E. coli GFP transformation: Difference between revisions

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= Genetic modification =


We're interested in doing genetic modification on micro-organisms at the London Hackspace. Needed for GFP experiment (that we don't already have):
Fridge?
Incubator.
Build or buy.
Cleaning:
Bleach, gloves, paper towels, hand cleaning gel.
Bunsen burner + fuel.
Glassware, pipettes, tools.
Ethanol.
Lots.
To apply for a license we will probably first need:
Written protocols for stuff, especially cleaning up.
Disposal.
Chemical.
Sterilisation.
Do we need incineration? Find a waste disposal company that will take biohazard material?
Restriction enzymes.
At least two. 
Pricey?
GFP plasmid.
Antibiotics.
Ampicillin.
Initial e. coli culture.
Growth medium/ broth.
LB.
Plates.
Standard 10cm plates.
Agarose.
Ice.
40C bath.
Proper storage.
Centrifuge
- bigger : hold
Miniprep kit
Maxiprep kit
== Growing competent E. coli ==
[http://wiki.london.hackspace.org.uk/w/images/a/af/DIYbio_Collaboration_protocols.pdf Here] is the protocol we used with UCL to grow competent cells during the making of the public biobrick. So we need to get E. coli, [http://www.sigmaaldrich.com/catalog/product/fluka/l3152?lang=en&region=GB LB] - £66, 0.1M CaCl2, M9 salts, MgSO4, [http://www.sigmaaldrich.com/catalog/product/sigma/l2025?lang=en&region=GB bacteriological agar solution] - £42.90, thiamine, D glucose and plates. All links are only the first ones I found, maybe cheaper options.
== Self-cloning ==
[http://www.ncbe.reading.ac.uk/ncbe/protocols/transformer.html Here's] a protocol to do a 'self-cloning' transformation, which we would not need a licence for, and so could do now to practise.
* plate spreaders
* petri dishes
* LB
* Agar
* Howard has a good protocol for transformation
== Legal ==
There are a number of things we have to do in order to set up a GM lab.
The relevant regulations are [http://www.legislation.gov.uk/uksi/2000/2831/pdfs/uksi_20002831_en.pdf The Genetically Modified Organisms (Contained Use) Regulations 2000] (PDF)
Here is a summary of those regulations: [[Project:Biohacking/Genetic_modification/Regulations_2000_summary|GMO Regulations 2000 summary]]

Latest revision as of 18:26, 2 April 2015