BISC 219/F10:Media Recipes

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Revision as of 10:59, 10 November 2010 by Tucker Crum (talk | contribs) (Ingredients and Stock Concentrations for some of the reagents used in Series 3)
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Ingredients and Stock Concentrations for some of the reagents used in Series 3

Reagent Composition
Solution 1* 50 mM glucose
10 mM EDTA (Ethylenediaminetetraacetic acid)
25 mM Tris, pH 8
Solution 2* 0.2 M NaOH
1% SDS (sodium dodecyl sulfate)
Solution 3* 2.7M Potassium Acetate
6.6M Acetic Acid
Luria-Bertoni Medium (LB) 1% Tryptone
0.5% Yeast Extract
1% NaCl
(2% agar in plates)
Nematode Growth Medium Lite(NGM lite)- Available in powder form from US Biological Cat. #N1005 ( 0.3% NaCl
1.7% Bacto-agar
0.25% Bacto-peptone
0.1% (vol/vol)cholesterol
Ampicillin* Stock = 50 mg/ml in water
use 1:1000 in liquid cultures
1:500 in agar
Tetracycline* Stock = 12.5mg/ml in 50% EtOH
use 1:1000 in liquid
1:1000 in agar
IPTG Stock = 0.5 M
use 0.5 mM in cultures
1% agarose gel 1% (w/v) agarose in 1X TAE buffer. (50X TAE Stock Solution= 242 g/L Tris Base (MW=121.1); 57.1 mL/L Glacial Acetic Acid; 100 mL/L 0.5 M EDTA; Add ddH2O to 1 Liter and adjust pH to 8.5 using KOH)
loading dye 0.4 % Ficoll 400, 1.8 mM EDTA, 0.55 mM Tris-HCl, 0.00117 % SDS, 0.025 % Bromophenol Blue pH 8.0 @ 25°C
1x PBS (phosphate buffered saline) 137 mM NaCl,2.7 mM KCl,4.3 mM Na2HPO4,1.47 mM KH2PO4,Adjust to a final pH of 7.4.

*NOTE: These concentrations are STOCK concentrations - not working concentrations!

Links to Labs& Project Info

Worm Info
Lab 1: Worm Boot Camp & Sex-Linked or Autosomal Start
Lab 2: Sex-Linked or Autosomal Finale
Background: Classical Forward Genetics and Gene Mapping
Lab 2: Mutant Hunt
Lab 3: Linkage Test Part 1
Lab 4: Linkage Test Part 2, Mapping and Complementation
Lab 5: Finish Complementation; Mapping Con't
Lab 6: DNA sequence analysis; Mapping Con't
Lab 7: Complete Mapping: Score
Schedule of Reverse Genetics Project
RNAi General Information
Media Recipes
Lab 5: Picking your gene to RNAi
Lab 6: Cloning your gene of interest
Lab 7: Picking your transformant
Lab 8: Plasmid purification and transformation
Lab 9: Induction of bacteria for RNAi
Lab 10: Scoring your worms and RNA purification

Lab 11: RT PCR reactions