A Teaching Module Integrating Literature and Genetic Engineering

Mari Knutson
Lynden Public High School
1201 Bradley Rd.
Lynden, WA 98264
Summer, 2004

WSU Mentor: Ryan Soderquist, Dr. James Lee
Department of Chemical Engineering
Washington State University
Pullman,WA 99164-2710

National Science Foundation Grant No. EEC-0338868 supports this project.

Introduction

Overview. This module is designed to encourage students to read and to introduce them to the biotechnology and principles of genetic engineering.

A fiction novel (Robin Cook’s “Chromosome Six”), a fictitious short story (“Bill Schwan’s “Ethics of the Prophets”) and a non-fiction news item will be used to pique curiosity and perhaps motivate students to read more about genetic engineering and it’s tremendous impact on society.

Laboratory exercises will include DNA extraction, comparison of chromosome banding patterns, transformation of bacteria using an engineered plasmid, and callus induction from seeds.

Scope. Pieces may be mixed and matched according to time constraints and laboratory accessibility. Activities are designed to be cost-effective and not to require extensive preparation on the part of the student or teacher. If all components are included, the module may be accomplished over six days. If the novel is omitted and just the short story used, the laboratory exercises may be accomplished in four days.

Chemical Engineering. Chemical engineers utilize chemistry to solve problems for industry. This module focuses primarily on the biochemical aspects of engineering. As science and technology seem to be producing information at astonishing speed, the ‘making it work’ is the job of the chemical engineer. The American Institute of Chemical Engineers (AIChE) has compiled a list of the “10 Greatest Achievements of Chemical Engineering”. The two on the list that pertain to this module are:

  1. The Human Reactor – Helping to improve clinical care leading to mechanical wonders such as artificial organs.
  2. Wonder Drugs for the Masses – Today’s low price and high volumes of antibiotics owe their existence to the work of chemical engineers.

When this list was made, the authors would surely have seen the engineering of transgenic organs as science fiction!

To see a summary of each of the ten achievements and to learn more about the field of Chemical Engineering visit:http://www.cems.umn.edu/~aiche_ug/history/h_whatis.html

Chemical Engineering Application and Inspiration. Chemical engineers at Washington State University (WSU) are working on lowering costs in the mass production of desirable proteins. Plant cell suspension cultures are being used as the host to produce recombinant proteins such as GM-CSF which is a human growth factor.

Problem Statement. Genetic engineering has outpaced the comprehension of most citizens. Ethical, legal and moral questions arise as the ‘remotely possible’ becomes probable. Students today will be faced with making decisions about how scientific knowledge is to be used and regulated. An understanding of how our perception of genetic engineering is arrived at is useful to students in determining if their perception has merit. An understanding of the actual principles and technology used to engineer pharmaceuticals, tissues and even new organisms will give students the tools to evaluate new information more critically. By combining fiction, non-fiction, and laboratory experience, students will have the opportunity to integrate literature and science.

The Essential Academic Learning Requirements. When students finish this module they will have achieved many of the items specified by the Washington State “Essential Academic Learning Requirements” (EALRs) and by national standards.

  1. The student understands and uses scientific concepts and principles.
  2. The student conducts scientific investigations to expand understanding of the natural world.
  3. The student applies science knowledge and skills to solve problems or meet challenges.
  4. The student uses effective communication skills and tools to build and demonstrate understanding of science.
  5. The student understands how science knowledge and skills are connected to other subject areas and real-life situations.

Background Information. According to the National Institute of Health, recombinant DNA technology is defined as: a body of techniques for cutting apart and splicing together different pieces of DNA. When segments of foreign DNA are transferred into another cell or organism, the substance for which they code may be produced along with substances coded for by the native genetic material of the cell or organism. Thus, these cells become "factories" for the production of the protein coded for by the inserted DNA.

“Biologics, which include protein hormones, engineered protein-based vaccines, and monoclonal antibodies, can precisely modify a patient's physiology, often with greater success and fewer side effects than traditional small-molecule drugs or vaccines. Indeed, early biologics—Amgen's (Thousand Oaks, CA) recombinant erythropoietin and Genentech's (S. San Francisco, CA) human growth hormone somatropin—have proven that these drugs can benefit huge numbers of patients and generate handsome profits. But biologics are fast becoming victims of their own success, and a looming deficit in biomanufacturing capacity threatens to restrict the expansion of the commercialization of this group of products.

