This page contains the internal lab protocols of the Cornell Food Safety Laboratory. These protocols are available for use to others, but there are no implied guarantees or assurances as to the validity of these protocols.
Below is a list of the current protocols for Cornell Food Safety. Below is a list of child pages where you can access PDF copies.
For any questions about these protocols, please contact Emily Wright at emw85@cornell.edu.
Current Cornell Food Safety Protocols & SOPS:
1-Animal Studies
1.1-Infection of Guinea Pigs with Listeria monocytogenes
2-Bacterial Growth & Preservation
2.1-Antibiotic Code System
2.2- Beckman Spectrophotometer DU-640 Protocol
2.3- Determining bacterial ODs using the Spectronic 20D+
2.4-Formulating media with growth inhibitors
2.4.1- Formulating media with growth inhibitors chart
2.5- FSL Isolate Designation, ID Assignment & Maintenance
2.6-Growth curve using the plate reader
2.7-Protocol for Freezing Bacterial Isolates
2.8-Spiral Plater & QCount
2.9-Standardized Growth Protocol for Bacterial Cultures & ODs
3-Bacteriophage
3.1- Listeriaphage isolation, purification, high-titer phage lysate preparation, and host range determination
3.2- Preparing High Titer Phage Stocks on M. smegmatis
3.3- Salmonella phage DNA extraction and PFGE
3.4- Salmonella Phage isolation, purification and host range characterization
3.5- Salmonella phage transduction
4- Commercial SOPs & LMT
4.1- actA PCR Assay to Differentiate Listeria monocytogenes
4.2- Generic Procedure for Automated Ribotyping Using the RiboPrinter
4.3- Molecular Subtyping of E.coli, Salmonella, and Shigella by Pulsed field Gel Electrophoresis (PFGE)
4.4- Molecular Subtyping of Listeria monocytogenes and Listeria species by Pulsed field Gel Electrophoresis (PFGE)
4.5- Procedure for Automated Ribotyping Using the RiboPrinter and Purified DNA Samples
4.6- Receiving External DNA
4.7-Receiving External Isolate
4.8- RiboPrinter Software Cheat Sheet
5-Lab Manuals & Safety SOPs
5.1-BSL1 &BSL2
5.2-Critical Laboratory Orientation
5.3-Exiting Student Checklist
5.4-Freezer emergency Plan
5.5- General safety procedures and handling and disposal of laboratory waste, including biohazard materials (BSL-1 and BSL-2 pathogens)
5.6- Laboratory Orientation Checklist
5.7-Offical Lab Manual
5.8-Poster Guidelines
5.9-Preparing Dangerous Goods for Shipping
5.10-Required and Suggested Readings
6- Microbial Detection & Isolation
6.1- Detection and Isolation of L. monocytogenes from Food and Environmental samples using the BAX system
6.2- Detection and Isolation of Listeria, Salmonella, Escherichia coli O157H7, non-O157 shiga toxin producing Ecoli (STEC) from Grazing Pastures, Pristine Environments an
6.3- Detection of Listeria monocytogenes from silage and other feeds
6.4- Detection of Listeria monocytogenes in milk
6.5- Detection of V parahaemolyticus O3K6 in oyster samples
6.6- Procedure to detect and isolate Listeria, Salmonella, Escherichia coli O157:H7, and non-O157 shiga toxin producing E. coli (STEC) from samples collected from pristine environments.
6.7- Surface swap detection for L. monocytogenes
7- Milk Quality
7.1- 2nd floor-16S GreenMaster Procedure
7.2- 16s Identification Using RDP database
7.3- 96 Well Plate Procedure
7.3.1- 96 well plate template
7.4- Checking Reports Protocol
7.5- Colony PCR
7.6- Dried milk — Enumeration of the specially thermoresistant spores of thermoresistant bacteria
7.7- Endospore Stain - cold
7.8- Endospore Staining -- Steam Method
7.9- Final rpoB GreenMaster Protocol
7.10- Growth Rate L.mono in Milk
7.11- Highly Heat Resistant Spore Pasteurization SOP
7.12- Identification of rpoB Sequences using Bioedit
7.13- Lab Pasteurization
7.14- PCR Procedure and Touble-shooting Guide for rpoB and 16S
7.15- Primer Ordering
7.16- Primer Reconstitution
7.17- Procedure for Lysis of Bacterial Cells
7.18- Purified Lysates Procedure
7.19- Spiral Plating for raw and powdered dairy products
7.20- Spore Count SOP
7.21- Spore Media Protocol
7.22- VSL Microbiological and Chemical Analysis
7.23-Sensory Analysis SOPs
7.23.1- Defective Milk, Reference Flavor & Basic Tastes Sample Preparation Protocol - in progress
7.23.2- MQIP VSL Sensory Analysis Panelist Training Protocol - in progress
