The Clark Family Story: Tracing the Cause of Hemophilia

The Clark Family Story: Tracing the Cause of

Hemophilia

by Trish Strohfeldt West Bend East High School, West Bend, Wisconsin In collaboration with the Centers for Disease Control and Prevention's

Nicole Dowling and Stacy League

This lesson is for a senior level biology course and emphasizes the relationship between DNA

sequences, mutations in DNA and the change in the resulting protein structure and function.

Hemophilia A will be used as a real life example of how a mutation in DNA results in a change in

a protein's structure and resulting ability to function. In order to complete these activities, the

students should have prior knowledge in DNA structure and mutations, protein synthesis,

Mendelian genetics and sex-linked traits.

Disclaimer: The findings and conclusions in this report are those of the author(s) and do not necessarily represent the views of the Centers for Disease Control and Prevention.

The Clark Family Story: Tracing the Cause of Hemophilia

by Trish Strohfeldt

West Bend East High School

West Bend, Wisconsin

Summary This lesson is for a senior level biology course and emphasizes the relationship between DNA sequences, mutations in DNA and the change in the resulting protein structure and function. Hemophilia A will be used as a real life example of how a mutation in DNA results in a change in a protein's structure and resulting ability to function. In order to complete these activities, the students should have prior knowledge in DNA structure and mutations, protein synthesis, Mendelian genetics and sex-linked traits.

Learning Outcomes ? The student will be able to relate a DNA sequences to the resulting protein structure in the factor VIII gene. ? The student will be able to demonstrate the use of restriction enzymes in the identification of a specific DNA sequence and explain how this can be used to diagnosis someone has having the hemophilia factor VIII mutation. ? The student will be able to demonstrate how a protein's structure and function can change with a change in the factor VIII DNA sequence (mutation). ? The student will be able to explain the genetics of hemophilia inheritance and affect on the blood clotting process. ? The student will be able to draw conclusions about the risk of passing on the mutated gene.

Materials 1. Photocopy of introduction and background activity-one per student 2. Photocopied DNA sequence templates-one set per group of 4 students 3. 15 feet of white paper (large roll butcher type) with base pair lengths marked (see diagram-Electro paper gel template) 4. Floralwire-40cm per student 5. Sharpie markers-1 per student 6. Computers with Internet access and RASMOL program 7. Biology textbooks to use as reference materials 8. Lab materials including tape scissors, wire, and markers

Total Duration 4 hours

Procedures

Teacher Preparation The teacher should prepare photocopies of handouts. Copies will need to be made of the following: Introduction ? "Pretest and Clark Family Background"; Step 2 ? "Student Template Factor VIII DNA Sequences" (one per group of 4-5), Hemophilia Factor VIII Mutation Activity Student Directions; Step 3 ? "Student Directions: Physical Protein Model Building", "ConclusionGenetic Counselor Report Rubric".

For Steps 2 and 3 organize lab materials (tape, scissors, wire, markers) at lab stations. A computer lab should be reserved for the Step 3 and the RASMOL program should be downloaded and tested. For the activity in Step 2, use large pieces of paper to create the electrophoresis gels with the + and ? ends, wells and base pair markers labeled on each. Use the "Electrophoresis Gel Template Paper" as a guide for drawing this.

This lesson plan should be taught after students have covered the basics of inheritance, including DNA structure, protein structure and synthesis, mutations, Mendelian genetics, and sex-linked traits. For additional information on these topics, visit the Web resources included below.

Web Resources Title: Online Mendelian Inheritance in Man-Hemophilia A URL: Description: This Web site is the OMIM hemophilia A entry that includes background information for the teacher on the biochemistry, clinical features and research.

Supplemental Document Title: Electrophoresis Gel Paper Template Description: This document is a scaled down drawing of what the teacher will need to create on large white paper for each lab group in order for them to complete the activity. Once the large versions of the gels are created the teacher may re-use these each time the lesson is done.

Introduction Before beginning the lesson, the "Pretest ? Hemophilia Lesson Plan" can be used to assess students' prior knowledge. The "Pretest Answer Key ? Hemophilia Lesson Plan" contains answer to the pretest for the teacher's reference.

Following the pretest, students will be introduced to hemophilia and its inheritance pattern. Using "The Clark Family Background" document, students will read a description of a family and their symptoms and use that information to learn about hemophilia. Students should take notes as the background information is discussed since they will need this information to complete later steps and to include in their conclusion paper. A Web resource is included below to supplement the conversation.

Web Resource Title: National Hemophilia Foundation URL: Description: This Web site provides background information on hemophilia A (Factor VIII) deficiency. Teachers can use this Web site to supplement their discussion on the Clark family and hemophilia.

Supplemental Documents Title: Pretest ? Hemophilia Lesson Plan Description: This file is the pretest that can be used to assess the students' knowledge of genetics, hemophilia proteins, and inheritance before the lesson.

