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AOHS Biotechnology

Lesson 8Bacterial Transformation

Students explore social and ethical concerns regarding recombinant DNA by learning about genetically modified foods. They also carry out a survey to find out people’s opinions about these products. Students acquire biotechnology laboratory skills by transforming E. coli bacteria with a recombinant plasmid.

Advance Preparation In Class Period 1, you will need access to the video “The Eyes of Nye – Genetically Modified

Foods” at http://www.youtube.com/watch?v=GKm2Ch3-Myg . If your school does not allow access to YouTube, you may wish to download the video to your

computer in advance using KeepVid (see www.keepvid.com) or a similar program. Gather all equipment and materials for the lab in this lesson according to Lab Resource 8.1,

Procedure: GFP Transformation Experiment. Follow the instructions to prepare for the lab according to Teacher Resource 8.3, Supplement: GFP Transformation Experiment. The lab occurs in Class Periods 2 through 5.

The amount of the time between Class Periods 2 and 3 and between Class Periods 3 and 4 must be sufficient to allow for the optimum growth of E. coli on petri dishes, which depends on the incubation temperature. At 37°C (98.6°F), the bacteria will be ready in 24‒36 hours; at 28°C (82°F), the bacteria will be ready in 48 hours; and at 21°C (70°F), the bacteria will be ready in 72 hours.

In Class Periods 4 and 5, students will need access to computers with word processing software and for Internet research (one computer per pair of students) as well as a printer.

Prior to Class Period 6, gather magazines with pictures of farms and/or food materials that students can use as visuals for their genetically modified food poster (optional).

This lesson is expected to take 6 class periods.

Lesson FrameworkLearning ObjectivesEach student will:

Summarize the steps in a bacterial transformation, including competency, recovery, and selection* Create recombinant E. coli

Copyright © 2014‒2016 NAF. All rights reserved.

http://www.keepvid.com/

http://www.youtube.com/watch?v=GKm2Ch3-Myg

AOHS BiotechnologyLesson 8 Bacterial Transformation

Demonstrate how transformants may be selected, including antibiotic resistance Evaluate the validity of results obtained during experimentation and compose a concluding

statement addressing possible sources of error and potential applications Explain key bioethics questions surrounding the application of recombinant DNA, including the

controversy associated with genetically modified crops*This is one of the 16 key learning objectives assessed by the NAFTrack Certification end-of-course exam for this course.

Academic Standards Apply the fundamentals of biochemistry, cell biology, genetics, mathematical concepts,

microbiology, molecular biology, organic chemistry and statistics to conduct effective biotechnology research and development of products (CCTC Standards HL-BRD, 2)

Demonstrate basic knowledge of recombinant DNA, genetic engineering, bioprocessing, monoclonal antibody production, nanotechnology, bioinformatics, genomics, proteomics and transcriptomics to conduct biotechnology research and development (CCTC Standards HL-BRD, 3)

Construct an explanation based on evidence for how the structure of DNA determines the structure of proteins, which carry out the essential functions of life through systems of specialized cells (Next Generation Science Standards 2013, HS-LS1-1, From Molecules to Organisms: Structures and Processes)

Demonstrate the principles of solution preparation, sterile techniques, contamination control, and measurement and calibration of instruments used in biotechnology research (CCTC Standards HL-BRD, 4)

Summarize and explain the larger ethical, moral, and legal issues related to biotechnology research, product development and use in society (CCTC Standards HL-BRD, 6)

Practice speaking and active listening skills (National Health Science Standards, 2.15) Apply personal safety procedures based on Occupational Safety and Health Administration

(OSHA) and Centers for Disease Control (CDC) regulations (National Health Science Standards 2015, 7.21)

Apply safety techniques in the work environment (National Health Science Standards 2015, 7.31) Observe all safety standards related to the Occupational Exposure to Hazardous Chemicals

Standard (Safety Date Sheets [SDSs] (National Health Science Standards 2015, 7.41) Comply with safety signs, symbols, and labels (National Health Science Standards 2015, 7.42) Evaluate roles and responsibilities of team members (National Health Science Standards 2015,

8.11)

AssessmentAssessment Product Means of Assessment

GFP transformation experiment conclusion (Student Resource 8.5)

Rubric: GFP Transformation Experiment Conclusion (Teacher Resource 8.4)

Genetically modified food poster (Student Resource 8.7)

Assessment Criteria: Genetically Modified Food Poster (Teacher Resource 8.5)

