BIO-RAD

Virus Detection and Transmission Kit
Catalog #17008261EDU

Instructor Guide
Note: Duplication of any part of this document is permitted only for noncommercial, educational use. Commercial use of
copyrighted content, including use in for-sale instructional materials, requires express permission from Bio-Rad Laboratories, Inc.
Contact us at explorer@bio-rad.com for more information.
For technical support, call your local Bio-Rad office, or in the U.S. call 1-800-4BIORAD (1-800-424-6723) option 2.

Dear Instructor
Thank you for fostering curiosity in our future scientists and citizens as
they prepare for a future of thinking critically, applying knowledge, asking
questions, and working collaboratively to solve real-world problems.
In 2020, the world changed for you and for your students as the
COVID-19 pandemic spread across the globe and into your community.
As scientists worked quickly to identify the virus responsible and create
tests and treatments, public health officials worked hard to decide thebest course of action to keep communities safe. The response to any
pandemic is an intricate and complex dance between establishedscience, constantly evolving data, and the desire to keep people
safe without unforeseen negative impacts on economies,social safety nets, and overall public health.
The Virus Detection and Transmission Kit teaches thecore skill of DNA gel electrophoresis within a context
that gives students a broad understanding of virology,
viral pathology, epidemiology, and an application of
PCR. It also offers insights into a range of professions
that intersect and work together during the response
to a viral disease outbreak. Students begin byworking through a fictional patient diagnosis and thenlearn how PCR can be used in pathogen detection.
They use agarose gel electrophoresis on simulated DNA
samples to determine who in a restaurant was infected bya novel virus. Students then apply their data to other pieces of
information to construct an evidence-based explanation for how thevirus was transmitted throughout the restaurant.
This kit offers you a choice of four different scenarios: two scenarios for acoronavirus, and two scenarios for a norovirus. The coronavirus activities
include an optional activity in which students explore the considerations
public health officials must make in response to an outbreak of a novel
virus. It is our hope you will find the scenario that best engages yourstudents and supports your curriculum.
We strive to continually improve our curricula and products, and your input
is extremely important to us. We welcome your stories, comments, andsuggestions.
Bio-Rad Explorer Team
Bio-Rad Laboratories
6000 James Watson Drive, Hercules, CA 94547
explorer@bio-rad.com

iTable of Contents

Contents
Before You Start
Kit Storage............................................................................................................................................. 1
Safety Guidelines................................................................................................................................... 1
Considerations for Sensitive and Complex Subject Matters.................................................................... 1
Kit Components..................................................................................................................................... 2
Introduction.............................................................................................................................................. 3
Solve the Mystery of How a Virus Spread through a Restaurant............................................................. 3
Resources............................................................................................................................................. 3
		 Student Guides.................................................................................................................................. 3
		 Answer Guide..................................................................................................................................... 3
		 Other Resources................................................................................................................................ 3
Kit Activity Overview............................................................................................................................... 4
		 Activity 1: Learning about Virus Biology and Virus Detection............................................................... 4
		 Activity 2: Detecting Infections............................................................................................................ 4
		 Activity 3: Building Transmission Models............................................................................................. 5
		 Activity 4: Mitigating Risk (CoV Activities, optional).............................................................................. 5
Activity Timelines.................................................................................................................................... 6
Curriculum Fit........................................................................................................................................ 6
		 Required Prior Knowledge.................................................................................................................. 6
		 Curriculum Fit and Topic Considerations............................................................................................. 6
Preparation Instructions
Select the Virus and the Transmission Scenario..................................................................................... 7
Prepare Reagent Stocks........................................................................................................................ 8
Aliquot Controls and Restaurant Staff and Patron Samples.................................................................... 8
Prepare Agarose Gels, TAE Buffer, and Stain (Fast Gel Protocol Only).................................................. 10
Activity 1: Learning about Virus Biology, Pathophysiology, and Detection..................................... 11
Activity 2: Detecting Infections............................................................................................................ 15
Activity 3: Building a Transmission Model.......................................................................................... 18
Activity 4 (Optional): Mitigating Risk.................................................................................................... 20
Appendix A
Ordering Information............................................................................................................................ 21
Appendix B
Fast Gel Protocol and Electrophoresis Preparation Instructions............................................................ 22
Appendix C
Activity 3 Restaurant Layout and Staff and Patron Cards..................................................................... 25
Appendix D
Resources........................................................................................................................................... 31

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Before You Start

Kit Storage

Safety Guidelines

When you receive the Virus Detection and Transmission Kit:

14°C
Record storage location and batch numbers from the product2

Store the DNA Electrophoresis Samples bag–20°C

in the freezer(–20°C).

4°C

labels.

For short-term storage, the reagent bag may be

5Store the remaining reagents andRoom Temp

4°C

4°C

If using UView Loading Dye and Stain, use
20°C
appropriate personal protective equipment whenusing a transilluminator, as UV light can irradiateskin and eyes.

!

–20°C

4°C
consumables at room temperature
4°C (~20°C).
4°C

Room Temp

20°C

Review this Instructor Guide –20°C
and included Answer Guide andchoose which of the four scenarios to run with your class.
4°C

6

Use special caution when working with hot molten
agar while preparing gels: wear heat-resistant
gloves, goggles, and a laboratory coat.

If using UView 6x Loading Dye and Stain,
store it in the refrigerator (4°C).

4!

If using Fast Blast DNA Stain, note it is not toxic but
will stain hands and clothes. Wear gloves and labcoats to prevent stains.

stored in the refrigerator (4°C) for up to 1 month.

3

Always follow basic laboratory safety guidelines:
wear gloves and safety goggles; do not eat, drink,smoke, Roomor Tempapply cosmetics in the work area. If any
gets into a student’s eyes, flush with water
4°C solution20°C
for 15 minutes.

Visit bio-rad.com/VDTkit to download the Student Guide foryour choice of activity (coronavirus or norovirus).

Considerations for
Sensitive and Complex
Subject Matters!

Some of your students may have personalconnections to the issues around the COVID-19
pandemic. They may know someone affected orthey may harbor misconceptions about the causesof illness or strategies undertaken to limit infection(for example, lockdowns, mask mandates, andvaccination). This kit offers a choice of focusing ona norovirus or coronavirus spread, so choose theactivity that you feel might be most appropriate foryour classroom.
Before addressing this sensitive topic in theclassroom, you may want to prepare yourself
to provide additional support for your students.
Appendix D has some resources to assist.

Technical Support is available at support@bio-rad.com or
1-800-4BIORAD, option 2.

1

Students likely have differing opinions on how the
COVID-19 pandemic was addressed, and thediscussions your students may have while going
through this set of activities provide opportunities
to discuss medical privacy, the consequences ofunlimited virus spread, the unintended consequencesof different mitigation measures, and the importantand varied contributions of various areas of expertise.
Be sure students understand that these topics are
not straightforward, and that research and policies
are in constant development and must change when
new information becomes available.

Before You Start

Kit Components
Each kit contains materials for eight (8) student workstations.
Virus Detection and Transmission Kit

Quantity

1.5 ml EZ Micro Test Tubes
Molecular Weight Ruler
DNA Sample 1
DNA Sample 2
Orange G Loading Dye, 5x

90
200 µl215 µl250 µl1 ml

Small Fast Blast DNA Electrophoresis Pack

Quantity

Fast Blast DNA Stain
Certified Molecular Biology Agarose
TAE Electrophoresis Buffer, 50x

100 ml5g
100 mlSmall UView DNA Electrophoresis Pack

Quantity

UView 6x Loading Dye and Stain
Certified Molecular Biology Agarose
TAE Electrophoresis Buffer, 50x

200 µl5g
100 mlRequired Materials (not included in this kit)

Quantity

20–200 µl adjustable-volume micropipet and tips
2–20 µl adjustable-volume micropipet and tips
or 10 µl fixed-volume micropipet and tips
Microcentrifuge tube rack
Horizontal gel electrophoresis chamberPower supply
UV transilluminator (if using UView 6x Loading Dye and Stain)
Marking pen
Waste container
Gel staining tray

1
8
8
8
4–8
1–8
1
8
8
8

Microcentrifuge tubes, molecular weight ruler, DNA Sample 1,
DNA Sample 2, and Orange G Loading Dye, 5x

Fast Blast DNA Stain, Certified Molecular Biology Agarose,and TAE Electrophoresis Buffer, 50x

UView 6x Loading Dye and Stain, Certified Molecular Biology
Agarose, and TAE Electrophoresis Buffer, 50x

Refer to Appendix A Ordering Information for ordering details for some ofthese required materials and for kit refills.

