Haynes:Protocols: Difference between revisions
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=Protocols= | =Protocols= | ||
==DNA Assembly== | |||
* [[Haynes:BioBrick Method short | Assembly of BioBrick Parts: an Overview]] | * [[Haynes:BioBrick Method short | Assembly of BioBrick Parts: an Overview]] | ||
* [[Haynes:Making BioBricks | Making Standardized DNA Parts]] | * [[Haynes:Making BioBricks | Making Standardized DNA Parts]] | ||
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==Cell Culture: Bacteria== | |||
* [[Haynes:ChemComp cells | Chemically competent cell prep]] | * [[Haynes:ChemComp cells | Chemically competent cell prep]] | ||
* [[Haynes:Glycerol Stocks | Glycerol Stocks]] - for long term -80°C storage | * [[Haynes:Glycerol Stocks | Glycerol Stocks]] - for long term -80°C storage | ||
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==Cell Culture: Mammalian== | |||
* [[Haynes:MamCultureMedia | Media formulas]] - cell line-specific formulas | * [[Haynes:MamCultureMedia | Media formulas]] - cell line-specific formulas | ||
==Protein== | |||
* [[Haynes:Bradford | Bradford assay]] - measure protein concentration | * [[Haynes:Bradford | Bradford assay]] - measure protein concentration | ||
==Real Time Quantitative PCR== | |||
* [[Haynes:UPLassay | Universal Probe Library (UPL) assay]]: for the Roche Light Cycler 480 | * [[Haynes:UPLassay | Universal Probe Library (UPL) assay]]: for the Roche Light Cycler 480 | ||
==Resources at OpenWetWare== | |||
* [http://openwetware.org/wiki/E._coli_genotypes E. coli Strains] | * [http://openwetware.org/wiki/E._coli_genotypes E. coli Strains] | ||
* [http://openwetware.org/wiki/Materials Materials]: buffer formulas; specific information on commonly used reagents (''e.g.'', LB broth, ampicillin, restriction enzymes, etc.) | * [http://openwetware.org/wiki/Materials Materials]: buffer formulas; specific information on commonly used reagents (''e.g.'', LB broth, ampicillin, restriction enzymes, etc.) | ||
==Software Guides== | |||
* [[Haynes:Jmol_Guide | Jmol commands: controlling PDB structure displays]] | * [[Haynes:Jmol_Guide | Jmol commands: controlling PDB structure displays]] | ||
Revision as of 16:36, 27 October 2012
Protocols
DNA Assembly
- Assembly of BioBrick Parts: an Overview
- Making Standardized DNA Parts
- Model Procedure for Assembling Parts: Classic Ligation for Beginners
- Gibson Assembly
Cell Culture: Bacteria
- Chemically competent cell prep
- Glycerol Stocks - for long term -80°C storage
- Transformation of E. coli with plasmid DNA
Cell Culture: Mammalian
- Media formulas - cell line-specific formulas
Protein
- Bradford assay - measure protein concentration
Real Time Quantitative PCR
- Universal Probe Library (UPL) assay: for the Roche Light Cycler 480
Resources at OpenWetWare
- E. coli Strains
- Materials: buffer formulas; specific information on commonly used reagents (e.g., LB broth, ampicillin, restriction enzymes, etc.)
Software Guides
Recipes
Click "show" to expand each recipe, and "hide" to, well, hide it.
Bromo-Blue/X-cyanol Loading Buffer, 10X
Formula: [30% glycerol; 12.5 mg/mL bromophenol blue; 12.5 mg/mL xylene cyanol]
Volume: 20 mL
- 50% glycerol, 12 mL
- Bromophenol blue, 250 mg
- Xylene cyanol, 250 mg
Bring up to total volume with dH2O.
Chromatin Prep Buffer A
Formula: [10 mM HEPES (pH 7.9); 10 mM KCl, 1.5 mM MgCl2; 0.34 M sucrose; 10% glycerol; 1 mM dithiothreitol; 1x protease inhibitor cocktail]
Volume: 100 mL
- 1 M HEPES pH 7.9, 1 mL
- 1 M KCl, 1 mL
- 1 M MgCl2, 150 μL
- Sucrose, 11.6 g
- 50% Glycerol, 20 mL
- 1 M Dithiotreitol (DTT), 100 μL
Bring up to total volume with dH2O. Store at 4°C. Add 100x protease inhibitor cocktail* (1:100) immediately before use.
LB Broth (Lennox)
Volume: 1 L
- Caesin tryptone, 10 g
- Yeast extract, 5 g
- NaCl, 5 g
- 1 N NaOH, 1 mL
Dissolve in 1000 mL dH2O. Autoclave to sterilize. Cool to room temperature before adding antibiotics. Shortcut: Use 25 g Acros LB Broth, Lennox (BP9722-500) granules instead of caesin tryptone, yeast extract, and NaCl.
Mammalian Cell Media
Volume: 500 mL
- Appropriate incomplete medium, 500 mL
- Fetal bovine serum (FBS), 50 mL
- Penicillin/ streptomycin (pen-strep), 5 mL
- Appropriate antibiotics, depending upon formula
Add FBS and pen-strep directly to the incomplete medium in the original stock bottle. Cap and invert to mix. Filter sterilize using a vacuum filtration unit.
