct sequence of the comparator part
we prepare more inoculations to purify tomorrow.
4/10/07
we are miniprepping and digesting the comparator device (pTet-LacI and pLac-TetR) with the fluorescences (CFP and YFP). we are purifying it and concentrating it so tomorrow we could ligate it.