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Before You Start

Introduction
Solve the Mystery of How a Virus Spread through a Restaurant
Two patients stumble into a hospital with symptoms associated with a newly emerging virus. Doctors diagnose the patients andfind one is infected with the novel virus. This person had eaten lunch at a local restaurant earlier that day, so public health officials
track down others who were also at the restaurant to determine whether and how the virus may have spread. Based on publishedviral transmission case studies, this hands-on lab activity puts your students into the roles of an emergency room (ER) doctor,
medical laboratory scientist, epidemiologist, and public health official as they analyze patient samples and use patient information todetermine how a novel virus strain spread through a restaurant.
In this laboratory activity, students analyze simulated samples collected from staff and customers at a fictitious restaurant where aviral transmission event took place.
• Use one of four different viral transmission scenarios: choose between two different viruses, each with two different		transmission scenarios• The patterns of infection among the customers in the restaurant vary between the scenarios to highlight different aspects of		 viral disease transmission• See the Preparation Instructions section in this guide for details of the four different scenarios• Select a scenario for your class before preparing reagentsYour students compile classroom sample data and propose explanations for how the virus was transmitted.
Resources
Student Guides
There are two Student Guides:• Student Guide (CoV) covers your choice of two coronavirus transmission scenarios• Student Guide (NoV) covers your choice of two norovirus transmission scenarios
Choose which virus and transmission scenario to run with your class. Each guide has the background information you and yourstudents will need. This Instructor Guide highlights key points to emphasize with your students and provides additional details to help
you address questions that may come up during class discussion.
Answer Guide
The Answer Guide provides extra details about the transmission scenarios that should be withheld from students to allow them toget the most out of the activity. Review the details in the Answer Guide about each transmission scenario to help you select the onethat would be best for your classroom.
Other Resources
Please refer to Appendix D Resources for additional information and instructional media.

3Before You Start

Kit Activity Overview
The Virus Detection and Transmission Kit allows you to conduct your choiceof four different virus transmission scenarios: two different transmission
scenarios for each of two different viruses (coronavirus or norovirus).

Activity 1
Learning about Virus Biology andVirus Detection

Fever
Congestion

Two patients come to the emergency room at AnyTown Hospital. Both are

Cough

aches
showing symptoms of a viral disease. Students assess the symptoms
Activityand
1: Learning About Virus Biology,Body
Pathophysiology,
and Detection

order a molecular test to assist them in their diagnosis.

Fever
Congestion

Review patient
symptoms, make ➜Patient Symptom Review
a hypothesis
Students evaluate the patients’ symptoms against a list of known
afflictions
about
cause

Cough
Body aches

and make a hypothesis about possible causes.

Discussviral biology,
pathophysiology,
Students read about and discuss viral biology, viral structure, and viraland molecular
Activity 1: Learning About Virus Biology, Pathophysiology,
andDetection
diagnostic
tests
pathophysiology (how viruses cause disease). These concepts are

Learning about Viruses, Pathophysiology, and Detection➜

important for understanding viral detection and modes of transmission.

Review patient
symptoms, makeaLoadhypothesis
restaurant
Students learn about molecular diagnostic tests in general, and
about
the
about
cause
staff andpatronFever

Activity 2: ActivityCongestion
2: Infection Detection

Patient Diagnosis

Cough
Body aches

components of a reverse-transcription polymerase chain reaction samples(RT-PCR)based test and agarose gel electrophoresis. They analyze provided data to

Run DNA gel
Discuss
electrophoresis

determine the infection status of the two patients.

Activity 3: Building Transmission Models

2

4

2

6

3

Activity 2: Activity 2: Infection Detection
A

1

B

2

4

D

8

Load restaurant
staff and patronsamples➜

Opioid drug prescription
drug prescription opioid
opioid drug prescription
prescription opioid drug
drug opioid prescription
drug prescription.

W1

WaiterW2

Waiter

7
2

5

3

1

3

E

F
4

Ent
ran

K1

4

6

1

6

1

2

5

Air conditioner

restaurant where the infected patient had been on the day of diagnosis.

5

C

7

1

Students analyze samples obtained from the staff and patrons of a local

Sample Analysis by Agarose Gel Electrophoresis

3

5

4

3

Air conditioner

Apply data torestaurant layout

Closed vents

Activity 2
Detecting Infections

viral biolog
pathophysioloand molecu
diagnostic te

ce

Cook

1,000 mg

Students perform agarose gel electrophoresis on staff and patron samples

Run DNA gelectrophoreand controls.
Claim

Collecting Class Data

Develop a modelDfor howtransmission
Reasoningoccurredoin theActivity
3: Building Transmission Models
restaurant
Evidence

Students combine their data into an analysis of the infection status of theentire restaurant.

Chain of
Infection
I

2
Closed vents

Activity 4:: Mitigating
Risk
2
1
4

B

Number of cases

W1
W2
Opioid drug prescription
drug prescription opioid
opioid drug prescription
prescription opioid drug
Without
drug opioid prescription
protective
measuresdrug prescription.

Waiter

Waiter

Entra

nce

K1

Cook

With
protective measures

1,000 mgTime since first case

4
3

5

C

7
8

1

4

2

6

1

7
2

5

3

1

E

Health caresystem capacity
2

F
4
1

6

5

Air conditioner

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3

D

Decide whether
and how to
mitigate further
spread

3

5

4

3

A

Air conditioner

Apply data torestaurant layout

4Before You Start

2
Closed vents

Closed vents

1

nce

W2

82

5

4
5

C

77

E

6

1

3

7

F
4

4

3

5

6

1

2

Entra

1,000 mg➜

Waiter

1

1

3

K1 W1
Cook Waiter

Opioid drug prescription
drug prescription opioid
opioid drug prescription
prescription opioid drug
drug opioid prescription
drug prescription.

Students learn the six components of a chain of infection.

W2

1

4

D

B

3

5

2

3

1

E

F
4

Waiter

2

Entra

1

6

nce

K1

Cook

1,000 mg

Claim

Creating a Transmission Model

Evidence

Students analyze the complete set of data from the restaurant andadditional restaurant information provided to propose an explanation for

Chain of
Infection
I

Reasoning

Claim

how the virus spread.

Evidence

Activity 4
Mitigating Risk (CoV Activities, optional)

Chain of
Infection
I

Number of cases

ReasoningStudents complete their investigation by considering whether and how toaddress the outbreak.

Health caresystem capacity

Without
protective measures

With
protective measures

Time since first caseStudents learn how public health officials develop strategies for mitigating
disease outbreaks in communities.
Developing a Response Strategy for the AnyTown Outbreak➜
Number of cases

Understanding Response Strategies to Disease Outbreaks

Health caresystem capacity

Without
protective measures

With
protective measures

Students develop their own strategies for mitigating spread of the virus inquestion.

5Time since first case5

Air conditioner

Opioid drug prescription
drug prescription opioid
opioid drug prescription
prescription opioid drug
drug opioid prescription
drug prescription.

Waiter2

452
6

Air conditioner

throughout the restaurant.

