Difference between revisions of "Team:CSU Fort Collins/NoteBook/JanThroughApr"
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</div> | </div> | ||
| + | <div class = 'Days'><h5>4/12/16:</h5></div> | ||
| + | <div class = 'Info'><h6></h6><ul><li> Reaction: PSB1C3 3.06 uL,Q1 2.5ul,Q2 1.78ul, 5x Infusion Mix 2ul, H20 .66ul</li> | ||
| + | <h6>Transformation</h6><li>Add 1 uL Infusion rxn to 50 uL Stellar Cells</li> | ||
| + | <li>Incubate 30 min on ice, 45 sec at 42C, 2 min on ice</li> | ||
| + | <li>Add 250 uL SOC media</li> | ||
| + | <li>1 hour outgrowth at 37C</li> | ||
| + | <li>Plate 50 uL on LB+Cam plates</li> | ||
| + | <li>37C overnight</li> | ||
| + | </ul> | ||
| + | </div> | ||
</html> | </html> | ||
Revision as of 22:47, 16 October 2016
CSU iGEM
January Through April
1/4/16:
- Ligated b0034 and c0061
- Transformed Ligated mixture
- Ran PCR results on a 2% gel
1/5/16:
- Digested LuxR with EcoRI and SpeI
- Performed Colony PCR on transformation from the 4th
1/11/16:
- slr0435 Excision from Synechocystis sp. PCC6803 with PCR using Phusion(size 276 bp).
- Ran PCR results on a 2% gel
- Digested I0462 and E0030 with EcoRI and SpeI, R0062 and B0015 with EcoRI and XbaI, B0034 with SPeI and PstI, and C0061 with XbaI and PstI.
1/12/16:
- Digested slr0435 and resuspended pSB1C3 with EcoRI and PstI
- Ligated pSB1C3 and slr0435
- Transformed ligation mixture
1/13/16:
- Colony PCR performed on slr0435+pSB1C3 transformed colonies
- Ran PCR results on 1% gel
- Digested I0462 and E0030 with EcoRI and SpeI, R0062 and B0015 with EcoRI and XbaI, B0034 with SPeI and PstI, and C0061 with XbaI and PstI.
- Ran on 1% gel
1/21/16:
- Digested I0462 and E0030 with EcoRI and SpeI, and C0061 with XbaI and PstI.
- Ran on 1% gel
- Phosphatase treatment
- Use Antarctic Phosphatase with 10x buffer. Need 1uL to treat 1ug of DNA
- Add 1uL phosphatase and 5uL 10x buffer to bring total rxn volume to 50uL
- Incubate at 37C for 1 hour, and heat inactivate at 70C for 5 minutes
1/25/16:
- Overnight ligated R0062 and I0462, B0034 and C0061, B0015 and E0030 at 16 C
1/25/16:
- Transformed the overnight ligation mixtures
2/6/16:
- PCRed out BBA_J23106 out of glycerol stock 19 using Q5 master mix 19rxns
2/9/16:
- Transformed Bba_K325909 from parts kit into DH5a cels
2/11/16:
- To confirm that Bba_K325909 are functioning we tried to induce expression using L-arabinose
- No luminescence was produced
2/16/16:
- Decided to order two gBlocks for quorum portion of the project
3/24/16:
- Set up O/N cultures of K1033929, K592009 and E1010 in 5 mL of LB with 5 uL of Cm34 antibiotic
3/29/16:
- Ran promoters Sll027, and LrtA in a 1% gel
Sll0027
LrtA
4/6/16:
- Ran Cpcg2 on 1% gel
4/12/16:
Prepare pSB1C3 vector for Infusion
- gBlocks (Quorum 1 and Quorum 2) have 15 bp homology with each other along with prefix and suffix at the entire block’s ends
- Digesting the vector with XbaI will leave 16 bp homology with Quorum 1
- Digesting the vector with SpeI will leave 20 bp homology with Quorum 2
- Digested c0061 with SPeI and XbaI
- Ran on 1% gel
4/12/16:
- Reaction: PSB1C3 3.06 uL,Q1 2.5ul,Q2 1.78ul, 5x Infusion Mix 2ul, H20 .66ul
- Add 1 uL Infusion rxn to 50 uL Stellar Cells
- Incubate 30 min on ice, 45 sec at 42C, 2 min on ice
- Add 250 uL SOC media
- 1 hour outgrowth at 37C
- Plate 50 uL on LB+Cam plates
- 37C overnight
Transformation
