IGEM:Cambridge/2008/Notebook/Turing Pattern Formation/2008/07/28: Difference between revisions
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==Wet Work== | ==Wet Work== | ||
* ''' | * '''Check plasmid ppL82''' | ||
We had 2 samples of ppL82 in DH5α in LB solution (~4mL), one from the frozen glycerol tube and one from a colony picked on a plate. | We had 2 samples of ppL82 in DH5α in LB solution (~4mL), one from the frozen glycerol tube and one from a colony picked on a plate. | ||
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- Digest | - Digest | ||
# For | # For biobricks I746001 and I746101, use digest enzymes EcoRI and SpeI, and 2μL of DNA | ||
#: Add 15μL of SDW, 2μL of 10X Fast Digest Buffer, 2μL of DNA, 1μL of EcoRI and 1μL of SpeI | |||
# For ppL82 (from plate and tube), use digest enzymes BamHI and PstI and add 15μL of DNA | |||
#: Add 2μL of SDW, 2μL of 10X Fast Digest Buffer, 15μL of DNA, 1μL of BamHI and 1μL of PstI | |||
# Microfuge tubes | |||
# Incubate 10 min at 37°C | |||
# Heat shock for 5min at 80°C | |||
- Gel : add 4μL of dye and 21μL of samples | |||
Revision as of 10:16, 29 July 2008
 Turing Pattern Formation
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Results from previous days
We wanted to check the size of our biobricks. For I746101, we have a band of about 3000kb, which is the size of the vectorm and a band of about 2000kb. The size of the biobrick is 2057kp, so it should be ok. For I746001, we hqve the same band of 3000kb (the vector) and a band of about 1000kb. The size of the biobrick is 896kb. So, the size of our biobricks are ok.
For Cm, 35μg/mL should be enough to kill B.S. For Amp, Wet Work
We had 2 samples of ppL82 in DH5α in LB solution (~4mL), one from the frozen glycerol tube and one from a colony picked on a plate. Normally plasmid and cells should not be kept in freezer. So, we want to extract this plasmid, check its size and then keep it in the fridge. We do that for the 2 different samples.
- Test the concentration of DNA in each tube
- Digest
- Gel : add 4μL of dye and 21μL of samples
- Plate a single colony (from 23/07/08) on a Cm plate - Incubate at 37°C
- Incubate at 37°C
-Pick e single colony of MC1061 (from 22/07/08) and put it in 10mL of LB (to make glycerol stocks) -Incubate at 37°C
- Add 100μL of LB+1A (mix of friday) and 10mL of LB - Incubate at 37°C Prepare Medium A- Add 81mL of Sterile Water, 10mL of 10X Medium A base and 9mL of 10X Bacillus salts
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