IGEM:Harvard/2006/NES Notebook: Difference between revisions
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# Initial_Streptavidin_Cloning pmid=2404273 | # Initial_Streptavidin_Cloning pmid=2404273 | ||
# Chimeric_Streptavidin_Proteins pmid=11036649 | # Chimeric_Streptavidin_Proteins pmid=11036649 | ||
# Ligand_Binding_Review pmid=11585052 | |||
# Technical_Details pmid=2025272 | # Technical_Details pmid=2025272 | ||
</biblio> | </biblio> |
Revision as of 17:31, 17 June 2006
To Order
- Spe1
- Competent Bacteria -- Ordered by Alain
- Small, Medium, Large, X-large Nitrile gloves
Introductory Materials
New Reporters
Due to issues with T7 promoter, the following biobricks were rehydrated 6/14
Constitutive (IPTG-inducable?) GFP
Constitutive (IPTG?) RFP
Constitutive GFP using TetR promoter. No experience; might not work.
Constitutive (IPTG?) YFP. Apparently gives uneven YFP expression
Transformed with 2ul of DNA on 15ul bacteria; twice for each plasmid. Took 3 ul from each incubation of and added into two two mixed SOC cultures. Let's see what happens when LacI GFP, RFP, YFP are co-cultured.
One of each LacI plasmid type, GFP, RFP, YFP, LacI were cultured overnight on plates treated with 10mM IPTG.
Microscope Notes
Overlap between FITC and YGFP blocks prevents differentiation between YFP and GFP. RFP can be differentiated between YFP/GFP very well.
YFP/GFP fades ~3 hours after being taken out of the incubator after overnight incubation. RFP was still strong at +3h.
IPTG doesn't make an apparent difference in fluoresence.
Streptavidin Research
- Sano T and Cantor CR. Expression of a cloned streptavidin gene in Escherichia coli. Proc Natl Acad Sci U S A. 1990 Jan;87(1):142-6. DOI:10.1073/pnas.87.1.142 |
- Sano T and Cantor CR. Streptavidin-containing chimeric proteins: design and production. Methods Enzymol. 2000;326:305-11. DOI:10.1016/s0076-6879(00)26061-8 |
- Pato C, Le Borgne M, Le Baut G, Le Pape P, Marion D, and Douliez JP. Potential application of plant lipid transfer proteins for drug delivery. Biochem Pharmacol. 2001 Sep 1;62(5):555-60. DOI:10.1016/s0006-2952(01)00708-0 |
- Sano T and Cantor CR. Expression vectors for streptavidin-containing chimeric proteins. Biochem Biophys Res Commun. 1991 Apr 30;176(2):571-7. DOI:10.1016/s0006-291x(05)80222-0 |