Oneill Lab:Chemicals
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Michael J. O'Neill Lab University of Connecticut Department of Molecular and Cell Biology
Common Lab Chemical Recipes
The recipes listed here can be found in the grey protocol binder
Phosphate Buffered Saline
- Add 1 tablet per 100 ml diH2O
- Autoclave
20x SSC
Recipe to make 4L
- Dissolve 701.28g NaCl and 352.92g NaCitrate
- pH to 7.0
- Bring final volume to 4L
- Autoclave if necessary
20x TBE-modified
Recipe makes 2L
- 649.096g Tris
- 185.24g Boric Acid
- 37.968g EDTA
- Bring final volume to 2L
50x TAE
- 242g Tris
- 37.2g EDTA
- 57.1mL Glacial Acetic Acid
- Bring final volume to 1L with diH2O
20% SDS
- 200g SDS / 1L
- Wear mask and safety goggles!
- Heat to 68°C to dissolve
0.5M EDTA
- Dissolve 186.1g EDTA / 1L
- pH to 8.0
- May take several mL of 10N NaOH to adjust pH
- EDTA will not all go into solution until pH is adjusted
- Autoclave if necessary
1M Tris
- Dissolve 60.55g Tris / 500 mL
- Adjust pH as desired (7.5-8.0 typically)
1M HCl
- Add 8.3mL 12M HCl / 100 mL
- Always add acid to water
1M NaOH
- Add 2g NaOH / 50 mL
- NaOH cannot be stored in glass containers
0.4M NaOH
- Add 32g NaOH / 2L
- NaOH cannot be stored in glass containers
Loading Dyes
6x OrangeG
- 50mg OrangeG
- 1.5g Ficoll
- Add MilliQ water to 10 mL
- Aliquot and store at 4°C
10x BlueJuice
- 3.25g Sucrose (65% Final)
- 50μL 1M Tris (10mM Final)
- 100μL 0.5M EDTA (10mM Final)
- 15mg bromophenol blue
- 15mg Xylene cyanol
- Bring volume to 5 mL with MilliQ water
- Aliquot and store at room temperature
5x RedJuice
RedJuice can be safely added to PCR reactions
Contents
- 60% Sucrose
- 1mM Cresol Red (FW= 404.4)
Preparation
- 12g sucrose
- Add MilliQ to 20mL and mix
- Pass through 0.45μm filter to sterilize
- Add 8.09 mg cresol red
- Aliquot and store at 4°C
RNA loading dye
- 50μL glycerol
- 25μL 1% bromophenol blue
- 25μL 1% xylene cyanol
- 5μL 0.5M EDTA
- 450μL formamide
- 160μL 37% formaldehyde
- 45μL 20x Running Buffer
PCR reagents
10x PCR Buffer
Contents
100mM Tris pH 8.4 500mM KCl 15mM MgCl2
Preparation
Makes 25mL
- 2.5mL 1M Tris
- 375μL 1M MgCl2
- 12.5mL 1M KCl
- 9.625mL diH2O
Aliquot and store -20°C
10mM dNTP Mix
Preparation
- 10μL dATP
- 10μL dCTP
- 10μL dTTP
- 10μL dGTP
- 60μL MilliQ H2O
- Vortex briefly
- Aliquot and store -20°C
Solutions for Southern Blots
Depurination Solution
Contents
1L 0.25M HCl
Preparation
- Dilute 22mL 12M HCl in 978 mL diH2O
- Label and store at room temperature
Denaturation Solution
Contents
0.5M NaOH 1.5M NaCl
Preparation
- 87.66g NaCl
- 20.00g NaOH
- Bring final volume to 1L
- Store in a labelled plastic bottle
- Should be made fresh
Neutralization Solution
Contents
1.5M NaCl 0.5M Tris 2mM EDTA pH 8.0
Preparation
- 87.66g NaCl
- 60.56g Tris
- 4mL EDTA pH 8.0
- 800 mL diH2O
- Adjust pH to 7.2
- Will take about 34 mL 12M HCl
- Bring volume to 1L with diH2O
Church's Solution
Materials
- 20% SDS
- 0.5M EDTA pH=8.0
- 1M Na2HPO4
- diH2O
Procedure
- Mix 35ml SDS and 0.2ml EDTA
- Add 50ml Na2HPO4
- Bring final volume to 100ml
Note: SDS will precipitate at low temperatures. We have found keeping Church's Solution at 37°C may help in limiting instances of blot disease