UBD Unit Title: DNA Fingerprinting

Grade Level: High School Biology Key Words: DNA, Replication, DNA Extraction, Gel electrophoresis Designed By: Noel Mellon in collaboration with Mt. Carmel Biology Teachers BRIEF SUMMARY OF UNIT: Playing roles as Forensic Evidence Specialists, students are asked to use numerous science skills to solve a crime. First, they must understand the importance, structure, and function of DNA. Students will be able to apply this knowledge and understanding to identify individuals. Also, students will be able to relate the limitations of this technique. Sterile technique training, biotechnology (DNA manipulation) and the exploration of other uses of Fingerprinting are other highlights of this unit. BIG IDEAS/THEMES: Unity and Diversity Technology and Society

Stage 1: Desired Results

California Life Science Standards 5. The genetic composition of cells can be altered by incorporation of exogenous DNA into the cells. As a basis for understanding this concept: a. Students know the general structures and functions of DNA, RNA, and protein. b. Students know how to apply base-pairing rules to explain precise copying of DNA during semiconservative replication and transcription of information from DNA into mRNA. c. Students know how genetic engineering (biotechnology) is used to produce novel biomedical and agricultural products. d.* Students know how basic DNA technology (restriction digestion by endonucleases, gel electrophoresis, ligation, and transformation) is used to construct recombinant DNA molecules. Understandings Students will understand DNA structure and function, Sterile technique, that each living thing has unique DNA, and applications that exploit this difference (DNA fingerprinting and its limitations). Essential Questions: Why is DNA important? What are common misconceptions about DNA? What are the strengths and weaknesses of using DNA fingerprinting for court cases? Students will know Structure and function of DNA How DNA unifies all living things yet is different in all living things Students will be able to: Extract DNA View differences among individuals using gel electrophoresis

Stage 2: Assessment Evidence

Performance Task: GRASPS (Goal, Role, Audience, Product/Performance/Purpose, Standards & Criteria for success are outlined below) Goal: The goal is to create a Forensic Specialist Report (figure out who committed the crime!). (P) As a Forensic Specialist, your tasks are to gather evidence, extract DNA, and run a DNA gel, understand the evidence, and make a final report. (R)(P) Your audience is the prosecuting attorney, defense attorney, judge and jury. (A) See following pages for unit materials. Key Criteria (S) Specialist Report includes: Evidence of your thorough preparation for court appearance: o Establish yourself as an expert --brief explanation of methods including safeguards against mistakes. o What did you find -- summary of your results including charts and data tables. o What do you conclude explanation of what the evidence indicates regarding guilt or innocence. Specialist Report is: 4 Thorough/complete Completely accurate, all facts and mechanics stated correctly Independently thought out and clear 3

Substantial Generally accurate With minimal assistance, reader able to follow Incomplete/partial Inaccurate, numerous errors detract from result With moderate assistance, lacks clarity, difficult to follow Misunderstanding Major inaccuracies Significant errors, considerable assistance, unclear

Other evidence: Quiz on terms Pre vs. Post test Self Assessment: KWL Chart and Writing Prompts

Stage 3: Learning Plan

W: Where Unit is going: In this unit, youll be solving a crime using a Forensic Science tool, DNA fingerprinting. In order to do this, youll need an understanding of DNA basics. 3 column notes including What do you Know, What do you want to Know, and What Misconceptions have you Cleared up? (KWL notes) H: Hook the Students: Show picture of DNA molecule Question prompts: where is DNA found? Show clip of CSI or other forensic science show Show small section of Talk Show, who is the father? Recent articles on Prison releases, OJ Use of evidence in trial Do DNA extraction as demo E: Equip: Lectures and activities to show DNA basics, interactive websites, puzzle pieces for replication, historical reading Watson/Crick, 3 column notes including What do you Know, What do you want to Know, and What Misconceptions have you Cleared up? (KWL notes) Explore: Article on Prison releases or similarempathy for being in prison for a crime they didnt commit Experience: Activities to go along with the lessons, activate prior knowledge on cells R: Rethinking the DNA whats possible and what isnt. 3 column notes including What do you Know, What do you want to Know, and What Misconceptions have you Cleared up? (KWL notes) E: DNA Fingerprinting (Stage 2) Assessment Student Reflections ideas about where they could go with this technology T: Reading, Computer, Hands-on Activities, Empathy, Lecture

