Team:Stockholm/1 October 2010
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==Andreas== | ==Andreas== | ||
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*Save pellet from the culture to purify protein | *Save pellet from the culture to purify protein | ||
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+ | ==Johan== | ||
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+ | Realized I had put in the wrong antibiotic in my overnight cultures (doh!). Made new overnight cultures for tomorrow. | ||
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+ | {{Stockholm/Footer}} |
Latest revision as of 21:07, 27 October 2010
Contents |
Andreas
LB agar plates
- 20 x 100 μg/ml Amp LB agar plates
Assembly of His⋅SOD⋅cCPP constructs
Digested pMA.His⋅SOD constructs for later assembly into cCPP plasmids, digested by Johan.
Digestion
pMA.His⋅ SOD | |
---|---|
10X FastDigest buffer | 3 |
DNA (3 μg) | 7.5 |
dH2O | 17.5 |
FD EcoRI | 1 |
FD AgeI | 1 |
30 μl |
- Incubation: 37 °C, 0:30
- Inactivation: 80 °C, 20 min
Stored in -20 °C for later ligation.
Nina
Miniprep
I performed a miniprep on IgG protease_Tra10_Ntermin#4 and Fusion_NS#2 according to the procedure that is described in protocols.
Send for sequencing
I sent two samples for seqeuncing. The mixture was 15 ul sample and 1.5 ul forward bank vector verification primer VF.
- IgG protease_Tra10_Ntermin#4 ASB0045 682
- Fusion_NS#2 ASB0045 681
Overnight culture
I inoculated 12 ml LB and 24 ul chloramphenicol with IgG protease_Tra10_Ntermin#6 again since I acidentally droped the previous sample and therefore lost it.
iGEM Uppsala collaboration
I drove to Uppsala and started a collaboration with their team. I obtained a construct that they want me/our group to PCR with verification primers (VF2 & VR) and run on an agarose gel. I in turn gave them the tyrosinase gene to also amplify via PCR but with own designed primers.
- Uppsala iGEM team's PCR master mix and prgm:
Mimmi
Over expression
- Start culture (9:30)
- 20ml LB + 200µl culture from ON
- At OD=0.6 add IPTG 1mM (checked at 12:00, allready too high OD, had to dilute...)
- Take samples at
- 0h
- 30min (50min)
- 3h
- Spinn down and resuspend in 100µl SDS-buffer
- dilute 3h samples 1:4
- heat at 95°C for 10min
- freeze
- Save pellet from the culture to purify protein
Johan
Realized I had put in the wrong antibiotic in my overnight cultures (doh!). Made new overnight cultures for tomorrow.
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