Galloway Lab Protocols
ehs_safety
  • General Lab Maintenance
    • Replacing microcentrifuge motor mounts
      • Required materials
      • Procedure
    • Cleaning and autoclaving guidelines
      • Washing glassware and tools
        • Glass bottles and flasks
        • Glass beads
        • Dissection Tools
      • Autoclaving
        • Tip boxes
        • Tube containers
        • Glass pipette containers
        • Toothpicks
        • Flasks
        • Dissection tools
        • Glass bottles (containing liquid or glass beads)
    • Cleaning BSCs
    • Cleaning microcentrifuges
    • Checking data backups
      • Onedrive backups
        • Once a year checks
      • NGS backups
        • Adding a new NGS dataset checklist
        • Once a year checklist
    • Incubator decontamination
      • Maui and Moana (Main TC)
      • Hei Hei and Pua (Main TC)
      • Te Kā (Quarantine) and Te Fiti (66-219)
    • Replacing the logged-in OneDrive user
    • Ordering
      • Direct ordering through Coupa
      • Requests through Quartzy
      • Bulk Plastics Order
    • Recycling
  • Instrument Use and Core Facility Access
    • Attune operation
      • Attune startup and shutdown
        • Quick startup checklist
        • Quick shutdown checklist
        • Initial inspection and startup
        • Performance test
        • Shutdown
      • Attune operation
        • Procedure descriptions
        • Weekly system cleaning
        • Every ~3-6 month system decontamination
        • Focusing fluid refill
        • Full Attune guides
      • User-replaceable parts
        • Main syringe
        • Sample Injection Probe (SIP)
        • Focusing fluid filters
      • Attune troubleshooting
        • Delay before events start running
        • Fluid leak from the autosampler SIP onto a running plate
        • Wells skipping and the autosampler fails to draw focusing fluid
        • Power cycling the Attune
    • Koch Flow Core
    • Animal Facility
    • Keck Imaging Facility
    • Myco Testing
      • Getting Access
      • Submitting Testing Request in iLab
      • Submitting Samples for Testing
      • Results
    • MIT SuperCloud Access
    • Keyence beginner’s user guide
      • Basic Operation
      • Multi-point capture
      • Z-stack
      • Stitching
    • Keyence Time Lapse Imaging
      • Setting up the incubation chamber
      • Using the Keyence Software
      • Tips and Tricks
  • Protocols
    • Biochemical and analytical protocols
      • Homemade EU click
        • Stock solutions
        • Add EU to cells
        • Dissociate
        • Fix cells
        • Permeabilize
        • Prepare click/label mix
        • Label and wash
      • HIVE scRNA-seq
        • Sample capture
        • HIVE processing
      • Immunofluorescent Staining
        • Solutions Required
        • Adherent Cell Staining
        • Cell Staining for Flow
      • ATAC-see
        • Transposase buffer preparation
        • ATAC-see
      • CellBaum User’s Guide
        • Local Usage
        • MIT Supercloud Usage
        • Data Analysis
      • CFSE Labeling
        • CellTrace dyes available in lab
        • Other materials needed:
      • HCR DNA-FISH
        • Fixation
        • Hybridization
        • Probe Preperation
        • Post-Hybridization
      • Halo-tag Labeling
        • Halo-tag cell permeable dyes available in lab
        • Other materials needed:
      • HCR RNA-FISH
        • Solution Detection Stage
        • Solution Amplification Stage
      • Live nuclear Staining using Hoechst
        • Solutions Required
        • Live Hoechst staining
      • Image segmentation and particle analysis in ImageJ
        • Defining regions of interest (ROI)
        • Quantifying nuclear-localized foci
        • Merging images in ImageJ
      • KAPA SYBR® FAST qPCR
        • Protocol
        • 1. Master Mix Preparation
        • 2. Reaction Setup
        • 3. qPCR
        • 4. Data Analysis
      • Magnetic bead clean-up for genomics libraries
        • Uses
        • Determine Bead Volume to use by size range you wish to recover:
        • Preparations:
        • Basic Bead Clean Protocol:
        • 2-step “Big” bead clean to remove large fragments >~350bp.