Copyright © 2014‒2016 NAF. All rights reserved. 2


Prerequisites Ability to measure accurately using micropipets Knowledge of bacteria culture technique Knowledge of sterile technique

Instructional Materials Lab Resource

Lab Resource 8.1, Procedure: GFP Transformation Experiment

Teacher Resources Teacher Resource 8.1, Answer Key: “The Eyes of Nye” Video Guide Teacher Resource 8.2, Presentation and Notes: Bacterial Transformation (includes separate

PowerPoint file) Teacher Resource 8.3, Supplement: GFP Transformation Experiment Teacher Resource 8.4, Rubric: GFP Transformation Experiment Conclusion Teacher Resource 8.5, Assessment Criteria: Genetically Modified Food Poster Teacher Resource 8.6, Key Vocabulary: Bacterial Transformation Teacher Resource 8.7, Bibliography: Bacterial Transformation

Student Resources Student Resource 8.1, Video Guide: “The Eyes of Nye” Student Resource 8.2, Survey Questions: Genetically Modified Food Student Resource 8.3, Survey: Genetically Modified Food Student Resource 8.4, Reading: Bacterial Transformation Student Resource 8.5, Lab Conclusion: GFP Transformation Experiment, Part 4 Student Resource 8.6, Opinions: Genetically Modified Food Student Resource 8.7, Poster: Genetically Modified Food

Equipment and Supplies LCD projector and computer for PowerPoint presentation An up-to-date video player on a computer ready to show a YouTube video Computers with word processing software and Internet access (one per pair of students) and a

printer Whiteboard, blackboard, or flip chart Spray disinfectant Equipment and materials for the lab as listed in Lab Resource 8.1, Procedure: GFP

Transformation Experiment A camera for students to record photographic evidence of their experiments Glue stick, one per four students, or tape



Scissors, one per student Small containers, eight, to hold the results from each of the eight survey questions Markers, one set per four students Chart paper, one sheet per four students Magazines with images of farms and/or food for posters (optional)

Lesson StepsStep Min. Activity

CLASS PERIOD 1

1 25 Video: “The Eyes of Nye”Students watch a video to learn more about genetically modified foods and the controversies associated with them.

Prior to showing the video, refer students to Student Resource 8.1, Video Guide: “The Eyes of Nye,” and read through the questions to familiarize students with the information they need to gather while watching it. Project the video, which is available at http://www.youtube.com/watch?v=GKm2Ch3-Myg .

If your school does not allow access to YouTube, you may wish to download the video to your computer in advance using KeepVid (see keepvid.com) or a similar program.

When the video is over, call on students to share their responses to the questions on Student Resource 8.1. Have Teacher Resource 8.1, Answer Key: “The Eyes of Nye” Video Guide, available as you listen to students’ responses.

In particular, have students discuss Question 7, “Did anything in this video sway your opinion about genetically modified food safety or labeling?” Have students articulate how and why their opinions have changed.

Close the activity by explaining that there is a range of opinions about the value of genetically modified foods and what impact they might have on people and the environment.

2 25 Survey Development: Genetically Modified FoodsIn this activity, students write survey questions aimed at uncovering beliefs about topics related to genetically modified foods.

Refer students to Student Resource 8.2, Survey Questions: Genetically Modified Food. Organize students into groups of three or four and tell the groups to choose two topics


http://www.youtube.com/watch?v=GKm2Ch3-Myg


Step Min. Activity

about genetically modified food from the list provided and draft one survey question about each topic. Briefly explain that a question can be designed in one of three types of response formats: short response, multiple choice, or an agree–disagree rating scale from 1 to 5.

Circulate around the room as students work on their questions. After 10 minutes, call on groups to share one of their two questions with the class. Record questions on the board and group them according to topic. (Topics are the value of GM foods, the impact of GM plants on the environment, how widespread GM foods are, the impact of GM plants on small farmers, safety of GM foods, how well GM plants and foods are regulated, the need for labeling GM foods, and how GM plants differ from plants developed by traditional breeding.)

Select eight of the students’ questions to use in the survey, one for each of the eight topics. Refer students to Student Resource 8.3, Survey: Genetically Modified Food, and have them record the eight survey questions on the resource. (Note: If the resource is printed out double-sided, two sets of data can be collected on one sheet of paper. This will allow students to cut out survey responses more easily when they review the data in Class Period 5.)

Tell students that for homework they will ask one peer and three adults (this can include parents and teachers) the eight survey questions. Encourage students to make an effort to interview adults that other students are not likely to interview―in other words, to try to ask family members, neighbors, or family friends rather than relying solely on teachers at the school whom other students might want to survey. The students’ goal should be to interview four people who have not been interviewed by anyone else for this survey. Students are required to turn in the data they collect from the survey in Class Period 5.

