Harrison Lab Protocols
These protocols are available on the lab website for general use. They are all tried and tested by me, so please use them in preference to other protocols that you have so that they can be easily trouble-shooted. If you have your own protocols that are simple and work well, please give me an electronic copy so that they can be included in the lab protocols.
Chemical Safety
Please check the safety advice on chemical bottles if you are using chemicals for the first time. Most of the chemicals in use are non toxic or irritant, and can be used without gloves in solution. Wear gloves and face mask for weighing out irritant and toxic chemicals. Please be particularly careful weighing out antibiotics, SDS, and other toxins, and wipe up all spills with water.
Phenol
Clean up phenol spills with PEG 300 and wash off with plenty of cold water.
DO NOT USE HOT WATER
Do not close tubes with liquid nitrogen ground tissue until the tissue is fully thawed
Ethidium bromide
Use lab operating procedures
Bacteria
Clean up spills with 70% Ethanol and 10% bleach
Cloning
Agrobacterium transformation;
Dephosphorylation of DNA;
Filling in ends of DNA;
Heat Shock Transform. E. coli;
HS competent cells;
Ligation of DNA.doc;
Preparation of Agrobacterium Electrocompetent cells
DNA
DNA Minipreps;
Moss DNA Extraction;
Random priming DNA;
Southern blotting;
Southern hybridizations;
A-tailing rpotocol
Histology
GUS staining.pdf;
Dexamethasone induction.pdf;
Embedding in plastic;
Immunolocalization (AP);
In situ protocol;
Moss fixation;
Preparation of fixative
Moss
Agrobacterium moss transformation;
Culturing moss;
Moss media;
PEG mediated moss transformation;
Protoplast Isolation;
Sporophyte induction;
Working with spores
PCR
3' RACE;
Clontech genome walking;
Colony PCR;
Degenerate primer design;
iPCR (Jill);
LA PCR (5' RACE);
PCR Genome walking;
Pfu PCR reactions;
RT-PCR.doc
RNA
Formaldehyde Gels (RNA);
Northern blotting;
Northern hybridizations;
RNA extraction (TRIzol)
Arabidopsis
Sterilization Arabidopsis seed