      • Nuclei Isolation for Downstream Genomics Applications
        • Nuclear Isolation
        • Recipes
      • Psoralen-qPCR supercoiling assay
        • Experiment setup
      • sci-ATAC-seq
      • Single cell qPCR
        • Single-cell acquisition
        • Lysis, reverse transcription (RT), and specific target amplification (STA)
        • Single-cell qPCR on Fluidigm Biomark
      • SingleShot RNA isolation and Reverse Transcription
        • Materials need for RNA isolation
        • Materials needed for cDNA synthesis
        • RNA isolation using SingleShot Cell Lysis
      • Snap-tag Labeling
        • Snap-tag cell permeable dyes available in lab
        • Other materials needed:
      • Titration of Tn5 transposition in MEFs
        • Part 1. Assembling the Transposase
        • Part 1.1 Prepping the Tn5 concentrations from Assembled Tn5 transposome per Transposition reaction (50uL)
        • Part 2. Transposition Reaction of Isolated nuclei
      • Western Blot
        • Lysis
        • Bradford Assay
        • Protein Gel Casting
        • Loading and Running the Gel
        • Coomassie Staining
        • Transferring the protein from the gel to the membrane
        • Antibody Staining
    • Cloning protocols
      • Gibson assembly
        • Protocol
      • Glycerol Stocking
      • Golden Gate assembly
        • Protocol for PCR fragments
        • Protocol for pPV plasmids
        • Reference for pPV connector sequences
        • Reference for entire Golden Gate workflow
      • Ligation assembly
        • Protocol
      • LR cloning
        • Protocol
        • Expected results
      • Oligo annealing for ligation cloning
        • Oligo phosphorylation
        • Oligo annealing
        • Combined oligo phosphorylation and annealing
      • Touchdown PCR
        • Touchdown PCR
      • Bacterial Transformation
        • Cloning Summary
        • Transformation (Chemically Competent Bacteria)
    • in vitro transcription
      • Design and preparation of DNA IVT template
      • Creating a new IVT template plasmid
      • Generating IVT linear DNA template from an IVT template plasmid
      • modRNA synthesis
      • IVT reaction
      • mRNA transfection
      • MessengerMax transfection protocol
      • StemMACS transfection protocol
    • Protein production
      • Protein gel casting
        • Required stock solutions
        • Casting protocol
        • Gel casting setup
      • Chitin-binding protein purification
        • Required solutions
        • Protocol
        • Dialysis and concentration protocol
        • Regeneration Protocol
      • Denaturing protein gel run and staining
        • Solutions required
        • Running procedure
        • Staining (Coomassie)
      • Protein expression
        • Protocol
      • His-tag protein purification
        • Source
        • Required solutions
        • Protocol
    • Safety
      • Virus safety
        • Control methods
        • Virus protocols
        • Exposure plan
        • Lessons learned from incidents
    • TC protocols
      • TC Basics
        • TC Best Practices
        • Adherent Cell Culture
        • Mammalian cells cryopreservation and plating from frozen stock
        • Transient HEK293T Transfection
        • Flow Cytometry
        • Glass slide cell culture
        • Culturing Human iPSCs
        • Plat-E Retroviral Transduction of MEFs
      • Virus
        • Virus production in HEK293T
        • Transduction with concentrated virus
        • Plat-E Retroviral Production
        • VSV production
        • MOI Calculation
      • SNAP circuit (Transient expression in 96-well plate version)
        • Materials
        • Procedure
      • Cell Line Creation
        • Day -1
        • Day 0
        • Day 1
        • Day 2-4
        • Day 5 - Week 2
        • Week 2 + 1 day
        • Week 3
        • Clonal selection or enrichment via flow sorting
        • Repicking
        • Genotyping your line
      • Electroporation of plasmids into MEFs
        • Overexpression
        • Plasmid Concentration Experiment
      • Transgene expression control through ribozyme switch (96-well plate version)
        • Materials
        • Day 1: Seed cells
        • Day 2: Transfection
        • Day 3 or 4
  • Recipes
    • Bacterial recipes
      • LB (Amp/Kan) plate
      • Antibiotic Stock Recipes
        • 1000X Ampicillin (AMP)
        • 1000X Kanamycin (KAN)
        • 1000X Chloramphenicol (Chlor)
      • Competent cells (NEB stable)
        • Day 1. Culture NEB stable cells for overnight.
        • Day 2. Grow the competent cells and stock the cells.
        • Day 4. Confirm the cell growth
      • DNA wash buffer
        • DNA wash stock solution (5x)
        • DNA wash buffer
      • Bacterial media recipes
        • TB (Terrific Broth)
        • 10x TB salts
        • SOC Media
        • LB Media
      • Other Solutions
        • Glycerol (60%)
        • 1xTAE buffer
    • FISH recipes
      • FISH buffer recipes
        • Probe Hybridization Buffer
        • Probe Wash Buffer
        • Probe Amplification Buffer
        • 5X SSCT
    • TC recipes
      • FBS heat inactivation
      • Common aliquot sizes for TC
        • 1,000X Neurotrophic factor stocks
        • RepSox
        • Fetal Bovine Serum (FBS) Aliquots
        • 100X Pen-strep aliquots
        • 50X Geltrex aliquots
      • TC media recipes
        • N3 media
        • MEF media
        • Breast cancer 1 media
        • Breast cancer 2 media
        • Breast cancer 3 media
        • Breast cancer 4 media
        • Epithelial cell 1 media
        • Epithelial cell 2 media
        • Glia media
        • Sorting/Collection media
        • Motor neuron dissociation media
        • Freezing media
        • HEPES-buffered DMEM
      • Other TC Solutions
        • Phosphate buffered saline (PBS)
        • 0.1% Gelatin
      • PEI Stocks
        • Materials
        • Dissolve and Aliquot PEI
        • Test PEI Ratio
        • Reference
      • Small molecule stocks for TC
        • Indole-3-acetic acid (Auxin, IAA)
        • Indole-3-acetamide (Auxin precursor, IAM)
        • 50mM O6-Benzylguanine (O6BG)
        • 25mM Guanine
  • Plot Gallery
    • High-quality PDF figure export
      • Intro
      • Problematic features
        • Transparency/opacity
        • Gradients/blurs
      • Final figure export
        • Illustrator
        • Matplotlib/Python
      • Preflight
    • Flow plots with adjunct histograms
    • Histograms
      • Function to make subplots for several variables
    • Mixed distribution + mean plots
      • Gating plot
      • Box plot with well means
      • Violin plot with well means
    • Multiple-axes plots
    • Testing matplotlib directives
      • Subheading
    • Notes on reproducing examples
  • Writing Guide
    • Additional resources
      • Big picture resources
      • Getting into the finer details
    • Points of style
      • Specific and nonspecific subjects
      • Hyphenation
      • Numbers in text
      • How to write gene and protein names?