Explain that the survey will help students learn about public opinion on topics related to genetically modified foods.

CLASS PERIOD 2

3 25 Presentation: Bacterial TransformationThis activity gives students an in-depth look at the steps involved in bacterial transformation. It also develops students’ listening and note-taking skills.

To prepare, make notes to guide class discussion using Teacher Resource 8.2, Presentation Notes: Bacterial Transformation. Have Teacher Resource 8.2, Presentation: Bacterial Transformation (separate PowerPoint file), ready to show as a full-screen slideshow using an LCD projector.

Post the following list of topics on the board or on a sheet of chart paper:

The steps of the transformation experiment The two samples of bacteria they will prepare The conditions under which each sample will be grown

Ask students to take notes on these topics in their notebook as they view the presentation.

Present the slideshow. Use the notes you prepared previously and the questions on the



Step Min. Activity

slides to encourage class discussion.

This presentation is duplicated as Student Resource 8.4, Reading: Bacterial Transformation. Students can read the presentation, answer the discussion questions in their notebook, and discuss their answers as a class. This student resource is also useful for review.

After the presentation, divide the class into pairs or groups of three and have them compare their notes for completeness and accuracy by referring to Student Resource 8.4. Invite each pair or triad to share their thoughts and questions.

To conclude, explain that the bacteria cell is able to make a protein from the plasmid gene because the DNA code is highly similar among all living organisms. Let students know they will be doing an experiment with this type of work during this class period (and for the rest of this lesson).

4 25 Lab: GFP Transformation Experiment, Part 1In this activity, students prepare for the transformation experiment by streaking petri dishes with bacteria that will grow into colonies for use in the transformation. Students also develop hypotheses for the growth of the two samples of bacteria under different conditions. The activity focuses on the following college and career skill:

Locating, evaluating, and applying information

Before class, prepare for the lab by following the instructions in Teacher Resource 8.3, Supplement: GFP Transformation Experiment. The equipment list and instructions for the lab are located in Lab Resource 8.1, Procedure: GFP Transformation Experiment.

Refer students to Part 1 of Lab Resource 8.1. Organize students into groups. Review how to streak the petri dish and explain how to avoid contamination. Remind students that streaking is a technique used to isolate a single bacterial cell that will then grow into a colony and that colonies are therefore “clonal.”

Instruct students to complete Steps 1 and 2, and then to streak starter dishes for the experiment according to Steps 3‒8. When students have finished streaking the dishes and placed them in the incubator (Step 10), have students review the pGLO plasmid map, which is reproduced in Step 12. Tell students they may also want to review Student Resource 8.4, Reading: Bacterial Transformation, as a reference.

Ask students to explain the sequences on the pGLO plasmid map. Assess if they know that GFP is the green fluorescent protein, bla is a protein that makes the bacteria resistant to ampicillin, and araC is a protein that regulates the transcription from the pBAD promoter which in turn drives the production of GFP, ensuring that bacteria only make it when fed arabinose.

Have students complete Steps 13‒15 of Lab Resource 8.1. Students will propose hypotheses about the growth of the transformation bacteria and the negative control bacteria. Again tell students they may want to review Student Resource 8.4, Reading: Bacterial Transformation.

Call on several students to share their hypotheses and explanations with the class.

Conclude by explaining that using antibiotic resistance to select cells is a widespread practice in biotechnology and biology research. Let students know they will be continuing the experiment in the next class period.



Step Min. Activity

Remind students that they should be gathering their survey results for homework. All survey results need to be completed by Class Period 5.

CLASS PERIOD 3

5 50 Lab: GFP Transformation Experiment, Part 2Students carry out a transformation of bacteria with the pGLO recombinant plasmid. The activity focuses on the following college and career skills:

Utilizing time efficiently when managing complex tasks Effectively using technology relevant to a profession

Refer students to Lab Resource 8.1, Procedure: GFP Transformation Experiment, Part 2. Point out the materials that students will need and briefly summarize the steps of the transformation experiment, including the preparation and treatment of the two bacteria samples. When students begin the experiment, circulate around the room to assist students as needed.

At the end of the activity, describe some of the potential uses of fluorescent proteins, which are enabling biologists to visualize biological processes such as protein synthesis, regulation of gene expression, cell movement, cell fate during development, and the formation of different organs. Explain that students are using GFP as a marker to identify a transgenic organism, which is the bacteria they are transforming.

