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If we are not teaching biotechnology to our students, then we are

By Cheryl Williams,2014-06-19 06:28
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If we are not teaching biotechnology to our students, then we are

Pat’s Biotech Lab Curriculum

    PATS BIOTECH LAB CURRICULUM

    If we are not teaching biotechnology to our students, then we are not preparing our students for their future.

Biotech Curriculum Spreadsheet

    My twin sons were born in 1974 with a genetic disorder, Phenylketonuria (PKU). Since that time I have made it my business to try to understand and keep up to date with the field of biotechnology. In 1990 I received a $30 000 grant from the Richard A. Lounsbury Foundation to start a biotech class and to provide training for other teachers. Since 1988, I spent my summers attending various biotech institutes, mainly at Cold Spring Harbor‟s DNA Learning Center, and doing research in various university labs such as Baylor College of Medicine where I was able to work on PKU. I frequently presented biotech workshops as a teacher. I present workshops all the time now as part of my position as Product Manager in the Biotechnology Department at Carolina Biological Supply Company. During my 27 years of teaching biology (and specializing in biotechnology for a decade), I counted on Carolina for practically everything I used. I feel well qualified to recommend the following (alternatives listed at end) biotech laboratory curriculum.

    Use the Carolina? catalog number that appears in parentheses after the boldface name of

    the product to search the website, www.carolina.com, or current catalog for further descriptions and

    prices.

    As a preface to my recommended labs let me list three EXCELLENT resource books that have great explanations, drawings, and extremely helpful appendices. Have your library purchase these, if necessary:

    DNA Science (21-2211) by David Micklos and Greg Freyer this was my main textbook for my

    Biotechnology class. The first half of the book contains informational chapters (with superb illustrations), and the last half contains 10 sequential labs, most of which are, in some form or fashion, part of my recommendations listed below.

    Recombinant DNA and Biotechnology (21-2214) by Helen Kruezer and Adrianne Massey. This

    book has many of the same labs as DNA Science but it also has numerous paper and pencil (dry) labs that are great to use as pre- and post-lab discussions and are also wonderful to use during many of the “down” times like waiting for gels to run;

    The Complete Idiot’s Guide to Decoding Your Genes (21-XXXX) by Mark Bloom and Linda

    Tagliaferro [available Spring 2001]. This book serves as a tremendous shelf reference. I highly recommend it as a basic reference for every school library.

    The main areas to cover in a biotech lab class are: 1) DNA structure & function; 2) basic lab techniques; 3) Transformation; 4) Electrophoresis; 5) Restriction analysis; 6) PCR; 7) Sequencing; 8) Bioinformatics; and, 9) Bioethics.

Pat Ryan Page 1 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

Pat’s Biotech Lab Curriculum

    1. First, do a DNA extraction. This lab lets your students see that DNA is “real stuff” and not just

    some pictures in a book. Let your students examine and scientifically observe strands of DNA

    they have spooled from an organism. This lab turns an abstract and somewhat mysterious

    concept into a concrete entity that your students could even demonstrate to their parents as a

    little “kitchen (bio)chemistry”. Considering consistently good results and ease of „teacher prep

    time‟, the best extraction lab I have experienced is with the Wheat Germ DNA Extraction Kit

    (15-4704, for a class of 30). Scientific observations students should glean from this lab include:

    1) DNA is linear;

    2) DNA has a white, cream color;

    3) DNA is soluble in water and insoluble in alcohol; and,

    4) If you use a pH indicator, DNA is acidic.

     Allow at least three class periods (I am figuring on 50-minute class periods):

    ; 1 for pre-lab discussion to review the structure and function of DNA;

    ; 1 for doing the lab; and

    ; 1 for post-lab results and discussion.

    2. Compared to reading and looking at pictures in a book, I had many more students say, “Hey, I

    get it now!” when using what I called the „Green Board‟ for them to learn DNA structure,

    replication, transcription and translation. The DNA Made Easy Kit (12-1040) provides a ‟hands-

    on‟ application of DNA structure/function, replication and protein synthesis. I would place my

    students in teams, demonstrate the processes, provide guided practice, and then assess the

    teams of students as they „taught‟ me the processes. They really liked being able to manipulate

    the pieces and work in teams and, also, it was an easy assessment for the teacher. One „Green

    Board‟ is enough for the whole school and I frequently brought it out for reviews.