W1

8

4

6C

7

3

1

D

the restaurant patrons and staff to figure out how the virus spread

Understanding the Chain of Infection

A

2

5

4

B

Air conditioner

Students integrate the diagnostic test data with other information about

4

2

6

33

1

3

5

4

3

A2

Air conditioner

Activity 3
Building Transmission Models

Before You Start

Activity Timelines
The activities in this kit are designed to take four 45-minute class periods as shown in Table 1.
Table 1. Alternative timelines.
Class Period 1

Class Period 2Class Period 3

Class Period 4(CoV Activities, Optional)

In-class or out-of-class

In-class

In-class

In-class or out-of-class

Activity 1
Part 1. Patient Symptom Review

Activity 2
Part 1. Pre-Laboratory Questions

Part 2. Learning about Viruses,
Pathophysiology, and Detection

Part 2. Sample Analysis by
Agarose Gel Electrophoresis

Activity 3
Activity 4
Part 1. Understanding the Chain Part 1. Understanding Responseof Infection
Strategies to Disease Outbreaks

Part 3. Patient Diagnosis

Part 3. Analysis

Part 2. Creating a Transmission
ModelPart 4. Collecting Class Data1

Curriculum Fit
Required Prior Knowledge• Cell structure and basic cell theory• Basic DNA, RNA, lipid, and protein structure and function• Roles of DNA and RNA in encoding heritable information• Basic principles of PCR and DNA electrophoresis• How to use a micropipet
Curriculum Fit and Topic Considerations• Biology and virology — students explore concepts surrounding basic virus structure and pathophysiology, including thefunction and interaction of viral proteins, lipids, and nucleic acids• Human body systems — the two viruses that are the focus of this investigation infect the human respiratory andgastrointestinal systems, which are also reviewed• Careers in the life sciences — throughout the activities, career highlights expose students to various career options and paths• Science practices — in these activities, students analyze data, incorporate knowledge of virology to deduce a mode oftransmission in a restaurant• Ethical, legal, and social issues — there are broader issues associated with viral outbreaks and a community response,
including health, social, and economic impacts

1

Depending on the gel staining method used, this step may move to class period 3.

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Instructor’s Preparation

Preparation Instructions
Preparation Step

Time Required

When to Begin Preparation

Select the virus and transmission scenario

5–15 minutes (reading)Prior to reagent preparation

Prepare reagent stocks

30 minutes

Up to a year before Activity 2 (store the stocks in the refrigerator
for one month or at –20°C for up to one year)

Aliquot controls and samples

30 minutes

Up to a year before Activity 2 (store the controls and samples in
the refrigerator for one month or at –20°C for up to one year)

Prepare agarose gels, TAE buffer, andstain

1 hourUp to a week before Activity 2 (store gels in a sealable plastic
bag with a few ml of buffer at room temperature for up to 1 day,or in the refrigerator for up to 1 week; store TAE buffer and Fast
Blast Stain at room temperature)

Select the Virus and the Transmission ScenarioFirst, select one of the virus and transmission scenarios described below. Then:• Print or download and review the Student Guide that corresponds to the virus you will be investigating• Label and dispense DNA samples using the table corresponding to the scenario you chose
The mode of transmission impacts the distribution of positive and negative test results in the restaurant. Select the virus andmode of transmission before preparing the samples. Refer to the Answer Guide for more details on the specific aspects ofeach scenario below.
Coronavirus (CoV)

Norovirus (NoV)

Analysis of transmission of a fictitious new strain of coronavirus
named PLS-CoV, which causes a new fictitious respiratory disease
called pneumonia-like syndrome, or PLS.

Analysis of transmission of a fictitious new strain of norovirus named
EBS-NoV, which causes a new fictitious disease called Extrusive
Bowel Syndrome, or EBS.

Scenario CoV 1(Indirect transmission through airborne aerosols)

Scenario NoV 1(Indirect transmission through contaminated food)This activity is based on a real event that occurred early in the
COVID-19 pandemic (Lu et al. 2020). At the time of the spreading
event, few details were known about how the virus spread. In this
scenario, staff and patrons of a fictitious restaurant were primarily
infected by inhalation of small, airborne droplets containing virus
particles.

In this scenario, patients were primarily infected indirectly bycontaminated food.

Scenario CoV 2(Direct transmission through airborne droplets)

Scenario NoV 2(Indirect transmission through contaminated food and fomites)

In this scenario, restaurant staff and patrons were primarily infectedby short-range, direct exposure to large droplets containing virus
particles.

In this scenario, patients were primarily infected indirectly by fomites(contact with contaminated surfaces) and contaminated food.

7

Instructor’s Preparation
Prepare Reagent Stocks

150 µl150 µl
75 µl

Reagent stock preparation will take 30 min and may be done up to a year
before Activity 2. Store the DNA sample stocks in the refrigerator for one

LoadingDyemonth or at –20°C for up to one year.
1.

GAPDH control stock (–). Label a microtube (–). Using a fresh pipet➜

Distilled water

(–)

DNA
Sample 2 Loadingdye
150 µltip each time, add the following and pipet to mix:

150 µl100 µl75 µl150 µl150 µl25 µl• 150 µl DNA Sample 2

75 µl

Loading
Dye
LoadingDye• 150 µl distilled water

Loading
DyeDistilled water(–)
DNA
Sample 2 Loading
(NTC)
dye
Loading(–)
DNA dyeSample 2 Loadingdye• 75 µl loading dye (Orange G or UView 6x Loading Dye and Stain)

Distilled waterDistilled water
100 µl2.

Virus + GAPDH DNA stock (+). Label a microtube (+). Using a fresh

➜

90 µl
150 µlpipet tip each time, add the following and pipet to mix:

45 µl
75 µl 25 µl

Loading
Dye
LoadingDye• 90 µl DNA Sample 1

LoadingDye• 90 µl DNA Sample 2

Loading
Dye➜

Loading
Dye Loading
Dye

DNA
Sample 1• 100 µl distilled waterDNA
Distilled
water
Sample2 water
Distilled
90 µl
100 µl 90 µl

25 µl 45 µl

LoadingDye

tip add the following and pipet to mix:(+)(–) (NTC)

50µlµl45

90 µl

No-template control (NTC). Label a microtube (NTC). Using a freshDistilled water(NTC)

Loadingdye
DNA
Loading
SampleDNA 1
dye Loading
Loading
Sample 2dye dye90 µl• 45 µl loading dye (Orange G or UView 6x Loading Dye and Stain)

3.

90 µl
150 µl100 µl

25 µl

Loading
Dye
MWR

(+)

Loadingdye

DNA
Sample 2
Distilled water
DNA(NTC) (+)
DNA dyeLoading
Loading
Sample 1 Loading
dye dyeMWR• 25 µl loading dye (Orange G or UView 6x Loading Dye and Stain)

Sample 2

50 µl
90 µl

4.

Molecular weight ruler (MWR). Using a fresh pipet tip, add

➜

90 µl

50 µl
45 µl
11 µl

LoadingDye

50 µl Orange G Loading Dye or UView 6x Loading Dye and Stain

MWR

Stock DNA samples(CCS, TTS, CTS)

LoadingDyeto the molecular weight ruler. Pipet to mix.

Loading
Dye
MWR

Loading
MWR dyeDNA
Sample 1

Loadingdye dyeLoading(+) MWR
MWR
CT: 8 tubes

MWR
50 11
µl µl

Aliquot Controls and Restaurant Staff and Patron Samples

Stock DNA samples(CCS, TTS, CTS)

11 µl

Labeling and aliquoting tubes will take 30 min and may be done up to one
year ahead of Activity 2. Store the DNA samples in the refrigerator for one

LoadingMWR
Dye

244–269
Stock DNA sam(CCS, TTS, CT

137–158
MWR

month or at –20°C for up to one year.

MWR

CT: 8 tubes

Loading
MWRdyeControls
5.

DNA
Sample 2

Label tubes and fill them according to the following table.

Quantity

Tube Label

Sample Stock

Volume

8

(–)

(–)

11 µl

8

(+)

(+)

11 µl

8

NTC

NTC

11 µl

8

MWR

MWR

22 µl

137
137
137
137
137
137
137
137
137
##

11 µl
137–158CT:
9 tubes

CC: 10 tubes

250
250
250
250

MWR
147
147
147
147
147
147
147
147
##

MWR

CT: 8 tubes
156
156
##

##

TT: 3 tubes

CC: 6 tubes##
244–269
CT: 14 tubes
Stock DNA samples(CCS, TTS,
CTS)
244–269

137–158
MWR

137
137
147
137
147
137
147
137
147
137
147
137
147
CT: 8 tubes
137
147
156
137
147
156
MWR
##
##
##
137
137
147
137
147
137
147
137
147
137
147
137
147
137
147
156
137
147
156
##
##
##
CC: 10 tubes
CT: 9 tubes
TT: 3 tubes
CC: 10 tubes

CT: 9 tubes

TT: 3 tubes

250
250
250
250
##

##

##
CC: 6 tubes##
CT: 14 tubes

CC: 6 tubes

137–158

22

CT: 14 tubes
244–2698

explorer.bio-rad.com

137
137
137
137
137

147
147
147
147

250
250
250
250

Instructor’s Preparation
Restaurant Samples
6. Choose the scenario you will be running with your class, then label
32 tubes as indicated in the following table.
• Omit the tube corresponding to Patient A for your scenario
		 (for example, for scenario NoV 1, omit E4)
7.