Let students know that in the next class period they will be examining and analyzing their colonies.

Remind students that they should be gathering their survey results for homework. All survey results need to be completed by Class Period 5.

CLASS PERIOD 4

6 15 Lab: GFP Transformation Experiment, Part 3Students examine and analyze experimental data, recording the results through drawings or photographs so they can refer back to these images when they work on the experiment conclusion.

Refer students to the first part of Lab Resource 8.1, Procedure: GFP Transformation Experiment, Part 3. Students examine the bacterial growth on their petri dishes and analyze the success of their bacterial transformation experiment. Students can record results on Lab Resource 8.1 by making a drawing; alternatively, a camera can be used to take pictures of the dishes.

Let students know that next they will analyze their transformation experiment.

7 15 Lab: GFP Transformation Experiment, Part 4Students analyze their colony observations and draw scientific conclusions based on



Step Min. Activity

their data about whether the experiment was successful and what they learned from each petri dish. Students also consider possible sources of error and applications of the results in order to prepare for writing their lab conclusion report.

Refer students to Student Resource 8.5, Lab Conclusion: GFP Transformation Experiment, Part 4. Have students work in their lab groups to analyze what they learned using the drawings or photographs of their colonies.

Ask students to explain how they were able to determine if their transformation experiment was successful. Then address common errors that can lead to inaccurate or missing results, such as contamination, improperly prepared media, and mistakes in the procedure steps.

Finally, instruct students to work individually to answer the remaining questions in Student Resource 8.5 and to make notes for the results and conclusion sections for the experiment. Remind students to refer to the rubric at the end of Student Resource 8.5.

Tell students that analyzing data and summarizing what they learned from an experiment are important lab skills. Tell students that next they will begin working on their written lab conclusion report.

8 20 Lab: GFP Transformation Experiment, Part 4 (Continued)Students complete the GFP transformation experiment by writing their report on the conclusion section. This activity focuses on the following college and career skill:

Demonstrating the ability to write effectively

Assign each pair of students to a computer and have pairs work together to type their lab report. Remind students to use their notes and the rubric in Student Resource 8.5, Conclusion: GFP Transformation Experiment Part 4. Tell students that they will begin typing up the report in this class period and complete it in the next class period.

Students can alternate typing and reviewing sections. Walk around the room while students work to help answer any questions and ensure students are following directions. Remind students to leave enough space in the report to add the drawings or photographs of their colonies after they print out their reports.

Tell students that communicating experimental results effectively is an important skill.

Remind students that they should be gathering their survey results for homework. All survey results need to be completed by Class Period 5, the next class period.

CLASS PERIOD 5

9 15 Lab: GFP Transformation Lab, Part 4 (Continued)Students complete the GFP transformation experiment by writing their conclusion section. This activity focuses on the following college and career skill:

Demonstrating the ability to write effectively

Assign each pair of students to a computer for this activity and have them work together to complete their reports. Remind them to use their notes and the rubric in Student



Step Min. Activity

Resource 8.5, Conclusion: GFP Transformation Experiment, Part 4.

Have students print out their lab reports and then add the drawings or photographs of their colonies. Distribute glue sticks to each student for this task. Assess the reports using Teacher Resource 8.4, Rubric: GFP Transformation Experiment Conclusion.

Finish the activity by asking students to talk about whether the process of writing a conclusion section has gotten easier with practice.

10 35 Research: Opinions on Genetically Modified FoodStudents gather information about public opinion, expert opinion, and their own opinions on a topic related to genetically modified foods. This information will be used to make a poster on GM foods. The activity focuses on the following college and career skills:

Demonstrating teamwork and collaboration Locating, evaluating, and applying information

Tell students that they will work in groups to research the range of opinions on genetically modified foods. Each group will focus on a genetically modified food topic that is associated with one of the survey questions, and they will prepare a poster to summarize and share what they learned.

Tell students that in this activity they will gather information about different opinions on their topic. Their first step is to cut up their survey results so that each group can look at one specific question. Distribute scissors to each student and set out eight containers, labeled 1 through 8. Have students cut up their survey results and put them in the appropriate container.

Divide students into groups of four and assign each group one of the eight questions (and its corresponding topic). Then hand out the appropriate container to each group.

Refer students to Student Resource 8.6, Opinions: Genetically Modified Food. Explain that there are three sections of this assignment. The first section, Public Opinion, relies on the survey results for their topic. The second section, Expert Opinion, involves doing research using the websites suggested in the resource. The third section, Personal Opinion, is about each student’s personal opinion.

