BISC 219/F10:Media Recipes

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(Plasmid Isolation and Transformation Solutions)
Current revision (09:56, 18 May 2011) (view source)
 
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== Plasmid Isolation and Transformation Solutions ==
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== Ingredients and Stock Concentrations for some of the reagents used in Series 3 ==
{| border="1"
{| border="1"
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! Solution !! Composition   
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! Reagent !! Composition   
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! Solution 1*   
! Solution 1*   
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! Luria-Bertoni Medium (LB)
! Luria-Bertoni Medium (LB)
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| 1% Tryptone <br> 0.5% Yeast Extract <br> 1% NaCl <br> 2% agar in plates
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| 1% Tryptone <br> 0.5% Yeast Extract <br> 1% NaCl <br> (2% agar in plates)
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|-
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! Nematode Growth Medium Lite(NGM lite)- Available in powder form from US Biological Cat. #N1005 (http://www.usbio.net)
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| 0.3% NaCl<br> 1.7% Bacto-agar <br> 0.25% Bacto-peptone <br> 0.1% (vol/vol)cholesterol
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! Ampicillin*
! Ampicillin*
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!IPTG
!IPTG
| Stock = 0.5 M <br> use 0.5 mM in cultures
| Stock = 0.5 M <br> use 0.5 mM in cultures
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|-
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! 1% agarose gel
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|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)
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|-
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! loading dye
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|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
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|-
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! 1x PBST (phosphate buffered saline with Tween20)
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|137 mM NaCl,2.7 mM KCl,4.3 mM Na2HPO4,1.47 mM KH2PO4, 0.25% -Adjust to a final pH of 7.4.
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|}
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'''*NOTE: These concentrations are STOCK concentrations - not working concentrations!'''
'''*NOTE: These concentrations are STOCK concentrations - not working concentrations!'''
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==Links to Labs& Project Info==
==Links to Labs& Project Info==
Series1:<BR>
Series1:<BR>
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[[BISC 219/F10: RNAi Lab 10 | Lab 10: Scoring your worms and RNA purification]]<br>
[[BISC 219/F10: RNAi Lab 10 | Lab 10: Scoring your worms and RNA purification]]<br>
[[BISC 219/F10: RNAi Lab 11 | Lab 11: RT PCR reactions]]<br><br>
[[BISC 219/F10: RNAi Lab 11 | Lab 11: RT PCR reactions]]<br><br>
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</div>

Current revision

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 (http://www.usbio.net) 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 PBST (phosphate buffered saline with Tween20) 137 mM NaCl,2.7 mM KCl,4.3 mM Na2HPO4,1.47 mM KH2PO4, 0.25% -Adjust to a final pH of 7.4.

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

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