      • References
    • Guiding principles
      • Tips
      • Outline
      • Active Voice
      • Editing and revising
    • What is a paper?
  • Bootcamp
    • IAP Bootcamp
      • Data analysis with Python
        • Why do data analysis with Python?
        • Introduction: the research problem
        • Good file layout practices
        • Oh yaml, my yaml
        • Loading flow data
        • Introduction to Pandas
        • Plotting with Seaborn
      • Day 0: Software and training setup
        • EHS setup and trainings
        • Software and webservices
      • Computational environment check
        • Initial environment check
        • Protocols check
        • Completion date
      • An introduction to Git
        • Acknowledgements
        • Motivation
        • Git by example
        • Basic Git terms
        • Downloading or starting a repository
        • Basic history terms and git status: What’s happening?
        • Branching out, adding, and checking out files
        • Snapshotting your work
        • Ignoring files
        • Recovering history: reverting commits and file-level checkouts
        • Working with others: remotes and inevitable merge conflicts
        • Conclusion
      • Software Tips and Tricks
        • One date format to rule them all/naming
        • Paper RSS feeds
        • Creating feeds from searches
        • Zotero
        • Better Quartzy
        • Regex help
        • Fonts
        • Activate SnapGene remotely
      • Startup checklist when working with repositories
        • New repository (Python)
        • New repository (Julia)
        • New repository (R)
        • Existing, non-data-driven repository (e.g. protocols)
        • Existing, data-driven repository (e.g. tangles_model)
      • On Terminals and Shells
        • Motivation
        • The basics
        • Interface basics
        • Starting off at home
        • Moving away from home
        • Relative and absolute paths
        • File operations
        • Editing and viewing files
        • Finding things in documents
        • Exercise
        • Extras
    • Gene expression modeling with ODEs
      • Tutorial 1: Modeling gene expression with differential equations
      • Tutorial 2: Examining parameters affecting our models
      • Tutorial 3: Michaelis-Menten Kinetics
      • Tutorial 4: Modeling repressors
      • Tutorial 5: Hill functions
    • “How to” Training Series
      • Tutorial 1: How to Write a Proposal
      • Tutorial 2: How to Read a Paper
      • Tutorial 3: How to Review a Paper
      • Tutorial 4: Graphic Figure Design
      • Tutorial 5: How to Compose Figures
  • Tech documentation
    • Repository setup
  • Contributor guide
    • Local environment setup
      • Text editor
      • git
      • Python/Sphinx setup
    • Standard workflow
    • Local building
      • Local previewing
      • In case of build errors
    • Online editing through Github
    • Repository layout
    • Basics of reStructuredText
      • Simple markup
      • Explicit markup
      • Admonitions
      • Code
      • Tables
      • References and links
      • Images
      • Math
Galloway Lab Protocols
  • General Lab Maintenance
  • View page source

General Lab Maintenance

  • Replacing microcentrifuge motor mounts
    • Required materials
    • Procedure
  • Cleaning and autoclaving guidelines
    • Washing glassware and tools
    • Autoclaving
  • Cleaning BSCs
  • Cleaning microcentrifuges
  • Checking data backups
    • Onedrive backups
    • NGS backups
  • Incubator decontamination
    • Maui and Moana (Main TC)
    • Hei Hei and Pua (Main TC)
    • Te Kā (Quarantine) and Te Fiti (66-219)
  • Replacing the logged-in OneDrive user
  • Ordering
    • Direct ordering through Coupa
    • Requests through Quartzy
    • Bulk Plastics Order
  • Recycling

© Copyright 2024, Galloway Lab. Shared under the Creative Commons 4.0 Attribution International license. Last updated on May 10, 2021.

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