Dispense 11 µl of each sample stock into each tube according to 		the table.

• Use only the column that matches your chosen scenario.
		 For example, for scenario NoV 1, tube A4, add 11 µl of (–) sample 				stockSeat

CoV Scenario

Seat

NoV Scenario

CoV 1

CoV 2

NoV 1

NoV 2

A1

(–)

(–)

A1

(–)

(–)

A2

(+)

(–)

A2

(–)

(+)

A3

(+)

(–)

A3

(+)

(–)

A4

(+)

(–)

A4

(–)

(–)

B1

(+)

(–)

B1

(–)

(+)

B2

(+)

(–)

B2

(+)

(–)

B3

(Patient A)

(–)

B3

(+)

(–)

B4

(–)

(–)

B4

(–)

(–)

B5

(–)

(–)

B5

(–)

(–)

B6

(+)

(–)

B6

(–)

(+)

B7

(–)

(–)

B7

(+)

(–)

B8

(+)

(+)

B8

(–)

(–)

C1

(–)

(+)

C1

(–)

(–)

C2

(–)

(–)

C2

(–)

(–)

C3

(+)

(–)

C3

(–)

(–)

C4

(–)

(–)

C4

(+)

(–)

C5

(–)

(–)

C5

(–)

(–)

C6

(+)

(–)

C6

(–)

(–)

C7

(–)

(+)

C7

(+)

(+)

E1

(–)

(–)

E1

(–)

(–)

E2

(–)

(–)

E2

(–)

(+)

E3

(–)

(+)

E3

(–)

(–)

E4

(–)

(+)

E4

(Patient A)

(–)

E5

(–)

(–)

E5

(–)

(–)

F1

(–)

(+)

F1

(–)

(+)

F2

(–)

(+)

F2

(–)

(+)

F3

(–)

(+)

F3

(–)

(Patient A)

F4

(–)

(–)

F4

(–)

(+)

F5

(–)

(–)

F5

(+)

(+)

F6

(–)

(–)

F6

(+)

(–)

W1

(–)

(Patient A)

W1

(–)

(–)

W2

(–)

(+)

W2

(–)

(–)

K1

(–)

(–)

K1

(+)

(–)

9

Instructor’s Preparation
Prepare Agarose Gels, TAE Buffer, and Stain(Fast Gel Protocol Only)
8.

Prepare eight 1% TAE agarose gels with 8 wells each. Use 1x TAE(not 0.25x TAE) to make the gels. See Appendix B for gel casting
instructions.
150 µl9.

150 µl➜Prepare 2.2 L of 0.25x TAE electrophoresis
buffer by adding 11 ml
75 µl

11 ml

2,200 mlof 50x TAE to 2,200 ml of distilled water to use with the Fast DNA
Loading
Dye

Gel Protocol.2

Distilled water

(–)

DNA
Note: the correct volumeof distilled water would be 2,189 ml, but
Sample 2 Loadingdye

50x TAE

Distilled water

500x Fast Blast

Distilled water

the difference does not affect results.
100 µl25 µl➜

10. If using, prepare 100x Fast Blast DNA stain for staining of agarose
LoadingDyegels. See Appendix B for instructions.
Distilled water(NTC)

Loading
Note: Fast Blast DNA stain
is not required if using UView 6x Loadingdye

Dye and Stain for visualization.
90 µl

90 µl

45 µlStaff and Patron Cards and
11. Print 8 copies of the Restaurant

8 Restaurant Schematics. Provide these to each student group
Loading
Dyefor Activity 3.

11 µl
DNA
Sample 1(+)

Loadingdye

DNA
Sample 2

12. Download and familiarize yourself with the Student Guide for thevirus you wish to focus on. Do not tell students which

CT

50 µl

transmission scenario they will be investigating — they will

CTS
CT: 8 tubes

apply their class results and what they know about the virus andpossible modes of transmission to deduce the mode of transmission.
Loading
Dye

MWR

Loading dye

22 µl

11 µl

MWR

MWR

MWR

###

11 µl
Stock DNA samples(CCS, TTS, CTS)

MWR: 8 tubes

MWR

48 tubes

MWR

CT: 8 tubes

MWR

244–269

137–158

DNA
Sample
1

2

137
137
137
137
137
137
137
137
137
##

147
147
147
147
147
147
147
147
##

Loading
Dye

DNA
Sample
Stock

250
250
250
250
156
156
##

##

##

This activity is designed to take advantage of Bio-Rad’s Fast Gel Protocols using 0.25x TAE
running buffer and higher voltages. See Appendix B for details and alternate electrophoresis
CC: 6 tubes
CT: 14 tubes
options with preparation
instructions.
CC: 10tubes
CT: 9 tubes
TT: 3 tubes

explorer.bio-rad.com

DNA
Sample2##

TT: 6 tubes

10Instructor’s Guide

Activity 1
Learning about Virus Biology,
Pathophysiology, and Detection
This activity begins with an introduction to two patients who come into
an emergency room with similar symptoms and conditions requiring
diagnosis. It concludes with a review of a molecular diagnostic test thatwill be used to help in diagnosis.
It covers background information about viruses and has multiple
curriculum connections to cell theory, biological molecules, thecentral dogma, body systems, polymerase chain reaction (PCR), andelectrophoresis.
Classroom Preparation• To save class time, assign the reading ahead of time and use the classtime to go over the Focus Questions• Prepare the slide deck and print the pages for Activity 1 from theStudent Guide for students, or electronically distribute by PDF

Part 1: Patient Symptom Review

Points to Emphasize• The spread of a novel virus is of particular concern because
the population will have no immunity against it, and the1.

Review the data shown in Figure 1 and Table 1 of the Student Guide withstudents. Elicit their thoughts, questions, and prior knowledge about
respiratory or gastrointestinal disease and the presentation of symptoms.

extent of disease and possible treatments may be unknown• Different diseases and medical conditions can have similar
symptoms, which can make diagnosis tricky• Doctors rely on their experience, patient history, and2.

As a class, discuss student observations and record their ideas. “Do
your observations justify ordering a diagnostic test? Why? What are thepossible outcomes of not obtaining an accurate diagnosis?”3.

Have students review the reading and answer the Focus Questions.

11

diagnostic tests in making a diagnosis

Instructor’s Guide
Part 2: Learning about Viruses, Pathophysiology,
and Detection
1.

Review basic facts about viruses, emphasizing virus structure.

2.

Ensure students understand (i) the difference between an enveloped andPoints to Emphasize• The diversity of viruses makes it difficult to make generalstatements about them
• Only a small fraction of viruses cause disease in animals,
plants, bacteria, or other organisms

unenveloped virion, (ii) enveloped viruses require an intact lipid envelope to
infect, and (iii) the genetic material within a virus can be DNA or RNA.
3.

Compare and contrast coronaviruses and noroviruses.

4.

Review the different types of molecular diagnostic tests, focusing on PCRmediated tests.

5.

As a class, review the Focus Questions.

6.

Point out the career highlights that appear throughout this activity.
Addressing a disease outbreak requires people from all sorts of different
professions to work together and contribute their expertise.

Part 3: Patient Diagnosis
1.

Review the RT-PCR diagnostic test for the scenario you will be using.

Points to Emphasize• The virus in question in these activities is an RNA virus. Thismeans the PCR-based diagnostic test requires a preliminary
step of reverse transcription2.

Reinforce that the test is looking for two different targets in a single tube.
This means it uses two different sets of primers (multiplex assay).

• Each sample will produce one or two different bands:
a band corresponding to human GAPDH (all samples
will have this, and it shows the test worked) and a band

3.

Review the controls that will be used.

4.