    3. Students need to have some expertise handling biotech equipment and instruments, and also with

    using sterile techniques. My advice is to take several class periods (3-5) for students to build

    confidence with their skills. The Sterile Technique Kit (21-1075, for 6 stations) is designed to

    familiarize students with the necessary skills for handling bacteria. Also, there is a Practice

    Pipetting Stations kit (21-1145, for 10 stations) for students to use to increase their pipetting

    skills. Part of the student practice should involve making electrophoresis gels. Students cold

    prepare the gels to be used in the next lab at this time.

    4. There are certain basic objectives scientists desire in doing research and also certain basic

    techniques they employ. The goal of a well-rounded biotech curriculum should be to expose

    students to as many of the basics as possible in order that they more fully understand and

    appreciate what is happening in a real research lab. One objective of a biotech lab is often to

    figure out how DNA is arranged. This is called „mapping‟ and is used with both large and small

    sections of DNA. Naturally, it is best to start students off with small DNA mapping problems. The

    Restriction Mapping of Plasmid DNA Kit (21-1175, for six stations) is a simple, yet

    educational, load-„n‟-go lab that yields consistently good results for students. In addition, after

    performing the lab, students must THINK and REASON and APPLY knowledge in order to correctly

    „map‟ their DNA. Again, set aside at least three class periods (same as above) to get the most out

    of this lab.

    Pat Ryan Page 2 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

    Pat’s Biotech Lab Curriculum

    5. If you don‟t teach any other biotech lab – teach transformation! Practically all of the

    major advances in genetic engineering in the fields of medicine, agriculture, ecology, and genomics are based on taking DNA from one organism and putting it into another organism and making it do what you want it to do! This genetic engineering process is what we call

    transformation the uptake and expression of a gene. You can make your students real gene

     with this lab!! By far, the very best transformation lab I have experienced and, believe jockeys

    me, I‟ve “done them all” – is the Green GENE Colony Transformation Kit (21-1082, for a six

    station kit). It is just so striking when students see a different color in their transformed cells

    they just know that something different has happened to their cells. And these transformants will be green without any special lighting or extra chemicals to add to the agar. Once again, this is the best transformation lab due to its consistently good results and minimal teacher prep time. The normal three class periods should be set aside for doing this lab with an extra day or two for teacher prep work (about 10 30 minutes each day). This lab is just really good science! Hardly

    any other lab will drive home the concepts of good experimental design and the use of controls in an experiment like this one. The results and discussion parts of this lab really cause the students to THINK and REASON. Also, there is a great math integration possibility when figuring the transformation efficiency. I regularly refer to this lab as “The Best Lab in the Book! “

    6. The DNA Restriction Analysis Kit (21-1106, for six stations) is, along with transformation,

    another classic biotech lab that introduces students to some of the fundamental techniques, skills, and ideas that have been and are currently used in research labs worldwide. In this lab students actually cut (restrict) DNA with several molecular scissors (restriction enzymes) and then analyze the results. The analysis and discussion of the results again requires students to THINK and REASON and to APPLY their knowledge. Students acquire the following skills:

     Handling DNA and restriction enzymes by pipetting ;

     Cutting DNA with restriction enzymes;

     Electrophoresis: Preparing, loading, running, and staining gels; and

     Analyzing results and determining base pair lengths for unknown DNA using semi-log graph

    paper.

