Team:Groningen/Notebook/11 August 2009
From 2009.igem.org
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+ | '''Plates''' | ||
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+ | LB-agar plates with 50 μg/mL ampicillin were freshly prepared, and one drop of each o.n. preculture was spotted and subsequently swiped three times with sterile needle. The plates were stored at 37°C overnight. | ||
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+ | '''Precultures''' | ||
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+ | Over night cultures in 5mL LB-amp<sub>50</sub> medium were prepared from the following colonies: | ||
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+ | → E.coli TOP10 pSB1AC3-med. const. promoter - GVP (amp.) (4x) (1 to 4) | ||
+ | → E.coli TOP10 pSB1AC3-low const. promoter - GVP (amp.) (4x) (5 to 8) | ||
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+ | and put in the 37°C waterbath at 200 rpm. | ||
===Transporters=== | ===Transporters=== |
Revision as of 09:30, 11 August 2009
Wet
GVP Cluster
- → DONE isolate plasmids from overnight precultures
- → DONE plate on cultures for storage in fridge (amp)
- → TODO cut plasmid and GVP containing plasmid for ligation (ligation tomorrow)
- → TODO purify wanted fragments (tomorrow)
- → DONE make o.n. precultures from plate colonies
Plasmid Purification
Plasmid isolation was performed on the cultures of E.coli TOP10 containing plasmids J61002 with high, medium and low constitutive promoters with the "Sygma-Aldrich™ GenElute™ Plasmid Miniprep Kit".
- From each tube 4mL of culture was collected in a 2.0mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded.
- Plasmids were eluted with 20μL MQ and stored in the fridge
Concentrations
Plasmid | Conc. ng/μL | 260/280 | 260/230 | -20 box (michael | Restriction Control |
pJ61002-BBa_J23100 (high) | 174.6 | 1.94 | 2.40 | E-8 | No |
pJ61002-BBa_J23106 (medium) | 149.2 | 1.95 | 2.41 | E-9 | No |
pJ61002-BBa_J23109 (low) | 152.8 | 1.94 | 2.38 | E-10 | No |
Plates
LB-agar plates with 50 μg/mL ampicillin were freshly prepared, and one drop of each o.n. preculture was spotted and subsequently swiped three times with sterile needle. The plates were stored at 37°C overnight.
Precultures
Over night cultures in 5mL LB-amp50 medium were prepared from the following colonies:
→ E.coli TOP10 pSB1AC3-med. const. promoter - GVP (amp.) (4x) (1 to 4) → E.coli TOP10 pSB1AC3-low const. promoter - GVP (amp.) (4x) (5 to 8)
and put in the 37°C waterbath at 200 rpm.
Transporters
Metal Accumulation
Vectors
Dry
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