Difference between revisions of "Davidson Missouri W/Davidson Protocols"

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# [http://www.bio.davidson.edu/courses/Molbio/Protocols/clean_short.html Ethanol Precipitate DNA (short protocol)]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/clean_short.html Ethanol Precipitate DNA (short protocol)]
 
# [[glycerolstocks How to Make Glycerol Stocks of Bacteria]]
 
# [[glycerolstocks How to Make Glycerol Stocks of Bacteria]]
−
 
  
 
'''B. Gel Electrophoresis and Purification'''
 
'''B. Gel Electrophoresis and Purification'''
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# [http://www.bio.davidson.edu/courses/Molbio/Protocols/molwt.html Calculate MWs]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/molwt.html Calculate MWs]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/gels2002/1kbladder.pdf 1kb MW markers]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/gels2002/1kbladder.pdf 1kb MW markers]
 +
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/MN_gelpure.html Macherey-Nagel Gel Purification (improved 260/230 ratios)l]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/Qiagen_gelpure.html Qiagen QIAquick Gel Purification]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/Qiagen_gelpure.html Qiagen QIAquick Gel Purification]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/QIAQuick_recycle.html Qiagen QIAquick Column Regeneration Protocol]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/QIAQuick_recycle.html Qiagen QIAquick Column Regeneration Protocol]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/gelpure.html ElectroElute Gel Purification]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/gelpure.html ElectroElute Gel Purification]
−
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/ligation.html Ligation Protocol]
 
  
 
'''C. Digestion, Ligation, Transformation'''
 
'''C. Digestion, Ligation, Transformation'''
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# [[Davidson Missouri W/Double Digest Guide| Double Digest Guide]]
 
# [[Davidson Missouri W/Double Digest Guide| Double Digest Guide]]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/SAP.html Shrimp Alkaline Phosphatase]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/SAP.html Shrimp Alkaline Phosphatase]
 +
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/ligation.html Ligation Protocol]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/Tranformation_list.html Choices for Transformation: Heat Shock vs. Zyppy]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/Tranformation_list.html Choices for Transformation: Heat Shock vs. Zyppy]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/Promegacompcells.pdf Heat Shock Transformation] OR [http://www.bio.davidson.edu/courses/Molbio/Protocols/transformation.html Short version of Heat Shock]
 
# [http://www.bio.davidson.edu/courses/Molbio/Protocols/Promegacompcells.pdf Heat Shock Transformation] OR [http://www.bio.davidson.edu/courses/Molbio/Protocols/transformation.html Short version of Heat Shock]

Revision as of 21:24, 13 March 2010

A. General Lab Information

  1. How to Keep a Lab Notebook
  2. Common molecular reagents
  3. Standard Assembly
  4. BioBrick Ends
  5. Compatibility of Plasmids
  6. Ethanol Precipitate DNA (short protocol)
  7. glycerolstocks How to Make Glycerol Stocks of Bacteria

B. Gel Electrophoresis and Purification

  1. Pouring an agarose gel
  2. Calculate MWs
  3. 1kb MW markers
  4. Macherey-Nagel Gel Purification (improved 260/230 ratios)l
  5. Qiagen QIAquick Gel Purification
  6. Qiagen QIAquick Column Regeneration Protocol
  7. ElectroElute Gel Purification

C. Digestion, Ligation, Transformation

  1. Digest DNA with restriction enzymes
  2. Double Digest Guide
  3. Shrimp Alkaline Phosphatase
  4. Ligation Protocol
  5. Choices for Transformation: Heat Shock vs. Zyppy
  6. Heat Shock Transformation OR Short version of Heat Shock
  7. Zippy Transformation
  8. TSS Competent Cell Transformation

D. Minipreps

  1. Choices for Mini-Preps: Promega vs. Zyppy
  2. Promega miniprep
  3. Zippy Miniprep

E. Making New Parts and PCR

  1. Building dsDNA with Oligos
  2. Setting up PCR mixtures
  3. PCR and Mg2+ concentration
  4. Making dsDNA Using Primer Dimers
  5. Clean and Concentrate DNA (after PCR, before digestion)
  6. Colony PCR to Screen for Successful Ligations

F. Expression of Phenotypes

  1. Using degradation tags on proteins such as GFP
  2. Genomic Insertion Protocol
  3. When inducing with IPTG, use 3 µL of stock (0.2 g/mL = 20% w/v) to every 1 mL of LB or other liquid.
  4. When inducing with Arabinose, use "2 µL of stock" (10% w/v L-Arabinose) "to every 1 mL" of LB or other liquid.

G. Computer Tools We Use

  1. All Sites In One Place
  2. Optimize your Gel
  3. Gene Splitting Web Site
  4. PCR Primers w/ BioBricks
  5. Promega Tm Calculator
  6. List of auto-inducers and their catalog numbers
  7. Sequencing at Agencourt Bioscience
  8. Sequencing at CUGI
  9. Analyzing Sequences with aPe
  10. Lance-olator Oligos for dsDNA assembly
  11. Access the GCAT-alog of Davidson and MWSU DNA Freezer Stocks
  12. Using Apes (A Plasmid Editor)
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