Students will need to divide up the work for the first two sections. Make recommendations about how you think students should split it up. For example, some groups might have two students work on the survey results and two students work on the research; other groups might put three students on the research and leave one to handle the survey results. Point out that students have only the rest of this class period to work on their research and survey results, so they need to use their time wisely.

While students work, circulate around the room and help them as needed.

When approximately 10 minutes remain in class, ask students to rejoin their groups to discuss and summarize what they learned.

Depending on the time available, have students complete their personal opinion in class or for homework. Tell students they will need to bring the completed resource to the next class period. You may also wish to encourage students to bring in images to use for their posters, either from magazines or pictures they printed off the Internet. This can make the following class period flow more smoothly.



Step Min. Activity

If time permits, you may wish to ask several students to share what they learned from their research or from the survey results.

CLASS PERIOD 6

11 50 Poster: Genetically Modified FoodsNow that students have organized their survey results and research, they think about how to communicate what they have learned about their topic in a poster format.

Ask students to sit with their poster group. Refer them to Student Resource 8.7, Poster: Genetically Modified Food. Review the assessment criteria at the end of the resource and answer any questions. Point out that students have only this class period to work on their poster.

Tell students to use Student Resource 8.7 to prepare for making their genetically modified food poster. Students will decide on a title, a visual, and a layout for their poster and record their decisions on the resource before making their poster. Each group needs to fill out only one student resource. The visuals on the poster can be student drawings, images they found for homework (if any), or materials you provide in class (if any).

When students have finishing planning, ask them to begin working on their posters. Have students refer to the information they collected on Student Resource 8.6, Opinions: Genetically Modified Food, and distribute poster paper and markers to each group. You will also need to provide scissors and tape or glue so students can attach their images.

Five minutes before the end of the period, have each group share their poster before displaying them in a common area. Assess the posters using Teacher Resource 8.5, Assessment Criteria: Genetically Modified Food Poster.

Close the activity by explaining that advances in technology usually affect society more directly than science does because technology addresses specific and immediate problems and needs. One role of a citizen in a technological society is to weigh the benefits and risks of different technologies in light of the common good. To do this, it is important to learn how to recognize both valid and unfounded claims.

ExtensionsContent Enrichment

Have students research and summarize the reasons for Food and Chemical Toxicology’s retraction of the study “Long term toxicity of a Roundup herbicide and a Roundup-tolerant genetically modified maize" (principal author Gilles-Eric Séralini).

Corn (also called maize) is the largest food crop in the world. Have students read the article “The Evolution of Corn” about corn genetics and how it was selectively bred in the Americas over thousands of years, available at http://learn.genetics.utah.edu/content/selection/corn/. Have students summarize the article.


http://learn.genetics.utah.edu/content/selection/corn/


STEM Integration Math: To compare how well different transformation experiments work, scientists calculate a value

called transformation efficiency. This is the amount of plasmid in micrograms (µg) per bacteria cell transformed. The typical calculation has four steps: o Determine the amount of plasmid in the cell culture by multiplying the concentration of the

plasmid by the amount added to the cell culture.o Determine the ratio of cell culture spread on a petri dish by dividing the volume of cell culture

spread on the petri dish by the entire volume of cell culture (the ratio may be converted to a percentage by multiplying by 100).

o Determine the amount of plasmid spread on the petri dish by multiplying the ratio, or percentage, of cell culture used by the amount of plasmid in the cell culture.

o Determine the transformation efficiency by dividing the amount of plasmid by the number of transformed cells that grew on the petri dish.

Have student groups calculate the efficiency of their transformation experiment. Students will need to review the procedure and their results to carry out the calculation for the two petri dishes that supported the growth of transformed cells. Tell students that the concentration of the plasmid solution they used in the experiment was 0.08 μg/μl. How similar was the efficiency between the two petri dishes inoculated with same amount of transformed bacteria? If the efficiency was not the same, what might have caused the difference?

Technology: If students have Internet access, consider asking them to make their poster using an online graphic design tool like Canva (www.canva.com) or CollageIt (www.collageitfree.com).

Additional Cross-Curricular Ideas Social Studies: Group students into pairs and have them read a specific Native American legend

associated with corn at http://www.native-languages.org/legends-corn.htm. Have each pair act out the legend they read. Conclude with a discussion about the role of legends, and ask students to consider how legends shape perception of the world, how they transmit societal values, and how they support an individual through life stages.


http://www.native-languages.org/legends-corn.htm

http://www.collageitfree.com/

http://www.canva.com/

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