O:

Day 1 (Suggested Monday) Where are we going? Forensic Science Lab Introduction Hook: TV Clip for Forensic Science (CSI) Start the KWL Chart 1 hour Day 2 Tuesday Demo of DNA extraction or show what extracted DNA looks like Show DNA molecule Lecture DNA basics (structure of the DNA), Historical Information 1 hour Day 3 (Suggested LAB day) DNA replication activities Empathy articles (Person released, convicted because of DNA Fingerprinting) 2 hours Day 4 Friday Setup the Forensic Science Lab Jigsaw, Gathering Information/ Interviews with Suspects Share data 45 minutes Day 5 Monday Gather DNA evidence (DNA extractions) Lab techniques (Labeling Properly) Start Report (Data Tables) 1 hour Day 6 Tuesday Webquest 1 hour Day 7 (LAB day) Pour Gels; while theyre solidifying (20 mins), do a quick lecture on gel basics: How to load samples (pipetting) Where to load samples (wells, the order of loading the samples) Also, Get samples ready for loading (label tubes) Load and run Gel While gel is running, give Assessment on DNA structure/ replication Document results 2 hours Day 8 Friday Complete Forensic Scientist Report

DNA Fingerprinting Teachers Notes and suggestions from field testing: It is possible to do the following in one Lab Period (110 minutes) or split into two 55 minute class periods A. Police Report Materials: Copy or copies of Police report (page 6) Teacher or students can read the police report aloud or in small groups. Teacher then discusses what needed to be done together in their individual groups: Interview suspects, gather samples from suspects, extract DNA from suspects, compare suspect DNA with DNA from crime scene. B. Interviewing the suspects Materials: Copies of suspect statements (pages 7 11) Chalk outline of victim with crime scene tape around crime scene Using chalk, draw an outline of a womans body on the sidewalk outside the classroom. Optional: Use Crime Scene Do Not Cross tape around the area. Tape suspect statements on doors or walls around the classroom. After gathering info/alibis, students can exchange notes in small groups, or have a class discussion regarding suspects, motives and alibis. Ask the class What can could be used to place a suspect at the crive scene? Hopefully one of them will say DNA fingerprinting! C. DNA extraction Materials: Copies of DNA extraction lab basics (page 14) Teacher answers on (page 13) Copies of Forensic Lab Request Form (page 15) Copies of DNA Extraction Procedure (page 12 Untoasted raw Wheat Germ (cells) labeled as different suspects and victim. (Suggested one set per table) Balance(s) to weigh 2 g wheat germ Warm water (50 60 C water bath) 70 % (or higher %) Isopropyl alcohol (approximately 2 16oz bottles per class of 32) Dish detergent (Dawn recommended) Test tubes (1 per student 25 X 150 mm, or 50ml capacity) Microcentrifuge tubes (1 per student) Glass stirring rods (1 per student) Thermometer (for warm water bath) Marker to label microcentrifuge tubes Students complete DNA Extraction Lab Basics first. Then they do the DNA Extraction Procedure. When theyve completed the procedure, students turn in their tubes and

FORENSIC REQUEST FORM. This form is to keep track of the samples and to simulate submitting the extracted DNA for processing (so they can load samples on a gel the next day) Discard these DNA samples and provide processed DNA samples (food color mixtures) to load on gels. Gel Electrophoresis (lab day 1 hour 45 minutes) Materials: Copies of DNA Electrophoresis Lab Procedure (1 per pair) page 19-20 Copies of Results from DNA Fingerprinting (page 21) Handout for each student Copies of Preparation for Court Appearance (page 22) one for each student (on the back of Resultsthis can be completed the next day) Gel Electrophoresis setups: I bought our original set from Wards. The gel chambers can hold 2 gels, stacked. So, for a class of 32 = 8 lab stations, I bought 4 gel chambers (each comes with 2 stackable gel trays) and 2 power supplies. Each power supply can run 2 gels. Item Power supply Dual Gel Electrophoresis chamber Cat # Quantity Price, ea. total 15W5340 2 439 878 36W5160 4 175 700