Review the basics of agarose gel electrophoresis and, as a class, answer
the Focus Questions.Activity 3

corresponding to a target gene from the virus (only infected
people will have this second band)• Both PCR products (amplicons) will be possible in a single
sample because the PCR test is a multiplex PCR test,
meaning it has two different sets of PCR primers to allow
amplification of two different targets in the same reaction

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12Instructor’s Guide

Instructor BackgroundFun Facts about Viruses
The number of viruses that exist on our planet is staggering to think about. Roughly 1031 individual viral particles (virions) inhabit theoceans alone at any given time — 10 billion times the estimated number of stars in the observable universe (Dance 2021). Within
this incredible number, there is also a lot of structural diversity — viruses exist in a wide range of sizes and shapes.
Points to emphasize with students:• It is difficult to make general statements about viruses• About 320,000 types of viruses are known to infect mammals; students will be focusing on only two in this activity• All viruses do have two main things in common, but there are seemingly endless variations on these two main commonalities:		 • All have a nucleic acid genome encased in a protein-based shell (capsid)		 • All depend on a living host to reproduce• Viruses tend to be characterized according to their shape, host cell, and/or genetic code (Pride 2020)• Only a small fraction cause disease: in animal, plants, and bacteria
Molecular Diagnostic Tests
Diagnostic tests for viruses are based on laboratory techniques that can identify the presence of a viral infection in a patient. These
tests can be divided into two main categories. The choice of test depends on the type of virus being detected, the stage of theinfection, and the clinical context.
Detection of Viral Nucleic Acids (RNA or DNA)
These tests use techniques like PCR and reverse transcription PCR (RT-PCR) to look for the presence of viral DNA or RNA in apatient sample. Highly sensitive and specific, these tests can often detect viral infections even before symptoms appear. They do,
however, require specialized equipment and often take longer to return results. Students may recognize these as the “PCR tests”
used to test for COVID-19.
PCR and Reverse-Transcription PCR (RT-PCR)
PCR and RT-PCR are techniques used to amplify and detect specific DNA or RNA sequences. In this lab activity, students visualize
the end-products of an RT-PCR test for a novel CoV or NoV. These types of tests are usually the fastest to be developed during an
outbreak of a novel virus because they require only the creation of specific DNA primer sets. Developing primer sets is simple andquick once the genomic sequence of a virus is available.
• PCR is used to amplify DNA sequences and so is used to detect DNA viruses. It uses a DNA template, which is		 denatured by heat, and then two primers are annealed to the template. The primers then serve as starting points for a
		 DNA polymerase enzyme to copy and amplify the DNA sequence between them. This process can be repeated multiple
		 times, exponentially amplifying the original DNA sequence, allowing its detection• RT-PCR is used to amplify RNA sequences and so is used to detect RNA viruses. RNA is first converted to DNA
		 using the enzyme reverse transcriptase. The resulting DNA is then used as a template for PCR using DNA primers

13Instructor’s Guide
Multiplex PCR
Be sure to point out to your students that the RT-PCR diagnostic test they are simulating is a multiplex RT-PCR. The test has two
different sets of PCR primers to allow amplification of two different targets in the same reaction.
In their scenario, a single reaction is being used to detect the presence of more than one gene in a sample: the human GAPDH
gene (all human samples should have this, showing the test worked) and a target gene from the virus (only infected people will
have this second product).
Students who have previous experience with gel analysis of a PCR may confuse the presence of two bands in this lab activity witha restriction digestion. The Focus Questions in Activity 1 walk them through the results they should anticipate, but it is a good idea
to repeat the concept of using two sets of primers in this fictitious test.
Detection of Viral Proteins or Patient Antibodies
These tests are used to diagnose past or present infections and can also help determine a patient’s immune status. The mostused tests are enzyme-linked immunosorbent assays (ELISAs) and lateral flow immunoassays (LFIs). Students may recognize thelatter as the rapid home tests used to test for COVID-19. Though often less sensitive and specific than PCR tests, these tests areconvenient; they do not require specialized equipment, and they offer a shorter time to results.

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14

Instructor’s Guide

Activity 2
Detecting Infections
In this activity, students use DNA gel electrophoresis to analyze the
PCR samples of all the people who were in the restaurant at the sametime as Patient A. They then combine their results with results from theirclassmates in preparation for Activity 4.
Classroom Preparation• Prepare student workstations (below)• Print the pages for Activity 2 from the Student Guide for students• Review the electrophoresis run conditions your students will use
		(Appendix B)
Student Workstation
Material

Quantity

Molecular weight ruler (MWR), 20 µl

1

GAPDH control (–), 10 µl

1Virus- and GAPDH-positive control (+), 10 µl

1No-template control (NTC), 10 µl

1Samples from restaurant staff and patrons, 10 µl

4

TAE electrophoresis buffer

300 ml100x Fast Blast DNA Stain (if using)

50 ml

1% TAE agarose gel with 8 wells4

1

Horizontal gel electrophoresis chamber

1Power supply (may be shared)

1

Microcentrifuge tube rack1

Micropipet and tips

1

Gel staining tray (optional)

1

Waste container

1

Activity 2 printout

1

3

3

4

Using 0.25x TAE buffer with a 1% agarose gel enables faster runs using higher voltage.
See Appendix B for details.
You may also use one gel with two rows of eight wells (2 x 8) for two student groups.

15

Instructor’s Guide
Part 1: Pre-Laboratory Questions
Students review their understanding of the diagnostic test and enter the
purpose for each of their samples into Table 3.

Part 2: Patient Sample Analysis by Agarose Gel
Electrophoresis
1.

Review the available materials and procedure with students.

2.

Introduce the expanded scenario: students will combine their results to get
a complete picture of virus transmission in the restaurant.

3.

Remind students to load 20 µl of MWR, 10 µl of control samples (–, +, andNTC), and 10 µl of each of their four participant DNA samples on their gel.

Points to Emphasize

4.

Ensure students record their sample numbers.

5.

Instruct students which electrophoresis conditions to use (voltage anderror is to incorrectly record what was loaded into each welltime settings; see Appendix B).

of a gel.

Accurate data recording is vital to any investigation. A common

6.

Have students conduct their investigations and visualize their DNA gels using
Fast Blast DNA Stain or UView 6x Loading Dye and Stain (see Figure 1,
below). Refer to Appendix B for visualization options and protocols.

Part 3: Data Analysis
This activity may be performed during the next class period asneeded to accommodate schedules and gel staining method used.
Have students sketch and interpret their gel results as directed in theStudent Guide.

Part 4: Collecting Class Data
This activity may be performed during the next class period asneeded to accommodate schedules and gel staining method used.
• Collect student data in a class data table or other shared space• Have students summarize the class data in the tables in the Student
		 Guide. Let them know which seat belonged to Patient A and have		 them indicate that spot as having tested positive

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16

Instructor’s Guide
Agarose Gel Electrophoresis and Different Gel Stains
In real life, PCR-based tests are performed using instruments and other PCR-based technologies that provide readouts indicating
infection status. For example, viruses are typically detected with quantitative real-time PCR (qPCR), which uses a different
amplification chemistry and a type of thermal cycler that provides real-time, quantifiable readouts of results.
In this lab activity, however, students perform agarose gel electrophoresis to separate and visualize the PCR products.
A few notes regarding this procedure:• 10-minute protocol − DNA separation can be hastened on an agarose gel using modified conditions and alternate reagents;
		 refer to Appendix B for options for running a gel in as little as 10 minutes• Smaller bands run together − short separation times and distances will affect resolution of smaller bands; for this reason,		 the smallest of the MWR bands may run together and appear as a single, fuzzy, or wider band
• Visualization/staining options − this kit offers two visualization options; both are nontoxic options to ethidium bromide:		 • Fast Blast is used to stain DNA after separation in a gel (it stains the entire DNA-containing gel). It is economical but
			 requires more time and requires use of loading dye, which is included with the kit
		 • UView 6x DNA Loading Dye and Stain includes the loading dye and stains the DNA prior to separation. It offers instant			 visualization using a UV transilluminator or lamp

Patient B

Patient A

NTC(+) Control(–) Control

MWR

Patient B

Patient A

NTC(+) Control(–) Control

MWR

The gel appearance will vary slightly, depending on the method used (Figure 1).

1,000 bp
700 bp500 bp200 bp100 bpFigure 1. Comparison of agarose gels visualized using Fast Blast Stain (left) and UView 6x Loading Dye and Stain (right). Both gels were run using the
10-minute run protocol. Note the smallest bands in the MWR have not resolved completely. Note also that the dye in the UView 6x Loading Dye and Stain separates
and creates its own dark bands which can mask bands in the MWR and samples if the separation time is insufficient.

