Rifampicin test #2

Piotr

Overnight inoculation and induction:
Top10
and Top10 carrying:
pMPM-T5-AID + 0.01% L-Ara
pTrc99a-AID + 0.5 mM IPTG
pMPM-T5-AID
pTrc99a-AID

Preparation of pMPMT5-AID+T7 and pMPMT5+AID-T7

Piotr, Michał K.

1. Design of primers for sequencing something we want from pMPM-T5.
   Primers:
   
   • pMPMT5seqF
   • pMPMT5seqR
   • AIDseqP

2. Isolation of hypothetical pMPM-T5 with transcriptional fusion.
3. Isolation of hypothetical pMPM-T5 with translational fusion
4. Control digest of pMPM-T5 with transcriptional fusion with EcoRI (EcoRI buffer).
5. Control digest of pMPM-T5 with translational fusion with BamHI (BamHI buffer).
6. Gel electrophoresis (no proper colonies found). Fig. 1.
Fig. 1. 1-DNA ladder;
2 and 3-hypothetical pMPM-T5 with transcriptional fusion digested with EcoRI (2 clones);
4-pMPM-T5-AID digested with EcoRI;
5-hypothetical pMPM with translational fusion digested with BamHI;
6-pMPM-T5-AID digested with BamHI.