Difference between revisions of "Part:BBa K374008:Experience"
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*pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP | *pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP | ||
| − | [[Image: | + | [[Image:Gifsy1_2GFP.png]] |
*pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter | *pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter | ||
*pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP | *pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP | ||
| − | [[Image: | + | [[Image:Gifsy1_2GFPBiomassvsTime.png]] |
*pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter | *pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter | ||
*pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP | *pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP | ||
Revision as of 21:56, 27 October 2010
This experience page is provided so that any user may enter their experience using this part.
Please enter
how you used this part and how it worked out.
Applications of BBa_K374008
Biolector experiment
The biobrick was cloned into the pSB4A5 plasmid, and a GFP reporter gene was inserted downstream of its pR promoter (pIGR01_04). The plasmid was transformed into E.coli (DH5 alpha) and measurements were done on this strain using a BioLector microreactor system. The BioLector is a microfermenting system that measures OD and fluorescence simultaneously. From this construct no GFP expression is expected as repression of the pR promoter by the GogR repressor is tight. As a reference the repressor was removed from the construct (pIGR03_11) allowing GFP expression from the pR promoter.
Procedure
- OD600 of a 10 mL O/N LB culture with required antibiotics was measured
- Culture was diluted to an OD600 of 0.05
- 1.5 mL of diluted culture was added to each of the 48 wells
- An Adhesive Gas Permeable Seal membrane was applied to the plate
- An Adhesive Seal for Evaporation Reduction on top of the Gas Permeable Seal was applied
- The microplate was incubated in the biolector and fluorescence and light scattering was measured under these conditions:
- 37 degrees Celcius
- sampling time 6 times
- fluorescence gain of 80
- biomass excitation 620 nm (light scattering)
- GFP filter was 486nm (ex) / 510nm (em)
Data
- pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter
- pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP
- pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter
- pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP
- pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter
- pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP
- pIGR01_4: pSB4A5 backbone with Gifsy1 pR and pRM promoters. GFP is expressed from the pR promoter
- pIGR03_11: pSB4A5 backbone with BBa K374008 followed by a GFP
Conclusion
User Reviews
UNIQab305b9672c66da4-partinfo-00000000-QINU UNIQab305b9672c66da4-partinfo-00000001-QINU