17Instructor’s Guide

Activity 3
Building a Transmission ModelIn this activity, students learn about the chain of infection. They then analyze
the classroom set of data from the restaurant, apply other patient andrestaurant information, and propose an explanation for how the virus spread.
Classroom Preparation• Print a restaurant layout and set of Restaurant Staff and Patron Cards
		 for each workstation (see Appendix C)• Ensure students have the class results from Activity 2

Part 1: Understanding the Chain of
Infection
1.

Review basic facts about the chain of transmission.

2.

Ensure students understand how a mode of transmission may relate backto the structure of the virus of interest.

Points to Emphasize
Differences in virus structure impact their modes oftransmission and infection. Enveloped viruses must maintainan intact lipid envelope and envelope proteins to infect a host
cell, which means the virion must remain hydrated. Enveloped3.

As a class, review the Focus Questions.

viruses, therefore, are more likely to spread through respiratory
droplets or aerosols or other bodily fluids like blood.

This review may also be done outside of class by the students in
preparation for Part 2.

Part 2: Creating a Transmission Model
1.

Have students apply their class results from Activity 2 to the restaurant
layout.

2.

Have students apply any additional information they may think is importantfrom the restaurant staff and patron cards to the restaurant layout.

3.

Who was Patient A? Invite students to explain why the Patient A sample
was not included in their class analysis. (Patient A was tested anddiagnosed at the hospital.)

4.

Have the students propose a mode and route of transmission of the virusof interest in the restaurant. Students should consider all information
provided, including the dimensions and features of the restaurant.

5.

Direct students to support their claim with evidence from the restaurant
and using other information they know about the type of virus.

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18

Instructor’s Guide
6.

Invite students to build a chain of transmission using as many of the points
in a chain as they can identify.

Points to Emphasize
Though students may be looking for a primary mode of7.

Ask students to examine the data and highlight any patterns or aspects ofthe data that support or conflict with their proposal.

transmission, emphasize that in most cases, transmission
can occur by multiple modes, for example fomite and foodcontamination, both aerosol and droplet transmission, droplet,and fomite, etc.

8.

Invite students to propose additional experiments epidemiologists might
use to test the model for how transmission occurred.

Instructor Background
About Coronavirus Transmission
This activity is based on a real-world case study of coronavirus transmission in a restaurant (Lu et al. 2020). Though coronaviruses
are often transmitted through respiratory droplets or aerosols (or both), other modes of transmission are also possible.
About Norovirus Transmission
Noroviruses are highly contagious; <100 viral particles may be sufficient to infect a person. They are transmitted primarily throughthe fecal–oral route, either by consumption of contaminated food or water or by spreading directly from person to person. Shellfish
and salad ingredients are the foods most often implicated in norovirus outbreaks, and many norovirus outbreaks have been traced
to food handled by only one infected person.
In Quebec, Canada, in 2017, an outbreak of norovirus sickened more than 700 people. The culprit was imported frozen raspberries,
so Canadian authorities issued a recall on raspberries and raspberry products from the region in question (Sherwood 2020).

19

Instructor’s Guide

Activity 4 (Optional)
Mitigating Risk
This activity correlates only to the coronavirus scenarios and is optional.
It explores the considerations public health officials must make indetermining whether and how to respond to the outbreak of a novel
coronavirus.
Classroom Preparation• This optional activity covers information about how communities
		 might respond to an outbreak of a novel virus• Prepare the slide deck and print the pages for Activity 4 from the		 Student Guide or distribute PDF electronicallyPart 1: Understanding Response
Strategies to Disease Outbreaks
1.

Show students the data in Figure 12 and Table 1 of the Student Guide.
Elicit their thoughts and questions about an epidemic curve.

2.

Have students discuss the impacts and potential risks of allowing
uncontrolled spread, especially of a novel virus, to answer the Focus
Question.

Part 2: Developing a Response Strategy
for the AnyTown PLS Outbreak1.

Review basic facts the prevention strategies in Table 1 to elicit theirthoughts and questions about the relative efficacy of each strategy
and how they might be combined.

2.

Answer the Focus Questions.

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20

Appendix A

Ordering Information
Kits and Consumables
Catalog #Description

17008261EDU

Virus Detection and Transmission Kit

17008251EDU

Virus Detection and Transmission Kit plus Small Fast Blast Electrophoresis Pack

17008241EDU

Virus Detection and Transmission Kit plus UView Electrophoresis Pack

1660450EDU

Small Fast Blast DNA Electrophoresis Reagents Pack, includes 25 g agarose powder, 100 ml 500x Fast Blast DNA Stain,
100 ml 50x TAE electrophoresis buffer

1660462EDU

Small UView DNA Electrophoresis Reagents Pack, includes 25 g agarose powder, 1 ml UView 6x Loading Dye and Stain,
100 ml 50x TAE electrophoresis buffer

1613116EDU

Certified Molecular Biology Agarose, 5 g

1613100EDU

Certified Molecular Biology Agarose, 25 g

1660742EDUTAE Electrophoresis Buffer, 50x, 100 ml

1665111EDUUView 6x Loading Dye and Stain, 200 µl

1665112EDUUView 6x Loading Dye and Stain, 1 ml

2239480EDU

1.5 ml EZ Micro Test Tubes, clear, 500

1613057EDU

1% TAE Mini ReadyAgarose Precast Gel, 7.1 x 10 cm, 2 x 8-well1613015EDU

1% TAE Mini ReadyAgarose Precast Gel, 7.1 x 10 cm, 8-wellEquipment
Catalog #Description

1660551EDUClassroom Adjustable-Volume Digital Micropipet, 2–20 µl

1660512EDU

Fixed-Volume Micropipet, 10 µl

1660506EDU

Professional Adjustable-Volume Digital Micropipet, 2–20 µl

1660507EDU

Professional Adjustable-Volume Digital Micropipet, 20–200 µl

1660531EDUUView Mini Transilluminator

1645050EDUPowerPac Basic Power Supply

1664000EDU

Mini-Sub Cell GT Cell Tank and Lid

1660481EDU

Green Racks, pkg of 521

Appendix B

Fast Gel Protocol and Electrophoresis Preparation Instructions
Fast Gel Protocol
There are multiple ways to hasten visualization of DNA bands on an agarose gel using modified conditions and alternate reagents.
Options, as well as the required materials and protocols, are provided below.
1. Cast 1% agarose gels with 1x TAE buffer.
Note: you may also use precast agarose gels (for example, 1613015EDU or 1613057EDU).
2. Prepare 0.25x TAE electrophoresis buffer.
3. Load samples, run gel using conditions in the table below, and visualize DNA using one of the stain options below.
Electrophoresis Buffer and Voltage

Electrophoresis Time

0.25x TAE at 300 V*

10 min0.25x TAE at 200 V

20 min1x TAE at 100 V

30 min* Requires power supply capable of voltages over 200 V, such as the PowerPac Basic Power Supply (1645050EDU).

