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Lab Floor Plan (with list of materials)
Detailed Lab Task Descriptions
General microbiology protocols
Media Recipes
Reagent Recipes
Working with Antibiotics
Freezing -80 Stocks
Freezing Aliquots
Competition Assays
Generic PCR
Gradient PCR
Running DNA Gels
Running SDS-PAGE Gels
Western Blot
Protein Purification
Protein Sample Concentration
Fixing Cells for Microscope/Flow Cytometry Work
Cloning and gene manipulation
Commonly Used Plasmids
Plasmid Purification
Digest and Ligation
Gel Purification
Creating Competent E. coli Cells
Transformation
Transformation — non-competent E. coli
Gibson Assembly
SOE PCR (Splicing by Overlap-Extension)
Qubit dsDNA Broad Range Assay
Preparing Sanger Sequencing (Eurofins)
Preparing Plasmid Sequencing (plasmidsaurus)
Creating Lac- E. coli Mutants
Streptococcus pneumoniae protocols
Dual Layer Assays
Streptococcus DNA Extraction - Genome Prep
Streptococcus CRISPR-Cas9 Editing
Streptococcus Transformation
Streptococcus Growth Curve Protocol
Streptococcus Growth Curve and Cell Count in Liquid Media
Log Phase Growth Curve and Cell Count in Liquid Media
Streptococcus Bacteriocin (Dual Layer) Assays - Original
Streptococcus Bacteriocin (Dual Layer) Assays - Early Producer
Streptococcus Bacteriocin (Dual Layer) Assays - Light and Normal Target Lawns
Streptococcus Bacteriocin (Dual Layer) Assays - Finding Producer-Resistant Target Bacteria
Streptococcus Bacteriocin (Dual Layer) Assays - Finding Producer-Resistant Target Bacteria (6-well plate version)
Streptococcus mutans protocols
Streptococcus mutans Growth
Streptococcus mutans Transformation
Myxococcus xanthus protocols
Media Protocols
Culture Cells from a Frozen Stock
Making a Broth Culture from an Agar Plate
Generating Frozen Stocks of Strains
Measure Absorbance of M. xanthus Culture
Generate St Curve for OD600 to Cells/mL conversion
Development Assay on Agarose Plates
Rehydrating New Primers
PCR Amplification from Genome
Ligation of PCR product into TOPO 2.1 vector
Transform competent E. coli cells
Colony PCR to confirm correct insert
Plasmid Isolation with BioBasic Miniprep Kit
EcoRI digest of plasmid
Plate Colonies Using CTTSA
Electroporation and Plating of M. xanthus transformants
M. xanthus genomic DNA extraction with Zymo (yellow) kit
Image Analysis in Fiji
Prepping a Submerged Culture
Heat Fixing and Staining
Propidium Iodide Staining on Agar Plates
Phage protocols
Media and Passaging
Plaque Assays with Soft Agar
The soft agar should be prepared first, likely at least an hour before you pour
- Put a heating block in the hood; set to 42C and place sterile 13mm test tubes in it to warm up; use the number of plates expected, plus 1 test tube.
- Make sure the water bath has enough water and is warming to 42C
- You will use about 4 mL of soft agar per plate, so calculate total volume of agar needed accordingly
- Always prepare enough agar for at least one extra plate
- Loosen the cap of the soft agar bottle and microwave the bottle of agar until it is fully liquid:
- Check for bubbling about every 15 seconds
- You will know when it is fully melted because:
- There will be no more “cloudiness” or bits
- Shaking or swirling will cause bubbles to rise directly to the top(not being blocked by solid material)
- After microwaving, place bottle into the preheated water bath
- After preparing agar, use a serological pipet to transfer 4 mL of soft agar into each 13mm test tube. The top layer on plates will consist of an equally sized layer made of the same broth and soft agar (0.8% agar) mixed with 60 uL of bacterial culture.
- Warm to 37 degrees and label plates with circles and dilution numbers to make plating easier
- Phage dilution will depend on goals of the assay but normally it consists of the undiluted sample and a range from 10^-2 to 10^-4
- Prepare for pouring the phage layer
- Prepare all phage pipettes
- Prepare filtered tips for phage
- Check that your dilutions are represent and organized
- Check that you collected all your warmed plates from the incubator
- Make sure your vortexer is in the hood, plugged in and the outlet is switched on
- Quickly but carefully (1 by 1):
- Pipette 60 uL of overnight bacterial culture into 13mm test tube
- Vortex for about 5 seconds (look for the tornado)
- Open pre-labeled plate and pour mixture onto the LB layer
- Swirl gently to allow soft agar to spread evenly, cover, then set aside
- Allow about 15 seconds for a poured plate to set before moving
- Plate 5 μl of each phage sample and dilution according to previously made circles
- After adding phage we need to allow liquid to dry since it is on top of the solidified soft agar. While it would dry faster if the plate was uncovered, we do not want the phage to contaminate the hood in any way, so let liquid dry with the lid on the plate.
- Allow to dry for about 1 hour to ensure no drying issues
- Place plates in 37C incubator and check the next day.
Serial Dilutions of Phage
Calculating Virus Titre
Measuring Burst Size
Interactions Protocols
Zone of Inhibition Assay
Remote Molecular Biology
Effect of Laboratory Protocols on Student Learning
Interesting Podcasts to Listen to When Doing Lab Work!
- This Week in Microbiology
- By Vincent Racaniello
- 5 stars! Amazing podcast to learn all about different types of microbiology research! This podcast goes pretty in depth into different current scientific papers so it is a great way to learn about current research.
- This Week in Virology
- By Vincent Racaniello
- Ologies
- By Alie Ward
- 5 stars! Great all around podcast made for a more general audience. There are many different episodes on scientific topics including Environmental Microbiology, Mathematical Biology, and much much more. This is a great podcast to explore different careers and listen to some amazing speakers.
- Overheard at National Geographic
- By National Geographic
- 4 stars! Great podcast to hear about the many wonders of the world. It also has shorter episodes and is a great way to learn about the world.
- Journey to the Micro Cosmos
- 3 stars! Short 10 minute episodes about cool microbes.
Waste Disposal
Streptococcus suis protocols
Streptococcus suis Transformation
Measuring Absorbance in Streptococcus
Streptococcus DNA Extraction
Streptococcus Competence Induction
Peptide Synthesis
Peptide Cleavage
Mass Spectrometery
Plate Reader Assay and Growth Curve
Measuring Competence : Fixation and Flow Cytometry
Arabidopsis thaliana protocols
Creating Sterile Agar Plates
Sterile Seeding Protocol
Germination Protocol for ''Arabidopsis thaliana'' Seeds in Non-Sterile Experiments
Growth Stage Phenotype Definitions
Growth Conditions for ''Arabidopsis thaliana''
Measuring Light with HOBO Data Loggers
Inoculation of ''Arabidopsis thaliana'' with Microbes
Removal and DNA Extraction of Phyllosphere Microbes
ARISA
Measuring ''A. thaliana'' Phenotype using FIJI by Hand
DNeasy PowerSoil Protocol
Fiji Measurement
Making Boxes
Growing ''A. thaliana'' for Seed Harvest
Growing ''A. thaliana'' in Cut Pipet Tips
Cambridge protocols
Storage buffer
transformation of R5(2)-mCh-FL-BST and
expression
lysis and immobilization
Bio320 Microbe Species Wikipedia Pages
Getting started with MediaWiki
Consult the User's Guide for information on using the wiki software.