    7. Polymerase Chain Reaction (PCR) is one of the most often used techniques in research labs today. This powerful procedure is what made the Human Genome Project possible. It allows scientists to amplify (make billions of copies) a sample of DNA with which they can then analyze and experiment. If you do not have the capacity to perform real PCR, then I advise you to do a simulation such as the PCR Forensics Simulation Kit (21-1210, for six stations) which

    reinforces the electrophoresis skills of your students and introduces the topic of PCR for discussion. The „forensics‟ angle is a good „hook‟ to spark student interest. Again, students will need to THINK and REASON and APPLY their knowledge. For the understanding of what PCR is really all about, the absolute best animation and explanation of the procedure can be found at the web site for Cold Spring Harbor‟s DNA Learning Center (http://vector.cshl.org). This site has numerous, fantastic

    animations for anything and everything you, and your students, ever wanted to know about DNA!!

    Pat Ryan Page 3 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

Pat’s Biotech Lab Curriculum

    8. If you are looking for a PCR lab that you can afford, look no further! For the simplest lab

    experience that conveys polymerase chain reaction procedures, I recommend the Lambda PCR

    Kit (21-1223, for six stations). Students do an actual PCR experiment without the need for an

    expensive thermocycler. This „foolproof‟ lab is a great “hands-on” way to understand the

    “magical” PCR process that occurs while a thermocycler is running. Students will perform a

    „timecourse‟ amplification of an 1106bp segment of the Lambda genome using two waterbaths. A

    timecourse process allows students to visualize the results of the PCR amplification procedure

    from cycle to cycle. Students should certainly better understand what‟s happening in a PCR

    experiment having completed this lab.

    One benefit of this lab is that it can withstand a great deal of student abuse. Another benefit of

    this particular PCR lab is that there is no need for expensive pipettors. I suggest that you use the

    Pipetting Device (21-1022, for a set of 5 syringes with 150 tips calibrated at 2 and 10 uL

    volumes) for a less expensive alternative.

************************************************************

     All of the following PCR labs utilize Ready-To-Go-Beads (RTGB) which eliminates much prep

    time and possibility for student error. To further reduce prep time and possibility for student error,

    I highly recommend using a thermocycler with a heated lid such as the OMN-E Thermal Cycler

    (21-6273). This is definitely a worthwhile investment considering the time and effort you and your

    students will put in to do a PCR lab.

Each PCR lab involves five basic components:

    ; Isolating DNA from human cells (hair sheaths or cheek cells);

    ; Utilizing specific primers to amplify the chosen locus by PCR.;

    ; Analyzing the amplified DNA by gel electrophoresis;

    ; Analyzing and discussing the results of the lab; and,

    ; Possible extension of the lab using computers BIOINFORMATICS!

A typical class schedule for a PCR lab might be:

    Day 1 Pre-lab discussion

    Day 2 DNA isolation

    Day 3 PCR amplification (samples typically run overnight)

    Day 4 Electrophoresis: load, run, stain, photograph gels

    Day 5 Results and Discussion

    Day 6 Extension activities on the DNALC web site

    Pat Ryan Page 4 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

    Pat’s Biotech Lab Curriculum

     If possible, try to do at least one of these three labs that involve actual

    PCR.

     These labs require the use of a thermocycler [PCR machine].

     If your school cannot afford a thermocycler of their own, I suggest you try to:

    A. Borrow the use of one from a nearby source (university, hospital, industry, or

    other school);

    B. Get your district to purchase one for the whole school district (you won’t need

    to use it everyday!);

    C. Get financial assistance from your state’s education department – remind them

    that technology money can be used for instruments other than just computers.

    9. The Human Mitochondrial DNA Kit AT (21-1238, for 25 reactions) should be the first if you

    are going to do all three PCR labs. This lab utilizes DNA from the mitochondria (mtDNA) in the students‟ cells. Because there are numerous mitochondria in each cell, there is a great quantity of

    DNA available for amplification which translates to mean that this lab can absorb more student abuse (mistakes and errors) than the following labs. Due to the greater amounts of DNA used, this lab lends itself well for „hand-cycling‟ and, also, the number of cycles for this lab can be shortened some if necessary. Students will extract and amplify a 460-nucleotide sequence of DNA from within the control region of the mitochondrial genome of their own cells. These control regions have been widely used to study human evolution. As a bonus, amplified student samples may be submitted to the DNA Learning Center‟s (DNALC‟s) Sequencing Service which will

    generate your students‟ individual DNA sequences and post the results at the web site for Cold

    Spring Harbor‟s DNA Learning Center (http://vector.cshl.org). Students can then compare and

    analyze their sequences with other students and populations across the U.S. and around the world. Additional extension activities such as, „Ancient DNA‟ and the „Mystery of the Romanovs‟, can be found at the DNALC web site.