You also need micropipets/tips (p20), agarose, and TAE buffer (or TBE buffer). We run the gels at 0.8% agarose. DNA samples / food colors: A: Congo Red + Bromothymol Blue (not food colors--these are dyes in our chemical stockroom) B: green C: red + yellow D: red + blue We used 1 part coloring and 3 parts water to dilute it. So for sample C: 2.5 ml red + 2.5 ml yellow + 15 mls water. Students load 10 microliters in a well. You can assign suspects or victims however youd like (A, B, C, D, plus criminal is one of these so youll have a match). Because all teachers are doing the lab each period, each teacher chooses a different suspect to be the criminal. To prepare the agarose, use 0.8 gram powder per 100ml 1X buffer. We use about 40 mls per gel and make 350ml of agar for each class (8 tables). Put the agarose and water in a flask and use a magnetic stirrer. Turn the temperature knob to 4-5 and once the cloudy solution turns clear, allow it to cool somewhat before pouring.

Tip: Reuse the agarose by cutting out the colored bands and reuse the agar that is left over. Pour agarose up to about half way up the comb (8 well comb). 1X Buffer solution can be reused a few times. Run from the negative to the positive (From black to red). Run to Red Voltage should be set to 125 volts. If you are pressed for time, use 135volts. If possible, rinse out the electrodes with distilled water and let air dry. This will stop the corrosion.

Preparation for Court Appearance / writeup (approximately 45 mintes) Suggestion: on the board, write terms youd like to see in student answers: equipment/solutions for the DNA extraction and DNA fingerprinting. You can review the terms for them such as: buffer, agarose, gel rigs, chambersetc. Suggestion: Pick 1 group to be the expert on DNA extraction, how fingerprinting works, and how criminal was determined. Suggestion each lab group will present to the teacher: Person 1 talks about the suspects and alibis and motives. Person 2 talks about the DNA extraction and how it was done. Person 3 talked about the DNA fingerprinting procedure Person 4 talks about results and conclusion.

DNA FINGERPRINTING
Goal: You are a forensic specialist. The goal is to create a report for the crime listed below. Your task is to gather evidence, extract DNA, and run a DNA gel, interpret the evidence, and make a final report for the court. Your testimony will be as an expert witness.

Case Study
The year is 2013. You have graduated from college and have just been hired at the San Diego Coroners Office as a Forensics Specialist. Your duties include DNA extractions for DNA fingerprinting. A crime has been committed near Mt. Carmel High School in Rancho Penasquitos. You have been assigned to review the evidence in the case and write a report for the court. You may be asked to testify as an expert witness. Because this will be a legal document, you must make sure you follow proper procedures with the highest level of accuracy. Make sure there is no cross contamination between samples and that you document each piece of data. Following is the initial report from the police regarding the crime.

Police Report
Press release information SUBJECT: DATE/TIME: LOCATION OF INCIDENT: VICTIM: Assault with blunt object. RPSD Case # J101114 _______________________ Call received _________ hrs. 13243 Stargaze San Diego, CA 92129 Cathy (last name withheld).

SYNOPSIS: The victim was reportedly last seen with her boyfriend following lunch at 1:00p.m. Her brother (Tom), who arrived at her house after she failed to pick him up at the airport from a 4:00 flight, discovered the victim. Her mother arrived at the home shortly afterwards. The responding officer stated that the victim was unconscious and bleeding. Paramedics stabilized the victim and transported her to the hospital. The officer noted that there was no indication of forced entry. In talking with the attending physician, the officer was told that the victim is suffering from blunt force trauma to the back of the head and will not be able to answer questions for some time. The victim also has numerous bruises and cuts on her arms and hands. The initial check up also revealed traces of torn skin under the victims fingernails.