Title: Pretest Answer Key ? Hemophilia Lesson Plan

Description: This file is the pretest answer key that can be used to discuss the students' responses to the pretest.

Title: The Clark Family Background Description: This file is the introduction information that gives the students background on the Clark family and the disorder of hemophilia.

Step 2

Duration: 45 minutes

Now that the students have been introduced to the Clark family and hemophilia, students will

explore the genetic underpinnings of the Clark family's disease. The teacher will organize the

students into groups of 3-4 and distribute one "Student Template Factor VIII DNA Sequences"

document to each group and the "Factor VIII Mutation Activity Student Directions" worksheet to

all students. Following the instructions in the worksheet, the students will cut apart each DNA

strand on the template, tape them into the correct order, and label each strand with the source

information. The actual DNA sequence for factor XIII can be found on the "NCBI Factor VIII F8

Entry" Web site included below.

Following the student directions, students will be given a restriction enzyme to cut the DNA sequences with. For the teacher's reference, a site that matches nucleotide sequences with restriction enzymes is included below. Answers for the restriction cuts are found in the "Factor VIII DNA Sequences Answer Key" along with other sequence information.

After they have cut the sequence with the restriction enzyme, students will compare the sequences on a simulated gel to determine which of the family members carry the Hemophilia A mutation, which family members have the disease, and which ones are free of the mutation. The gel will be copied onto the large electrophoresis gel drawn by the teacher in the Teacher Preparation Step. "Hemophilia Factor VIII Mutation Activity Answer Key" provides answers to the questions found in the student directions worksheet.

Web Resources Title: NCBI Factor VIII F8 Entry URL: &val=67083285 Description: This Web site will provide you with actual DNA sequence nucleotide bases for the factor XIII gene.

Title: NEBcutter V2.0 New England Biolabs Inc.

URL:

Description: This Web site allows you to input any DNA nucleotide sequence to find

restriction enzyme cut sites along the sequence.

Supplemental Documents Title: Student Template Factor VIII DNA Sequences

Description: This is the document that contains each family member's DNA strands,

which each group will cut out to analyze. Each group will need one copy of this file.

Title: Hemophilia Factor VIII Mutation Activity Student Directions Description: This document is the student version of the activity and post activity questions that are to be completed as students complete the activity.

Title: Hemophilia Factor VIII Mutation Activity Answer Key

Description: This document is the teacher's answer key to the student activity.

Title: Factor VIII DNA Sequences Answer Key Description: This document is the teacher answers to the DNA sequences and where the restriction enzyme cuts each DNA strand.

Step 3

Duration: 1 hours, 30 minutes

To further their exploration of the genetic underpinnings of the Clark family's hemophilia, the

students will use the DNA sequences found in the "Student Template Factor VIII DNA

Sequences" document from Step 2 to determine the effect of the mutation in the factor VIII gene

on the resulting factor VIII protein structure. Using the instructions "Student Directions: Physical

Protein Model Building", students will begin by transcribing and translating their DNA into

mRNA, amino acids, and shapes. Then, each student will create a physical model of one of the

family member's factor VIII protein and compare the structures. Finally, the students will use a

molecular visualization tool called RASMOL to look at the structure of a part of the factor VIII

protein. Information about downloading and using RASMOL is found in the Web resources for

this step. The student directions worksheet can be evaluated using the "Student Directions:

Physical Protein Model Building Answer Key". In addition, the mRNA, amino acid, and shapes

can be evaluated using the "Factor VIII DNA Sequences Answer Key" found in Step 2.

Web Resources Title: RASMOL Quick Start Page URL: Description: This Web site provides an introduction to using the molecular visualization tool RASMOL. Students can use it before the lesson or during the lesson.

Title: RASMOL Download Page URL: Description: This Web site provides information on downloading the free version of RASMOL. Once downloaded, RASMOL can be used to view and modify coordinate files of molecular structures.

Supplemental Document Title: Student Directions: Physical Protein Model Building Description: This worksheet directs the students to create the physical model of the mutated and normal version of the factor VIII protein to compare the structure and then infer the effects on the function of the protein in the blood coagulation pathway. Students will transcribe the DNA of each family member into mRNA and then translate into amino acids. The student will then use wire to create a physical model of the protein.

Title: Student Directions: Physical Protein Model Building Answer Key Description: This document is the answer key to the questions on the student version of the activity.

Conclusion

Duration: 45 minutes

After students have found the gene responsible for the disorder and have found out which

members of the family are affected and which members are carriers in the previous steps the

students will then act as genetic counselors. As genetic counselors they will prepare a report

for the Clark family that includes the information on living with hemophilia, the affected gene and

location, mode of inheritance and risk of future offspring in the family (parents and children)

developing hemophilia. To create this report they should use what the have learned in previous

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