    10. The Human Alu Insertion Polymorphism Kit AT (21-1232, for 25 reactions) allows students

    to visualize their own DNA and to compare their „DNA Fingerprint‟ with other classmates. The lab checks for the presence (+) or absence (-) of a ~300bp transposable Alu element located on

    chromosome 16. This lab lends itself to population studies, Hardy-Weinberg distributions, and human evolution studies. Facilities for these extension studies can be found at the web site for Cold Spring Harbor‟s DNA Learning Center (http://vector.cshl.org). If I could do only one PCR lab, I

    would do this one.

    11. The Human VNTR Polymorphism Kit AT (21-1235, for 25 reactions) again allows students to

    examine their own DNA. This lab checks for a VNTR (variable number of tandem repeats) polymorphism which is caused by short, repeated copies of a 16-nucleotide sequence at the pMCT118 locus on chromosome 1. Differences in the number of repeated units produce longer or shorter alleles, which show up in the electrophoresis gel as bands of various lengths. Because there are 29 known alleles, a class of students shows a variety of different genotypes. This illustrates the use of DNA fingerprinting to identify individuals in court cases and disasters. This lab is more sensitive than the Human Alu Insertion lab, so, if you can do both, be sure to do this one last so that student skills and techniques will be a little sharper.

    Pat Ryan Page 5 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

    Pat’s Biotech Lab Curriculum

    Any well-rounded BIOTECH curriculum must also include investigations into the field of Bioinformatics. This new field of study involves the use of computers to analyze and try to make sense of all the information generated by the Human Genome Project and other sequencing efforts. Scientists that use laptops instead of test tubes will make many important discoveries in the future! The term „gene mining‟ will signify prospectors (scientists) using computers instead of pick axes to retrieve „gems‟. Any and all of the three PCR labs listed above can be utilized to give your students the much-needed experience of analyzing databases for necessary information by utilizing the extension activities with the DNA Learning Center‟s website.

    If you are unable to perform any of the PCR labs, the Carolina? Webcutter Kit (21-1195,

    for six stations) is designed to bring your biotech curriculum into the computer age. Your students will experience how computers and databases are used in a biotech research lab. This kit involves 3 modules as follows:

     Module I students do two dry labs involving DNA sequencing;

     Module II students log on to the internet to use the Carolina? Webcutter software to

    determine the identity of their gene fragment and its restriction map; and, Module III students perform restriction analysis to confirm the identity of their unknown

    gene (“gene mining”).

    Pat Ryan Page 6 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

Pat’s Biotech Lab Curriculum

     Additionally, any well-rounded Biotech curriculum must also include activities for students to explore the field of Bioethics. With each passing day, more biotech discoveries are made that lead to opportunities, possibilities, and decisions for our students to explore. To prepare for the world in which they live, your students need to actively investigate some new situations in a scientific manner, striving to make logical decisions and conclusions. Your students will also need practice placing their prior knowledge, beliefs and emotions in a proper perspective when investigating bioethical issues.

     Probably the best sources for these issues are current newspapers, magazines, or internet news sites. I modified a “Reading Reaction” form (included in this packet) that I was introduced to

    at a workshop several years ago at Cold Spring Harbor. This is a two page “teacher-friendly” form

    that allows for quick assessment. I maintained a small library of articles in my classroom from which students could choose their weekly reading reaction assignments. They were also free to obtain articles on their own. Be sure to stress that their readings must be „substantive‟ in nature and not just a quick paragraph or two. I required that at least a portion of their assignments had to come from scientific journals or magazines such as “Scientific American”. The DNA Learning Center‟s home page provides an excellent source of biotech articles under the heading of “Gene News”. I have included some other great websites that you and your students can explore.