TASK: Your first job is to interview those close to her to determine possible motives and suspects. Your office has decided that it would be more efficient for each of you to interview the people separately. The people to be questioned include the brother, the boyfriend, the mother, the delivery boy and any neighbors, who were home at the time.

BOYFRIEND (Jake): Cathy and I went to lunch at Fridays to celebrate her recent promotion. She made fun of me for cutting myself shaving this morning and we talked about going to her cousins upcoming wedding. She said wed have a blast with all the relatives we havent seen in awhile. I looked forward to seeing them, except her brother Tom, I bet it was him. Theyve never been too close and things seem to be worse since they had a fight about their uncle. I took her home after lunch and left.

BROTHER (Tom): I was waitin at the airport from 4:00 to 5:00 for Cathy to come pick me up. She was late, I figured it was traffic, so I wasnt really worried. I was more concerned with the scratch I had gotten on my arm on the plane. The cart always gets you when you sit on the isle. I called home and her cell and got no answer. Before taking a taxi, I called my mom to see if she had heard anything from Cathy. I was shocked to hear she was attacked. [Angry] I bet it was her boyfriend! Hes always made a point of trying to separate Cathy from her family and friends. Cathy has told me before that she has to watch what she says, cuz he gets mad for no apparent reason.

NEIGHBOR(Sam): I was working in the garden when I heard Cathy and her boyfriend pull up. They didnt seem too happy, but Ive heard them argue lots of times. I didnt listen cause the roses were scratching the heck out of me. I saw Cathy use her keys and go into the house at about 1:00. I went inside because my phone rang. About 20 minutes later, a grocery delivery truck drives up and I saw the delivery guy, Joe, unloaded several boxes of groceries. I dont know if he went inside or not, usually he brings the groceries in and puts them in the kitchen when he delivers groceries to my house. Joe has told me before how much he admires Cathy and is always asking about her and the fights with her boyfriend. I believe he has a crush on her.

MOTHER (Beth): I arrived home at 5:30. I was headed to the bathroom to clean the scratch the dog gave me, when I saw Tom hovering over Cathy in the other room. He had already called 911. Cathy was bleeding heavily from the back of her head and was unconscious. [quietly] Cathy and Tom have never been that close, they fight a lot, but it seemed to get worse after my brother discussed his new Will with the family. They have to divide his estate and its a lot of money.

Delivery Boy: (Joe) Yea, I delivered the groceries just like I always do. Cathy was fighting with her jerk of a boyfriend as usual so I just took the stuff to the kitchen. I was hoping hed leave so I could talk with her shes a babe. When I was done I went out the back door. Look, I even scratched my arm on the screen thats loose. They should get that thing fixed. I didnt hear anything when I walked to the truck. Then, I just finished my other stops.

DNA Extraction Procedure

NOTE: All new forensics technicians (students) should complete pre-lab form
DNA Extraction Lab Basics before beginning the lab. Materials: warm water 50-60 C. No warmer than this! skin cells detergent isopropyl alcohol (must be cold) large test tube microcentrifuge tube glass stirring rod thermometer Forensic Lab Request Form Procedure: 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. In a large test tube, put 2 grams of cells collected from suspects. This should be logged into evidence and labeled. Add approx 20 ml of warm tap water 50-60 C. Do not let water go above this temperature range. Stir constantly for 3 minutes with glass stirring rod. Add 1ml of detergent, one drop at a time (to reduce the amount of foam). Stir gently for 5 seconds each minute for 5 minutes. Be very careful not to create bubbles or foam. Slowly add 20 ml of cold isopropyl alcohol by pouring down the side of the tube. Alcohol should sit on top, not mixing with the water. DNA will be collecting at the water-alcohol interface. Let the mixture sit for a few minutes. Make observations. While you are waiting, label microcentrifuge tube and fill halfway with cold isopropyl alcohol. After 15 minutes, the DNA will collect between the solution and the alcohol. You may use a DNA collecting tool (paper clip hook) to gently remove DNA and transfer it into microcentrifuge tube. Submit sample to the Transformation Technician (teacher) with your completed Forensic Lab Request Form.