Fast Blast DNA Stain: Prepare DNA samples before electrophoresis with 5x Orange G Loading Dye. After electrophoresis, stain
gels with Fast Blast DNA Stain and visualize the next day.
UView 6x Loading Dye and Stain: Prepare DNA samples for electrophoresis using 6x UView Loading Dye and Stain. After gel
electrophoresis, visualize instantly with a UV transilluminator or a handheld UV lamp in the dark.
Preparing Agarose Gels
For each class of eight workstations, prepare either (i) eight 7 x 7 cm gels with one 8-well comb, or (ii) four 7 x 10 cm gels with two
8-well combs. Use the volumes and quantities of reagents shown in the table below.
Volumes and Quantities of Reagents for Agarose Gels.
Number of Gels

1

4

8

16

Purified water, ml

39

156

312

624

50x TAE, ml

0.8

3.2

6.4

12.8

Agarose, g

0.4

1.6

3.2

6.4

Total, volume of molten agarose, ml

40

160

320

640

Purified water, ml

49

196

392

784

50x TAE, ml

1.0

4.0

8.0

16.0

Agarose, g

0.5

2.0

4.0

8.0

Total, volume of molten agarose, ml

50

200

400

800

1% TAE Agarose Gel (7 x 7 cm) — serves one workstation

1% TAE Agarose Gel (7 x 10 cm) — serves two workstations

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22

Appendix B
Prepare Molten Agarose
1. Add the appropriate amount of agarose powder and then the liquids to a suitable container; fill to less than 75% of thecontainer volume. Swirl to mix.
Note: If using an Erlenmeyer flask, place a small, inverted 25 ml flask over the opening to minimize evaporation. If using abottle, loosen the cap to allow air and steam to escape.
2. Place the agarose solution into the microwave. Microwave for 3 min. Continue to boil in 30 sec increments until the solutionboils and all agarose has dissolved.
Caution: Always wear heat-protective gloves, goggles, and a lab coat while preparing agarose gels. Hot molten agarose cancause severe burns.
3. Let the agarose cool to 60°C before pouring the gels.
Cast Agarose Gels
There are a variety of ways to cast agarose gels. This section outlines the tape method. Consult the instruction manual for yourhorizontal electrophoresis system for alternate methods.
1. Firmly seal the ends of a gel tray with standard laboratory or masking tape (not regular sticky tape).
2. Place the comb into the appropriate slot in the gel tray. If pouring a double-well gel, place a comb at one end of the tray andanother in the middle.
3. Once the molten agarose has cooled to at least 60°C, pour enough agarose to cover ¾ of the way up the gel comb teeth or toa depth of 0.5–0.75 cm.
4. Allow the gel to solidify at room temperature for 10–20 min; it will be opaque when ready to use.
5. Carefully lift the combs straight up to remove them. Carefully remove the tape.
6. Store gels in a sealable plastic bag with a few ml of buffer at room temperature for up to 1 day, or in the refrigerator for up to 1 week.
Preparing TAE Buffer
Though 1x TAE buffer is usually used both for gel casting and as running buffer, the electrophoresis time can be reduced by running
the gels with 0.25x TAE buffer at 200 or 300 V. When using this faster protocol, gels should still be cast using 1x TAE buffer.
You will need 300 ml buffer for each gel. Combine distilled water with the volume of 50x TAE buffer indicated and mix well.
Volumes and Quantities of Reagents for Agarose Gels.
Number of Electrophoresis Chambers

1

4

8

16

274

1,094

2,189

4,378

0.25x TAE Buffer
Purified water, ml
50x TAE, ml

1.4

5.6

11

22

Total, volume of 0.25x TAE buffer, ml

275

1,100

2,200

4,400

Note: If you are reusing 0.25x TAE buffer between classes, ensure the buffer is at or below room temperature before reuse. If thebuffer starts out warm, it may become hot enough to melt the agarose gel during a high-voltage run.

23

Appendix B
Visualizing DNA
UView 6x Loading Dye and Stain
No additional preparation is needed when using UView 6x Loading Dye and Stain.
• Replace Orange G Loading Dye with UView 6x Loading Dye and Stain when preparing DNA samples• Directly after electrophoresis, carefully place gels on a UV transilluminator, lower UV shield, and turn on UV light to visualize• Cell phones can be used to capture gel images: Turn off the flash and place transilluminator in a dark room or area
Fast Blast DNA Stain
1. Prepare 100x Fast Blast DNA Stain according to volumes in the table below.
100x Fast Blast DNA Stain Preparation.
Number of 7 x 10 cm Gels to Stain

1

4

8

16

500x Fast Blast DNA Stain, ml

10

40

100

200

Distilled water, ml

40

160

400

800

Total volume of 100x Fast Blast DNA Stain, ml

50

200

500

1,000

2. Add ~50 ml of 100x stain per gel in a gel staining tray and gently rock for 2–3 min.
3. After 2 min, pour off stain and retain for future use.5 Staining longer will increase background.
4. Rinse gel with tap water for 30 sec to 1 min or until all surface stain is removed.
5. Cover the gel with a large volume of tap water and gently rock to destain. DNA will be visible as dark blue bands against alighter blue background after a few hours, with contrast gradually increasing overnight. Destain overnight for best results.
Note: Using 100x Fast Blast DNA stain prevents the small DNA fragments from diffusing in the gel during an overnight destaining.

5

100x Fast Blast DNA Stain can be reused at least six times.

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24

Appendix C

Restaurant Layout
Several independent air conditioners were running in the room, which created isolated and recirculating streams of air. The
windows to the outdoors were closed. Note no one was seated at Table D.
Windows2

1

WaiterA

D

W1

Waiter

Ent
ran
ce

Cook

W2

K1

3

4
3

2

3
1

4

4

2

B

E

6m

5

7

6

8

5

1

1

2

1

6

3

C

2
7

F
5

4

6

3

5

4

Air conditioner

Air conditioner
5m

25

Appendix C

Restaurant Staff and Patron Cards
Patient Background

Patient Background

Name: Tav (male) 26

Name: Aidan (male) 27

Seat: A1

Seat: A2

Back story: Roommate of A2. Lives in aneighboring town. Met friends for lunch.

Back story: Roomate with A1. Lives in aneighboring town. Met friends for lunch.

Restroom: N/A

Restroom: 12:48 pm

Food at restaurant: Onion rings, mushroom sandwich, soft drink

Food at restaurant: Clam chowder, onion rings, iced tea

Patient Background

Patient Background

Name: Ezri (female) 28

Name: Madison (female) 26

Seat: A3

Seat: A4

Back story: Friend of A1 and A2.
Lives nearby. Met friends for lunch.

Back story: Friend of A1–A3.
Lives nearby. Met friends for lunch.

Restroom: N/A

Restroom: N/A

Food at restaurant: Raw oysters, mushroom sandwich, water

Food at restaurant: Fish tacos, french fries, soft drink

Patient Background

Patient Background

Name: Connor (male) 68

Name: Arabella (female) 65

Seat: B1

Seat: B2

Back story: Father of B3.
Lives nearby. Met friends for lunch.

Back story: Mother of B3.
Lives nearby. Met friends for lunch.

Restroom: 12:31 pm

Restroom: N/A

Food at restaurant: Clam chowder, garlic pasta, hot tea

Food at restaurant: Shared raw oysters with B3, pasta, iced tea

Patient Background

Patient Background

Name: Zach (male) 23

Name: Maciah (male) 39

Seat: B3

Seat: B4

Back story: In town visiting family.
Lives in another region. Arrived in town on June 2.

Back story: Brother of B3, married to B5.
Lives in the region, but not nearby.

Restroom: N/A

Restroom: N/A

Food at restaurant: Shared raw oysters with B2, fish tacos,
onion rings, iced tea

Food at restaurant: Shrimp cocktail, cheeseburger, soft drink

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Appendix C

Patient Background

Patient Background

Name: Riley (female) 38

Name: Kaley (female) 10

Seat: B5

Seat: B6

Back story: Spouse of B4.
Lives in the region, but not nearby.

Back story: Daughter of B4 and B5.
Lives in the region with her parents, but not nearby.

Restroom: N/A

Restroom: 12:45 pm

Food at restaurant: Vegetable soup, white fish fillet, iced tea

Food at restaurant: Split shrimp cocktail, garlic pasta, lemonade

Patient Background

Patient Background

Name: Camielle (female) 66

Name: Jake (male) 66

Seat: B7

Seat: B8

Back story: Aunt of B3 and sister of B1.
Lives nearby.

Back story: Uncle of B3 andbrother-in-law of B1. Lives nearby.

Restroom: N/A

Restroom: N/A

Food at restaurant: Garden salad, clam chowder, hot tea

Food at restaurant: Fried oyster sandwich, soft drink

Patient Background

Patient Background

Name: DaeLynn (female) 69

Name: Matteo (male) 14

Seat: C1

Seat: C2Back story: Mother of of C4 and C5.
Lives with C3 in a neighboring region. Visiting family.

Back story: Child of C5. Lives with C5.
Restroom: 1:08 pm

Restroom: 12:15 pm
Food at restaurant: Fish tacos, french fries, lemonade
Food at restaurant: Shrimp cocktail, mushroom sandwich, water

Patient Background

Patient Background

Name: Jackson (male) 70

Name: Devlin (female) 42

Seat: C3

Seat: C4

Back story: Grandfather. Lives with C1
in a neighboring region. Visiting family.

