Team:Cambridge/Notebook/Week7

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Week 7

Monday

Wet Work

Amplification

Colony PCR of 8 colonies picked from 391 in pSB3K3 transformants, 8 colonies picked from 374 in pSB3K3 transformants, 8 colonies picked from 384 in pSB3K3, and 1 colony picked from 381 in pSB3K3 transformants confirmed to contain an insert of the right length. Overnight cultures of each of those colonies as well. No 374 constructs were found to contain inserts of the right length, however 4 384 constructs were of the correct length. The 381 construct was confirmed once again to be the correct length.

Overnight,

  • Restirction digests of pSB3K3, 380, 382, 385, and 390.
  • Colony PCR of 10 new 391 in pSB3K3 transformants, another 374 in pSB3K3 transformants

MelA BioBrick

PCR done of the origonal MelA and the first MelA intermediate (without the second PstI site), to create the DNA for a PstI restriction digest. After PCR the DNA was run on a CYBRsafe-stained gel and then extracted with the QuiGen kit. A Restriction digest was carried out using the fastdigest buffer and PstI and left for three hours.

We also tested the Primer C again, as it didn't work last time. Three PCR's were done:

  • Primer F and Primer C (same aliquot as used last time, to check it wasn't just an experimental error on my part)
  • Primer F and a new aliquot of Primer C
  • Primer F and revortexed and re-aliquoted primer C from the origonal vial

None of these produced any result - there is clearly an error in the primer that I ordered.

Dry Work

We found the phzM and phzS pigments in the registry!

  • BBa_I723024 (phzM)
  • BBa_I723025 (phzS)

This means we can now start working on these pigments, connecting them to processing and logic systems.

Development of Matlab programs to automatically process plate reader data was continued.

Tuesday

Wet Work

Amplification

Colony PCR of 391 in pSB3K3 showed none had the correct insert. Ligations of 380, 382, 385, 390 into pSB3K3, transformed into Top10.

Dry Work

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