DNA Extraction Lab Basics

Teacher Answers
New Employees must file this form as requested 1. What Does the abbreviaion DNA stand for? Deoxyribonucleic acid 2. What has DNA? All living (or previously living) substances have DNA. Plants, animals, fungi, bacteria, viruses. 3. Give two ways that DNA is universal? All DNA has the same basic structure regardless of what type of life. DNA is used to pass on information to new progeny. 4. How does DNA differ among different species? Different species have different amounts of DNA etc...Ill let you bio teachers answer that one. 5. How is DNA Different within the same species? Base pairs differ, creating different genes with different functionality etc... another one for you. 6. What is the purpose of the detergent? The detergent acts as a catalyst to break open the cell and nuclear membranes and helps separate hydrophobic substances. 7. What is reason for adding the isopropyl alcohol? Which layer is the DNA Present in? The isopropyl alcohol and water will separate and the DNA will precipitate into the isopropyl alcohol.

DNA Extraction Lab Basics

New Employees must file this form as requested to ensure familiarity with lab procedure. 1. What does the abbreviation DNA stand for?

2. What has DNA?

3. How is DNA in all organisms similar?

4. How is DNA different among different species?

5. How is DNA Different among the same species?

6. What is the purpose of the detergent?

7. What is reason for adding the isopropyl alcohol?

FORENSIC LAB REQUEST FORM Submit DNA samples for Processing

1. In order to submit samples to the DNA lab for testing, fill out the form below. Make sure that all samples are properly labeled. Investigators Date Name Victims Is Victims DNA Name/Case No. being tested? First Sample Relation to Victim Suspect Name Second Sample Relation to Victim Suspect Name Third Sample Relation to Victim Suspect Name 2. Is there a possibility of cross contamination from any of the above DNA sources? Circle Yes/No If so which ones? Explain how this could occur.

3.

List the three parts of each nucleotide unit? A) B)

C)

4. Which nitrogenous bases should be present in this sample?

5. Which of the above bases will be present in equal amounts?

Understanding DNA Basics KWL notes: (Students will divide their paper into three columns: 2 small, one larger) What do you Know? What do you Want to Know? What have you Learned? (Cleared up misconceptions)

Empathy articles http://www.signonsandiego.com/news/state/20031117-1557-ca-lacipeterson.html Scott Peterson DNA evidence allowed http://www.nctimes.com/articles/2003/11/20/news/inland/11_19_0322_40_20.txt Crowe/Tuitt case http://www.miami.com/mld/miamiherald/7447889.htm Dru Sjodn dna found in car

DNA FINGERPRINTING WEBQUEST: NOVA KILLERS TRAIL

1. Go to http://www.pbs.org/wgbh/nova/sheppard/analyze.html 2. Read through the information on creating a DNA fingerprint and answer the questions on your answer sheet. 3. Open Part 1: It Takes a Lickin (a bit corny I know, but go ahead & do it anyway, its fun & informative!) 4. Open Part 2: DNA Fingerprinting at the NOVA Lab. This is great as you manipulate the tools necessary to run a fingerprint. **AFTER EACH STEP, WAIT FOR A POP UP WHAT HAPPENED TO OPEN. ANSWER QUESTIONS ON YOUR ANSWER SHEET.** 5. Open: Science in the Courtroom. Make sure you answer questions. 6. Open: Cleared by DNA, & check out the number of people that have been released from prison because of new technology. Answer question on answer sheet. 7. Open: Chronology of a Murder, and read about the Sheppard case.