7.23.3- Pouring instructions - computer set-up - short version
7.23.4- Sensory Analysis SOP
7.23.5- VSL Sensory Analysis Protocol - in progress
7.24-SOPs for USDEC
7.24.1- 16S rDNA PCR-Go Taq
7.24.2- HHR Spore Count SOP
7.24.3- Dried milk — Enumeration of the specially thermoresistant spores of thermoresistant bacteria
7.24.4- Spiral Plating for raw and powdered dairy products
7.24.5- Spore Count SOP
8-Molecular Biology Techniques
8.1-Characterization & DNA Amplification
8.1.1-PCR
8.1.1.1-General PCR Methods & Guidelines
8.1.1.1.1-16S rDNA PCR-Go Taq
8.1.1.1.2-16S rDNA PCR-modified for Taq gold
8.1.1.1.3-Basic GoTaq PCR Protocol
8.1.1.1.4-Colony PCR Protocol
8.1.1.1.5-Guidelines for Primer Design
8.1.1.1.6-Primer Naming, Reconstitution & Storage Protocol
8.1.1.1.7-Primer Reconstitution
8.1.1.1.8-Sample Submission to BRC
8.1.1.1.9-Ten Things That Can Kill Your PCR
8.1.1.1.10- Submitting sequences to GenBank
8.1.1.2-Listeria PCRs
8.1.1.2.1-Ten gene MLST Listeria
8.1.1.2.1.1-New MLST Scheme
8.1.1.2.1.2-addB PCR with GoTaq
8.1.1.2.1.3-ldh PCR with GoTaq
8.1.1.2.1.4--lmo490 PCR with GoTaq Hotstart
8.1.1.2.1.5-lmo1555 PCR-GoTaq
8.1.1.2.1.6-lmo2763 PCR-GoTaqhotstart
8.1.1.2.1.7-pbpA PCR-GoTaqhotstart
8.1.1.2.1.8-pbpA PCR-J1-208 GoTaqhotstart
8.1.1.2.1.9-polC PCR-GoTaq
8.1.1.2.1.10-prs PCR-GoTaq
8.1.1.2.1.11-rarA PCR-GoTaq
8.1.1.2.2- PCR by Gene Name
8.1.1.2.2.1- Fact Sheet on ActA Differentiation PCR & Interpretation of Results
8.1.1.2.2.2- Gap PCR protocol for Listeria monocytogenes
8.1.1.2.2.3- Hly ab gene PCR for Listeria monocytogenes
8.1.1.2.2.4- inlA ORF & Gene Fragment PCR & Sequencing
8.1.1.2.2.5- Modified SigB PCR Protocol-Taq Gold 10uM primers
8.1.1.2.2.6- Multiplex PCR for inlH screening
8.1.1.2.2.7- Nested hly PCR for Listeria monocytogenes
8.1.1.2.2.8- Protocol for sequencing actA gene most polymorphic region
8.1.1.2.2.9- Protocol for Sequencing actA gene region with best discrimination
8.1.1.2.2.10- Protocol for Sequencing inlA gene region with the best discrimination
8.1.1.2.2.11- Prs PCR Protocol for Listeria monocytogenes
8.1.1.2.2.12- PurM PCR Protocol for lineage II Listeria monocytogenes
8.1.1.2.2.13- PurM PCR Protocol for Listeria monocytogenes
8.1.1.2.2.14- RibC PCR Protocol for Listeria monocytogenes
8.1.1.2.2.15- SigB PCR Protocol GoTaq
8.1.1.3-Salmonella PCRs
8.1.1.3.1-fimA PCR & DNA sequencing of Salmonella enterica
8.1.1.3.2-manB PCR & DNA sequencing of Salmonella enterica
8.1.1.3.3-mdh PCR & DNA sequencing of Salmonella enterica
8.1.1.3.4-7 Gene MLST for Salmonella
8.1.1.3.4.1-7 gene MLST for Salmonella- Primers and PCR
8.1.1.3.4.2-Salmonella 7 gene MLST Template
8.1.1.3.5-PCR detection of the presence or absence of antibiotic resistance genes in Salmonella
8.1.1.3.6- InvA colony PCR for Salm 4-13-10
8.1.1.3.7-PCR identification of common Salmonella serogroups
8.1.1.3.8-PCR determination of H1 and H2-antigens for Salmonella
8.1.1.4-PCRs for Other Organisms
8.1.1.4.1- E. coli 6-gene multiplex PCR
8.1.1.4.2- ipaH colony PCR for Shigella
8.1.1.4.3-SodA PCR Protocol for S. agalactiae
8.1.2-Phenotypic Characterization
8.1.2.1-Restriction Digest and Cloning of PCR amplified 16S rDNA
8.1.2.2-Semi-Quantitative Hemolysis Assay for Listeria
8.1.3-RLFP
8.1.3.1-Polymerase Chain Reaction- Restriction Fragment Length Polymorphism
8.1.4-Southern Blot
8.1.4.1-Southern Blotting by Electrophoretic Transfer and non-radioactive Detection
8.1.4.2-Southern Blotting
8.1.4.3-SouthernBlot Hybridization
8.2-DNA Labeling & Modification
8.2.1-3' end labeling of oligonucleotides
8.2.2-Kinasing and DIG labeling of LCR primer
8.2.3-Poly-T tailing of oligonucleotides
8.2.4-Protocol for Fluorescein labeling of oligonucleotides carrying an aminolink
8.2.5-Protocol for Geneclean (USB)
8.3-DNA Preparation, Purification & Quantification
8.3.1-Chelex DNA extraction from blood
8.3.2-Chelex DNA extraction from semen
8.3.3-Detailed Preparation of Chromosomal Listeria DNA (5_05_2011)
8.3.4-DNA Preparation from Listeria using Guanidinium Isothiocyanate
8.3.5-Listeria Lysis Methods and Procedures
8.3.6-PCR product purification using ExoSAP
8.3.7-PCR Purification Protocol for DNA Sequencing
8.3.8- Preparation of chromosomal Listeria DNA according to Flamm et al.