The current standard technology in biomanufacturing, which uses cultured Chinese hamster ovary (CHO) cells in bioreactors, presents major difficulties for companies seeking to scale up. Because nutrients, heat, and gases must diffuse evenly to all cultured cells, the laws of physics set strict limits on the size of bioreactors. Building more bioreactors multiplies costs linearly. A CHO cell–based biomanufacturing plant can cost upwards of $250 million, and an error in estimating demand for, or inaccurately predicting the approval of, a new drug can be incredibly costly. To compound the problem, regulators in the United States and Europe demand that drugs be produced for the market in the same system used to produce them for the final round of clinical trials, so companies have to build facilities for drugs that might not be approved.” (Dove, 2002)

Table 1 shows many of the biotech companies actively pursuing biomanufacturing and the approximate costs.

Alan Dove’s article “Uncorking the biomanufacturing bottleneck” can be viewed in the nature biotechnology archives at http://www.nature.com/nbt/ under August 2002.

A real fear is that microbes or animal cultures will harbor pathogens. Plant cell cultures do not harbor human pathogens and the productions costs are low but keeping genetically modified pollen from being blown to other plants is worrisome.

“Plant cell media are composed of simple sugars and salts and are therefore less expensive and complex than mammalian media. Consequently, purification of secreted protein is simpler and more economical. Additionally, plant cell proteins are likely to be safer than those derived from other systems, since plant cell pathogens are not harmful to humans.” (James and Lee, 2001)

Researchers at WSU are using a four-step process to move desirable genes into plant cells.

“The generation of transgenic plant cells involves 4 basic steps (figure 1). The first step is to subclone the gene of interest into a binary Ti plasmid that contains genes for both antibiotic selection and bacterial propagation of the plasmid and for selection of transgenic plant cells. The second step involves the transformation of the bacterial vector, Agrobacterium tumefaciens, with the newly constructed vector. The third step involves the transformation of a tobacco cell line. With A. tumefaciens and selection of transgenic cells, which grow as clumps of cells (calli) on agar plates containing the appropriate antibiotic for selection. The fourth step involves the screening of the individual calli for production of the transgenic protein. A final step involves subcloning the calli cells and selecting for the highest producing lines.” (Magnuson, Wang, James, An, Reeves, and Lee, 2002)

Plant suspension cultures are obtained and propagated as calli (undifferentiated cells) which are similar to mammalian stem cells. Stem cells have the remarkable potential to develop into many different cell types in the body. Serving as a sort of repair system for the body, they can theoretically divide without limit to replenish other cells as long as the person or animal is still alive. When a stem cell divides, each new cell has the potential to either remain a stem cell or become another type of cell with a more specialized function, such as a muscle cell, a red blood cell, or a brain cell. Figure 2 shows how stem cells may be used to repair specific tissues.

Scientists are currently investigating the potential of various adult stem cells. According to Rosenthal (2003), Table 2 indicates the most common cell types being studied.

Science fiction writers have popularized the suggestion that these cells could someday produce organs for transplantation. Now, it seems, they may not be so “wacky”!

“Stem cells have been found that can be successfully transplanted from one species to another without immune system rejection, a discovery that could bring stem cell treatments for brain disorders closer.

Researchers from Kansas State University, publishing in the journal Experimental Neurology, report that they have xeno-transplanted umbilical cord matrix stem cells from a pig into the brain of a rat without the rejection of the foreign cells by the rat's immune system.

Umbilical cord matrix stem cells are extracted from a material called Wharton's jelly, a gelatinous connective tissue that helps maintain the umbilical cord's structure and protect its blood vessels.

While they aren't sure why or how, the researchers found that the transplanted cells survived for six weeks undetected, without rejection and without the use of any drugs to suppress an immune response.

It is common for the immune system to reject foreign cells, especially those from other species, and the response poses serious limitations on the success of cell and organ transplants -- especially xeno-transplants.

To counteract immune system rejection, patients are usually put on immunosuppressive therapy. But often, complications arise from immune suppression or from secondary effects of immunosuppressive drugs.