NAME: _________________________________

PERIOD _____

DNA FINGERPRINTING WEBQUEST: NOVA KILLERS TRAIL

1. What can a DNA fingerprint be used for? 2. What do restriction enzymes do? Give an example Are all DNA fragments alike? Explain 3. What is the function of the agarose gel? 4. What is meant by electrophoresis? Which moves faster, a larger or smaller fragment? Why? 5. What if the function of the nylon membrane? 6. Why are probes added? What happens to the excess probes? 7. When the film is developed what occurs?

8. Who licked the lollypop? SCIENCE IN THE COURTROOM 9. What are Judge Chernoffs reasons judges should be tutored in DNA analysis? (x4)

10. What will the biggest challenge that science & law will have to face regarding DNA? CLEARED BY DNA 11. What was the longest sentence served? _____ Do you think this person should have received some type of reparation for being wrongly incarcerated? CHRONOLOGY OF A MURDER What were some of the injustices during the investigation and court case?

Pouring an Agarose Gel: 1. Snap the end dams onto each end of the gel casting tray. The end dams seal the ends of the tray.

2. Insert the comb into the slots near the end of the tray. The comb should be placed with the 8-well side down.

3. Get 25mls of liquid agarose from your teacher and pour it slowly and carefully into the tray.

4. Allow the agarose to solidify. It will look opaque in approximately 20 minutes. 5. When the agarose is solid, remove the comb by carefully pulling it straight up. 6. Remove the end dams carefully. Loading your samples: 1. Using the micropipet, remove 10ul of DNA sample from you sample tube. 2. Transfer the sample to a gel well, taking care not to pierce the bottom of the well with the micropipet tip. 3. Each person in the group will load a sample. Be careful not to mix up samples! 4. When ALL samples are loaded, carefully bring your tray to the gel chamber. Running the gel: 1. Make sure that the power supply and cords are away from moisture. 2. Place each loaded gel (still on the gel tray) in the center of the electrophoresis chamber. Position the wells of the gel near the negative (black) electrode. Since DNA is negatively charged the tray is placed in the tank with the wells closest to the negative electrodes. DNA will migrate across the gel toward the positive electrode.

3. Two gels will be put into one chamber.

4. If Running Buffer is NOT already in the gel, slowly pour Running Buffer to one side of the gel. The level of buffer in the chamber should be above the surface of the top gel. 5. Wipe up any spilled liquid or buffer before proceeding. 6. Make sure the cover is dry, and lock it securely onto the chamber. 7. Make sure plugs are dry. Connect the red (positive) cord to the red electrode terminal on the chamber. Connect the black (negative) cord to the black electrode terminal on the chamber. 8. Plug in the power cord to the wall. 9. Turn on the power supply. The red power light will illuminate. Set the voltage adjustment knob to125V. You should see bubbles forming along the electrodes. Removing the gel: 1. Turn off power to chamber. Unplug cord from wall. 2. Wait 10 seconds, then disconnect the cords from the electrodes. 3. Remove safety cover. 4. One person from two groups will put on a glove. That person will carefully remove gel trays from chamber and then carefully push gel off of tray onto a 3X5 card. 5. Document results on Forensic Lab Request Form. Clean-up: 1. Rinse off gel trays, combs, and dams. Carefully dry with paper towel. 2. If teacher requests, rinse chamber but do NOT dryallow to air dry by setting it upside down on a paper towel. 3. Give gel to teacher.

FORENSIC LAB FORM

Results from DNA Fingerprinting

Fill out the form below. Submit with your court appearance preparation sheet.

Explain how cross contamination of DNA sources can occur.

What steps did you take to prevent cross contamination in your samples.

Preparation for Court Appearance

Submitted by ________________________ Date _____________

Attach the Forensic Lab Request Form to this sheet. 1. As a Forensic Evidence Specialist, explain how you gathered DNA samples and then used the process of gel electrophoresis to produce DNA fingerprints.

2. What were your findings? (Summary of your results including charts and data tables)

3.

What do you conclude? (Your explanation of what the evidence indicates)