Team:Cambridge/Protocols
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Revision as of 12:41, 19 September 2012
Contents |
Protocols
Safety
See our Safety Page for Associated risk assessments for the protocols below and MSDS sheets for the reagents we have used.
Methods
Construct production
- Gel Electrophoresis A technique for separating DNA strands of different lengths.
- Gel Extraction of DNA A technique used to isolate a desired fragment of intact DNA from an agarose gel following agarose gel electrophoresis.
- Gibson Assembly A technique for ligating multiple DNA fragments in one step, compatible with standard assembly.
- PCR using a high temperature DNA polymerase A method for amplifying a section of DNA.
- Making a Biobrick A final step to put your construct into the standard biobrick format to be submitted to the registry.
Transformation protocols
- Chemically competent cells generation A technique to produce e.coli cellsreceptive to chemical transformation.
- Electocompetent cells generation A technique to produce e.coli cells receptive to transformation by electroporation
- Electroporation A method for transforming appropriately competent cells with plasmid DNA using electricity.
- Glycerol stocks A technique for long term storage of cells at -80 degrees without losing cell vitality.
- Transformation of Bacillus subtilis A technique used to introduce foreign DNA into competent Bacillus cells.
- Transformation of Escherichia coli A method for transforming competent E.coli with DNA
Construct verification
- Colony PCR PCR with cells as a template. Useful for checking the length of an insert in an introduced plasmid.
- Restriction Enzyme Digest A method for creating a restriction map of a plasmid.
- MiniPrep - DNA extraction A method used to extract DNA from bacterial cells.
Ribosense testing
- β-galactosidase assay Assay to qualitatively measure the amount of the enzyme β-galactosidase being produced by a population of cells. Useful as a reporter system.
- Miller assay Assay to quantify the amount of the enzyme β-galactosidase being produced by a population of cells. Useful as a quantitative reporter system.
- Production of magnesium free cells. Technique to produce cells that contain little or no magnesium, to control for this variable in tests involving the magnesium riboswitch.
Ratiometrica testing
- IPTG induction A technique for inducing the pSPANK promoter in E.coli cells.
Sporulation and Germination
- Imaging spores A method for seeing vegetative cells and bacillus spores under the microscope.
- Producing high yields of spores A method for obtaining high yields of spores from B. Subtilis
Miscellaneous
- BioBricks Resources for manipulation of BioBricks and information regarding the distribution.
- LB Agar Plates preparation A method used to prepare agar plate to culture common bacteria.
- SDS PAGE protein analysis A method used to separate polypeptides of different lengths.
Recipes
- M9 minimal medium A recipe for making M9 minimal medium for use when autofluorescence must be avoided.
- Medium B A recipe for making medium B.
- CCMB80 for chemically competent cells generation A recipe for making CCMB80
- TAE buffer for gel electrophoresis A recipe for making 10xTAE buffer
- SOB growth medium A recipe for making SOB growth medium
- CDSM A Chemically Defined Media to induce sporulation in B. Subtilis
- Germination Media A Chemically Defined Media to induce Germination in B. Subtilis
Lab supplies
- [http://www.bioc.cam.ac.uk/stores/ BioPath Stores]
- [http://www.plantsci.cam.ac.uk/camonly/stores/stock.xls Plant Department Stores Catalogue xls (Updated 2nd July 2012)]
Other resources
File:Concentration calculator for lazy scientists.xls
[http://meta.wikimedia.org/wiki/Help:Contents Media wiki editing]
[http://blast.ncbi.nlm.nih.gov/Blast.cgi BLAST at NCBI]
[http://www.finnzymes.fi/tm_determination.html finnzymes Tm calculator]
[http://mfold.rna.albany.edu/?q=mfold mfold, DNA or RNA folding forms on the left hand side]