    Additionally, I highly recommend that you and your students visit the DNA Learning

    Center‟s website concerning Eugenics (http://vector.cshl.org/eugenics.html). The intriguing story of

    America's embrace of eugenic engineering has been largely hidden in historical archives and scholarly publications.

    It is imperative that we learn from our past mistakes, especially when dealing with such a powerful force as biotechnology. The study of the Eugenics Movement in America should be an eye-opening and very interesting topic that allows your students to experience the history and social interpretation of modern science. This, of course, would be a terrific unit to integrate with history and/or social studies classes.

     Last, but not least, a well-rounded Biotech curriculum needs to have an assessment tool to determine what your students have learned. I modified a biotech vocabulary assessment

    instrument that I picked up in a workshop. I used this as both a pre-test and a post-test. The students knew that they would see the very same exam at the end of the course and were encouraged to compile definitions/explanations in a lab book throughout the course which they could then use during the post-test.

     I have included a spreadsheet of the reagents, equipment, supplies, and consumables necessary for the labs on this list. Do not hesitate to contact me if you have questions or comments concerning any of the above ideas and materials. Best wishes for a productive school year!

Pat Ryan 8/2/00

    Pat Ryan Page 7 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

Pat’s Biotech Lab Curriculum

    OPTIONAL LABS / ACTIVITIES

1. You might consider substituting the Onion DNA Extraction Kit (21-1133, for 15 extractions) for

    the Wheat Germ DNA Extraction for the purpose of demonstrating to your students that they can

    extract and observe DNA from a common “grocery” organism such as an onion. The downside to

    this lab is that it is stinky smelly, and requires a little more teacher prep time. Still, you can get

    great results and make a point. Also, this is a lab that your students could take home and do in

     with their parents using dish soap (with a little salt) as a lysing agent, Adolph‟s Meat the kitchen

    Tenderizer? as a protease, and ethanol to pull the DNA out of solution. Talk about great school /

    home involvement!

    2. If you have the time and capability, you can make your own “practice pipetting stations” by

    purchasing Knox Gelatin? at the grocery and forming it in any convenient sized container (such

    as petri plate tops and bottoms or Tupperware? containers). “Practice wells” can be punched in

    after the gelatin sets up with a pencil or other appropriate sized device.

    3. Restriction mapping problems are, in essence, logic problems. There are numerous paper & pencil

    problems available. I include some with this document.

    4. Do not fail to do a transformation lab! There are numerous types listed in the Carolina catalog,

    but again, for simplicity and results, I highly recommend the GREEN Gene lab!!

    5. Carolina has a really great working and, relatively inexpensive group of Exploring

    Electrophoresis experiments. These can run on battery power (1 5 nine volt batteries) or, I

    recommend, a permanent Exploring Electrophoresis Power Supply (21-374, for running two

    gel boxes). Each kit contains agarose, TBE buffer, Carolina BLU stain, DNA samples (enough for

    10 separate experiments) and 5 sets of apparatus which include the following:

    ; Gel boxes;

    ; Electrical leads;

    ; Gel box electrodes;

    ; Combs;

    ; Pipetting devices; and

    ; Sets of DNA and enzymes.

    Exploring Restriction Analysis and Electrophoresis of DNA (21-1010) [students

    actually „cut‟ DNA using restriction enzymes] is a good substitute for the DNA Restriction

    Analysis Kit and does not require expensive gel boxes or power supplies.

    Exploring Electrophoresis and Forensics (21-1014) [students just “load „n‟ go”

    using pre-digested DNA] is a good substitute for the PCR Forensics Simulation Kit.

    After an initial purchase, which yields 5 sets of apparatus, you can just order the refill kits or

    build up a good supply of apparatus in just a year or so. This would be my ‘kit of

     preference’ if all I wanted to do was to teach the concept of electrophoresis.

Pat Ryan

    Carolina Biological Supply Company

Pat Ryan Page 8 6/19/2012

    Carolina Biological Supply Company 800.227.1150 x5963 pat.ryan@carolina.com

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