8.3.9-Preparation of chromosomal Listeria DNA
8.3.10-Quantification of Purified DNA by NanoDrop
8.3.11-Single Stranded DNA preparation from M13
8.3.12-Small-scale phenolchloroform extraction of chromosomal DNA from Listeria monocytogenes
8.4-DNA Recombination
8.4.1-Cloning of DNA Flanking Tn917-LTV3
8.4.2-Cloning of DNA Fragments
8.4.3-Cloning PCR products for Sequencing using pCR 2.1
8.4.4- Cloning Protocol for Transformation of E. coli DH5aF
8.4.5- Electroporation L mono
8.4.6- Electroporation of E coli
8.4.7- Electroporation of Mycobacterium paratuberculosis
8.4.8- General Cloning DNA
8.4.9- GUS promoter fusions in Listeria monocytogenes
8.4.10- Integration of pPL2 derivatives into L. monocytogenes 10403S
8.4.11- Listeria Genomic Alterations
8.4.12- Overview-Making a mutant in Listeria monocytogenes
8.4.13- Part II-Making a mutant in Listeria monocytogenes
8.4.14- Part III-Making a mutant in Listeria monocytogenes
8.4.15- Part IV-Making a mutant in Listeria monocytogenes
8.4.16-pMJG2 vector map
8.4.17-pMJG3 vector map
8.4.18-pMJG4 vector map
8.4.19- Procedure for cloning with pDH32
8.4.20-Whole Mutant- SoeingPCR for L. mono
8.5-Microarrays
8.5.1-Analyzing Microarrays
8.5.2-Creating Microarrays
8.5.3-Labeling RNA with Cy3Cy5 and Hybridization for Microarrays
8.5.4-Microarray Slide Blocking
8.6-Protein Methods
8.6.1-Assay for beta-galactosidase activity of B. subtilis transformed with lacZ reporter plasmid
8.6.2-Beta Galactosidase Assay
8.6.3-His tagged sigma factor protein purification protocol
8.7-RNA Prep & Analysis
8.7.1-DNase treatment and phenolchloroform extractions for high purity RNA
8.7.2-Isolation of RNA from Listeria monocytogenes infected mammalian cells
8.7.3-Preparation of infected tissue culture cells for bacterial RNA collection
8.7.4-Preperation of Listeria RNA
8.7.5-RNA extraction, clean-up and storage using Turbo DNase for TaqMan
8.7.6-RNA Prep with RNeasy Midi Kit instructions
8.7.7-RNeasy kit total RNA isolation from Listeria monocytogenes with on-column DNase treatment
8.7.8-RT-PCR for the detection of cytokine mRNA
8.7.9-Synthesis of large amounts of RNA
8.7.10-Taqman- Quantitative Real-Time RT-PCR Protocol
9- Food Microbe Tracker
9.1- Food Microbe Tracker Batch Upload & Help Guide
10- Tissue Culture
10.1-Caco-2
10.1.1- Listeria monocytogenes Caco-2 cell invasion assay
10.1.2- Maintenance of Caco-2 Cells_22NOV10
10.1.3- Salmonella Caco-2 cell invasion assay
10.2-Hela
10.2.1- Culturing Conditions, Splitting and Freezing of Hela
10.2.2- Tissue culture assay to quantify bacterial adhesion to HeLa cells
10.3-J774
10.3.1- J774 cells Culturing Conditions, Splitting and Freezing
10.3.2- Label Listerial Protein J774 Cells
10.3.3- Labeling of listerial proteins in J774 cells
10.4-L2 Mouse
10.4.1- Listeria monocytogenes Intercellular Growth Assay in L2
10.4.2- Mouse L cells Culturing Conditions, Splitting and Freezing
10.4.3- Passage of Mouse L Cells for Plaque Assay
10.4.4- Passage of Mouse L Cells for Plaque Assay
10.5-Other Cell Lines
10.5.1- Culturing Conditions, Splitting and Freezing of L929
10.5.2- Growth of Primary Sheep Neurons in Cell Culture
10.5.3- L. monocytogenes Infection of Primary Neuronal Cell Culture(DRG)
10.5.4- Lmono Intracellular Infection Assay in Fish Cells
10.5.5- PC12 cells Culturing Conditions, and Freezing
10.6-Tissue Culture Media & Reagents
10.6.1- Preparation of 0.1% Trypsin-EDTA_22NOV10