Back story: Sister of C5. Lives nearby.
Restroom: N/A

Restroom: N/A
Food at restaurant: Fried oyster sandwich, seasoned frenchfries, water

27

Food at restaurant: Calamari, garden salad, mushroom sandwich,
iced tea

Appendix C

Patient Background

Patient Background

Name: Hollis (female) 45

Name: Conway (male) 17

Seat: C5

Seat: C6

Back story: Mother of C2 and C6.
Lives nearby.

Back story: Child of C5. Lives with C5.
Restroom: N/A

Restroom: 12:20 pm
Food at restaurant: Cheeseburger, seasoned fries, lemonade
Food at restaurant: Calamari, roast chicken, hot tea

Patient Background

Patient Background

Name: Delaney (female) 47

Name: Tim (male) 54

Seat: C7

Seat: E1

Back story: Friend of C5.
Lives in a neighboring town.

Back story: Works nearby with everyone
at table E. Lives in a different region. Eating before a work function.

Restroom: 12:36 pm

Restroom: N/A

Food at restaurant: Garden salad, tuna sandwich, iced tea

Food at restaurant: Vegetable soup, roast chicken, iced tea

Patient Background

Patient Background

Name: Nick (male) 36

Name: Kiki (female) 42

Seat: E2

Seat: E3

Back story: Works nearby with everyone
at table E. Lives in neighboring town. Eating before a work function.

Back story: Works nearby with everyone
at table E. Lives nearby. Eating before a work function.

Restroom: 12:53 pm

Restroom: N/A

Food at restaurant: Clam chowder, french fries, hot tea

Food at restaurant: Shrimp cocktail, hot pastrami sandwich

Patient Background

Patient Background

Name: Savannah (female) 43

Name: Neenah (female) 38

Seat: E4

Seat: E5

Back story: Works nearby with everyone
at table E. Lives in a different region. Eating before a work function.

Back story: Works nearby with everyone
at table E. Lives in a neighboring town. Eating before a work function.

Restroom: 12:09 pm

Restroom: N/A

Food at restaurant: Raw oysters, mushroom sandwich, soft drink

Food at restaurant: Vegetable soup, hot pastrami sandwich, water

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Appendix C

Patient Background

Patient Background

Name: Ricardo (male) 17

Name: Alex (male) 17

Seat: F1

Seat: F2

Back story: Attends a school near therestaurant. Friends with F2–F6. Lives nearby. Celebrating thebirthday of F3. Ate a cupcake from F2.

Back story: Attends a school near therestaurant. Friends with all at table F. Lives nearby. Celebrating thebirthday of F3. Brought homemade cupcakes from home to share.
Ate one himself.

Restroom: N/A

Restroom: 12:26 pm

Food at restaurant: Tuna sandwich, soft drink

Food at restaurant: Cheeseburger, split an order of onion rings,
soft drink

Patient Background

Patient Background

Name: Anna (female) 18

Name: Stefan (male) 17

Seat: F3

Seat: F4

Back story: Attends a school near therestaurant. Friends with all at table F. Lives nearby. Celebrating
her birthday with friends. Ate a cupcake from F2.

Back story: Celebrating the birthday of F3.
Ate a cupcake from F2.
Restroom: 1:03 pm

Restroom: N/A
Food at restaurant: Vegetable soup, split onion rings, water
Food at restaurant: Fish tacos, split an order of onion rings,
lemonade

29

Patient Background

Patient Background

Name: Naija (female) 17

Name: Brooklyn (female) 18

Seat: F5

Seat: F6

Back story: Attends a school near therestaurant. Friends with all at table F. Lives nearby. Celebrating thebirthday of F3. Ate a cupcake from F2.

Back story: Attends a school near therestaurant. Friends with all at table F. Lives nearby. Celebrating thebirthday of F3. Ate a cupcake from F2.

Restroom: N/A

Restroom: N/A

Food at restaurant: Garden salad, split an order of fries,
lemonade

Food at restaurant: Vegetable soup, split an order of fries, water

Patient Background

Patient Background

Name: Elijah (male) 23

Name: Dannie (female) 24

Seat: W1

Seat: W2

Back story: Arrived at 11 am.
Worked lunch shift, primarily serving tables E and F. Began feelingsick midway through the shift and went home at 1:15 pm.

Back story: Arrived at at 10:45 am for an
11:30 am open. Prepped dining room. Worked all day, primarily
serving tables A, B, and C.

Restroom: N/A

Restroom: 1:30 pm, 4:26 pm

Food at restaurant: N/A

Food at restaurant: N/AAppendix C

Patient Background
Name: Anthony (male) 33
Seat: K1
Back story: Arrived at work at 8:30 am.
Received food shipment. Prepped ingredients. Cooked all meals.
Restroom: 9:30 am, 11 am, 4:56 pm
Food at restaurant: N/A

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Appendix D

Resources
Organization

Link/ReferenceType of Resource

Description

Bio-Radbio-rad.com/vdtkit

Supplementary
documentation

Instructor Guide, Student Guide,bulletins

bio-rad.com/classroomresources

Supplementary
documentation

YouTube playlists, supplementaryresources

Answer Guide (included with kit)

Supplementary
documentation

Answers to Focus Questions

CDC NERD Academy Student Quick Learn: How Does
Disease Spread? (youtu.be/1QLgXzyXOH0)

Video

Describes how infection prevention
specialists use information on thetype of infectious agent, how it isspread, and common ways it exits
and enters its hosts to create achain of infection model

Lesson 1: Introduction to Epidemiology, Section 10:
Chain of infection(cdc.gov/csels/dsepd/ss1978/lesson1/section10.html)

WebsiteDescribes the chain of infection

COVID-19 Outbreak Associated with Air Conditioning in
Restaurant, Guangzhou, China, 2020

WebsiteDescribes the transmission case andrestaurant layout upon which these
kit activities are based

Viruses: Molecular Hijackers (youtu.be/wUgEhfo_qxU)

Video

Describes viruses, what they are andhow they cause disease

Introduction to Virology and Viral Classification(youtu.be/lADC0C_WeH0)

Video

Describes viruses, what they are,how they were discovered, variationsin nature

Routes of Viral Transmission(youtu.be/S5egQXgBZ6c)

Video

Describes various modes and routesof transmission

Stevens, H (2020)

Website withsimulator

Offers a deeper description of theepidemic curve

Web simulation

Describes several settings foroutbreaks studied by healthauthorities with conclusions
that offer insights into mitigation
strategies

Centers for
Disease Control
and Prevention(CDC)

wwwnc.cdc.gov/eid/article/26/7/20-0764-f1
Professor Dave
Explains

Washington Post

Why outbreaks like coronavirus spread exponentially, andhow to “flatten the curve”(www.washingtonpost.com/graphics/2020/world/corona-simulator/)
El PaisSalas J and Jafra M (2020)
An analysis of three Covid-19 outbreaks: how they
happened and how they can be avoided(elpais.com/especiales/coronavirus-covid-19/an-analysisof-three-covid-19-outbreaks)

31

Appendix D
Bibliography
Cassedy A, et al. (2021). Virus detection: A review of the current and emerging molecular and immunological methods, Front Mol
Biosci 8: 637559.
Hardstaff JL, et al. (2018). Foodborne and food-handler norovirus outbreaks: A systematic review, Foodborne Pathog Dis 15:
589–597.
Jayaweera M, et al. (2020). Transmission of COVID-19 virus by droplets and aerosols: A critical review on the unresolved
dichotomy, Environ Res 188: 109819.
Li Y, et al. (2021). Probable airborne transmission of SARS-CoV-2 in a poorly ventilated restaurant, Build Env 196, 107788.
Louten J (2021). Virus transmission and epidemiology, Essential Human Virology 2016: 71, 92.

References
Dance A (2021). The incredible diversity of viruses, Nature 595: 22–25.
Pride D (2020). Viruses can help us as well as harm us, Sci Amer 323, 46–53.
Lu J, et al. (2020). COVID-19 outbreak associated with air conditioning in restaurant, Guangzhou, China, 2020, Emerg Infect Dis
26, 1628-1631.
Sherwood D (2020). How a Chilean raspberry scam dodged food safety controls from China to Canada, Reuters 6 October 2020.

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