Something about from pig umbilical cord matrix, however, allows them to be ignored by the immune system. And because a subset of the transplanted stem cells respond to the chemical environment of the brain and develop into cells commonly found in the nervous system, they could eventually be used to treat human brain disorders.

"Specifically, the umbilical cord matrix cell source may offer us a basis for treating nervous system disorders like Parkinson's disease," says neuroscientist Mark Weiss.

As evidence from previous studies shows that human umbilical cord matrix cells can differentiate into nervous system tissue, Kansas State researchers are now extending the findings to test human transplant suitability.” (Hunter, 2003)

Materials to obtain before starting the module. One paperback copy of “Chromosome Six” per class involved in the module. You will be tearing it apart so a cheap, well-used copy is preferable. Directions for reading this book in a collaborative, quick manner are included in Appendix A.

One copy per student of “Ethics of the Prophets” found in appendix B.

Bacterial transformation supplies (micropipets, E. coli, plasmid). Any ‘glow in the dark’, ‘green gene’ or x-gal transformation kit will work. I recommend kit IND-9 from Modern Biology, Inc. which sells for about $75.00 per kit. http://www.modernbio.com/ind-9.htm Once you are familiar with the techniques and materials required, you can order by item and save money.

Make student copies of “Comparison of Human and Chimpanzee Chromosomes” found at ENSIweb (Evolution and Nature of Science Institute) for each student. Several hi-liters in different colors for student groups of two to four are helpful. http://www.indiana.edu/~ensiweb/ The lesson may be found by clicking ‘The Lessons’, ‘Evolution’, ‘List of Titles’, ‘Comparison of Human and Chimpanzee Chromosomes’. Student pages are in pdf format.

High School teachers within the state of Washington can obtain lab equipment, such as micropipets, through the equipment loan program at Washington State University. Information can be found at this address on the internet http://www.sci.wsu.edu/bio/equiploa.html

Prerequisite Knowledge and Skills. Students should have experience with the idea of sterile technique. Keeping work areas clean and being aware of possible sources of contamination will facilitate the lab work. Students should practice using micropipettors to transfer solutions, streaking agar plates, using pipettes (eye droppers) and handling microcentrifuge tubes. A basic understanding of DNA replication, plasmid structure and protein synthesis is useful but not essential.

Daily Activities

Each day’s activities are designed for 85 minutes periods although the module could be divided into shorter periods.

Day One - Start by introducing the novel (Appendix A) and going through two rotations of reading. This should take about 30-35 minutes. The short story (Appendix B) may be used here if you are omitting the novel.) Discuss scientific terms and procedures mention in the section (10 minutes). Use it to introduce the DNA extraction from Kiwi. (Appendix C) The kiwi extraction takes about 40 minutes.

Day Two – Go through two more rotations of the novel. Each rotation should take about 12 minutes. This would be a good place to discuss the difference between bonobos and chimpanzees. The activity, “Comparison of Human and Chimpanzee Chromosomes” asks students to consider the relationships between humans, chimpanzees and bonobos and takes about 50 minutes. If more information about bonobos would be helpful, I recommend the video from National Geographic, “New Chimpanzees” (1995).

Day Three - Go through two more rotations of the novel. Review/introduce plasmids and DNA structure and function. Practice biotechnology techniques (Appendix D). Set up Callus Induction experiment (Appendix E). This experiment is long-term and can take a month for callus to form. It also is susceptible to contamination by fungi or bacteria. If you choose to include it, it is a good example of plant cells that are similar to mammalian stem cells.

Day Four – Bacterial Transformation (Appendix F) lab IND-9 usually takes the entire 85-minute class period. Students will already have covered approximately 300 pages of the novel at this point. I have included student pages for inserting the pGreen plasmid from Carolina Biological, Inc. into E. coli colonies picked from a petri dish.

Day Five – Go through as many rotations as need in order to finish the novel. Have students finish focus questions and discuss or turn them in. Check on bacterial transformations. Collect class data (colony counts) for the bacterial transformations and discuss the results. Depending on which kit (plasmid system) you choose, colonies may need an extra day or two. The Ind.-9 kit from Modern Biology, Inc. usually is ready the next day if you have an incubator.