July 3
Checked Gibson transformations
Plated remaining transformed cells
Made new dilutions of original enrichment cultures (BPA, nonylphenol, 17-alpha-ethinyl-estradiol, DDT)
Results
There was no growth on the transformed Gibson plates. Since we did a small reaction, we used all of the reaction in transforming competent cells. The Gibson assembly likely failed, as one of the negative control plates had a single colony. However, we have had trouble with our competent cells in the past and will try plating what we have left.
Gibson Transformations
| Plate |
Number of Colonies |
| pNT001 + |
0 |
| pNT001 - |
0 |
| pNT002 + |
0 |
| pNT002 - |
1 |
July 4
Results
After plating the remaining 450 ul of our transformed cells:
| Plate |
Number of Colonies |
| pNT001 + |
10 |
| pNT001 - |
0 |
| pNT002 + |
1 |
| pNT002 - |
0 |
This Week
Start enrichment cultures from a previous iteration, as the latest dilutions remain clear.
Design experiments to test for degradation of BPA by pNT001 and pNT002 using the HPLC
Get the BioBricks we ordered from the parts registry ([http://partsregistry.org/Part:BBA_K364309 K364309], [http://partsregistry.org/Part:BBA_K364329 K364329], [http://partsregistry.org/Part:BBA_K364327 K364327], [http://partsregistry.org/Part:BBA_K364307 K364307])
Transform these into cells, miniprep and sequence
Continue with enrichment cultures
Put LA River soil DNA into fosmids
Work on Project Description and Safety Proposal
Once we've sequenced ER ([http://partsregitry.org/Part:BBa_K123003 K123003]), start making test plasmids to see if it works.
July 5
Start overnight cultures of Gibson transformations
July 6
Miniprep Gibson transformations and send off for sequencing
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