Difference between revisions of "Davidson Missouri W/Davidson Protocols"
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# When inducing with IPTG, use '''3 µL of stock''' (0.2 g/mL = 20% w/v) '''to every 1 mL''' of LB or other liquid. | # When inducing with IPTG, use '''3 µL of stock''' (0.2 g/mL = 20% w/v) '''to every 1 mL''' of LB or other liquid. | ||
# When inducing with Arabinose, use "2 µL of stock" (10% w/v L-Arabinose) "to every 1 mL" of LB or other liquid. | # When inducing with Arabinose, use "2 µL of stock" (10% w/v L-Arabinose) "to every 1 mL" of LB or other liquid. | ||
| + | # When inducing with 3OC6 (HSL), use a '''2000 fold dilution of a 10 mg/mL stock solution'''. We have dissolved in EtOH which is not the best - degrades with time. Keep this cold. | ||
# [http://partsregistry.org/AHL List of auto-inducers and their catalog numbers] | # [http://partsregistry.org/AHL List of auto-inducers and their catalog numbers] | ||
Revision as of 18:32, 25 March 2010
A. General Lab Information
- How to Keep a Lab Notebook
- Common molecular reagents
- Standard Assembly
- BioBrick Ends
- Compatibility of Plasmids
- Ethanol Precipitate DNA (short protocol)
- glycerolstocks How to Make Glycerol Stocks of Bacteria
B. Gel Electrophoresis and Purification
- Pouring an agarose gel
- Calculate MWs
- 1kb MW markers
- Macherey-Nagel Gel Purification (improved 260/230 ratios)l
- Qiagen QIAquick Gel Purification
- Qiagen QIAquick Column Regeneration Protocol
- ElectroElute Gel Purification
C. Digestion, Ligation, Transformation
- Digest DNA with restriction enzymes
- Double Digest Guide
- Shrimp Alkaline Phosphatase
- Ligation Protocol
- Choices for Transformation: Heat Shock vs. Zyppy
- Heat Shock Transformation OR Short version of Heat Shock
- Zippy Transformation
- TSS Competent Cell Transformation
D. Minipreps
E. Making New Parts and PCR
- Building dsDNA with Oligos
- Setting up PCR mixtures
- PCR and Mg2+ concentration
- Making dsDNA Using Primer Dimers
- Clean and Concentrate DNA (after PCR, before digestion)
- Colony PCR to Screen for Successful Ligations
F. Expression of Phenotypes
- Using degradation tags on proteins such as GFP
- Genomic Insertion Protocol
- When inducing with IPTG, use 3 µL of stock (0.2 g/mL = 20% w/v) to every 1 mL of LB or other liquid.
- When inducing with Arabinose, use "2 µL of stock" (10% w/v L-Arabinose) "to every 1 mL" of LB or other liquid.
- When inducing with 3OC6 (HSL), use a 2000 fold dilution of a 10 mg/mL stock solution. We have dissolved in EtOH which is not the best - degrades with time. Keep this cold.
- List of auto-inducers and their catalog numbers
G. Computer Tools We Use
- Optimize your Gel
- Gene Splitting Web Site
- PCR Primers w/ BioBricks
- Promega Tm Calculator
- Lance-olator Oligos for dsDNA assembly
- Access the GCAT-alog of Davidson and MWSU DNA Freezer Stocks
- Sequencing at Agencourt Bioscience
- Sequencing at CUGI
- Analyzing Sequences with aPe
- Using Apes (A Plasmid Editor)
