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Team:DTU-Denmark/Notebook/12 June 2013
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==Main purposes today== | ==Main purposes today== | ||
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New PCR amplification of USER fragments | New PCR amplification of USER fragments | ||
==Procedure== | ==Procedure== | ||
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===PCR=== | ===PCR=== | ||
MasterMix x 17 was made | MasterMix x 17 was made | ||
Revision as of 10:18, 26 July 2013
12 June 2013
Contents |
208
Main purposes today
New PCR amplification of USER fragments
Procedure
PCR
MasterMix x 17 was made to each tube 3ul of Fw and 3 ul of rv primer was added as well as 1ul DNA template
every fragment was run 2 min 98 C followed by 35 cycles of
10 sec 98 C
1 sec 65 C ramp by 0.1C/s down to
10 sec of 55C
3 min 72C
after repetitions, 5 min of 72 C
prepared
- 1l of 10x TBE
- 200ml of 2% agarose solution and added 10 ul EtBr
- casting 3x 2% gels and 1x1% gel
- 200ml EDTA stock solution 0.5 M from powder, dissolved at pH 8 and later equilibrated to pH 7
- 200ml LB medium w. agar and kanamycin
- autoclaved 1.5l of LB medium
- new batch of competent cells
- glycerol stock from I14032
run gel of todays PCR
got SFGFP for TAT and SFGFP for Sec
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