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Team:Stockholm/29 July 2010
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It looks like I got a correct band on both samples, however the 2.5 min looks better than the other. Unfortunately I also have a lot of unspecific binding. | It looks like I got a correct band on both samples, however the 2.5 min looks better than the other. Unfortunately I also have a lot of unspecific binding. | ||
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| + | ---- | ||
| + | ===Sequencing of SOD=== | ||
| + | |||
| + | I prepared tubes of SOD for sequencing: | ||
| + | |||
| + | 15 ul plasmid from a mini-prep and 1.5 ul (10uM) primer. Preferably the forward primer (VR) of the vectors verification primers. | ||
| + | |||
| + | * SOD A: ASB0045 179 | ||
| + | * SOD B: ASB0045 180 | ||
Revision as of 16:11, 31 July 2010
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Andreas
CPP troubleshooting
Analyzed sequencing results of our constructed CPPs (LMWP and Transportan 10). It seems like our target vector recircularizes instead of ligating with our CPP primers. Me, Johan and Nina are working on troubleshooting this, however it looks like we will soon have to synthesize our genes, since our primer ligations just don't seem to work.
Nina
Protein expression of IgG protease from pEX
BL21 cultures of IgG in pEX and bFGF in original vector from plates used to inoculate for IPTG induction:
A streak of colonies from the IgG plate was inoculated in two separate falcon tubes containing 12 ml LB and 24 ul ampicillin (50 mg/ml). ~10 colonies from the bFGF plate was inoculated in two separate falcon tubes containing 12 ml LB and 24 ul ampicillin (50 mg/ml). This was incubated with shake in 37 °C until OD reaches around 0.6.
PCR on Tyrosinase
I have had trouble obtaining a band representing tyrosinase on a 1 % agarose gel after running a PCR. Therefore I ran two new PCR:s, one with an annealing time of 2.5 min and another with 3 min.
1 = 2.5 min
2 = 3 min
DNA Ladder: FastRuler™ Middle Range, ready-to-use, 100-5000 bp Fermentas
It looks like I got a correct band on both samples, however the 2.5 min looks better than the other. Unfortunately I also have a lot of unspecific binding.
Sequencing of SOD
I prepared tubes of SOD for sequencing:
15 ul plasmid from a mini-prep and 1.5 ul (10uM) primer. Preferably the forward primer (VR) of the vectors verification primers.
- SOD A: ASB0045 179
- SOD B: ASB0045 180
