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SOP120.001.02 Rev. D Effective 02/03/2003 Page 1 of 1

10004530

Life Science GroupMATERIAL SPECIFICATION

PART NUMBER

EFFECTIVE DATE: 04/03/06 DCO #: BP-006621 Rev: A

MATERIAL DESCRIPTION:)

Manual for Comparative Proteomics Kit I: Protein Profiler Module

COMPUTER FIELD DESCRIPTION (30 CHARACTERS):

MANUAL,COMP PROT KIT I: PP MOD

ATTRIBUTES

FORMULA: N/A MOLECULAR WEIGHT: N/A

CAS #: N/A

SPECIFICATIONS: SIZE: 8.5 X 11

PAPER: 20 # RECYCLED PAPER OR EQUIVALENT

INK: BLACK THROUGHOUT, DOUBLE SIDED

PAGES: 114 pagesincluding cover

BINDING: COVERBIND

METHODOLOGY: N/A

PACKAGING: N/A

SHIPPING: Material shall be packaged and shipped in a manner that will insure protectionof the material from the adverse effects of moisture, dirt and other forms ofcontamination or the adverse effects of temperature.

STORAGE: Ambient X 4C 20C 70C Other

QUALIFIED PARTS

VENDOR VENDOR CODE VENDOR PART NO. COMMENTS

BIO-RAD MAIL ANDCOPY CENTER

LASTING IMPRESSIONS

OFFCO

ACTION PRINTERS

GILMAN ST. PRESS

FUHRMAN PRINTING

002401000

V0095220

V00601343

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V00007671

Primary vendor

Equivalent Substitutions Authorized: Yes X No

Special Instructions:

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Biotechnology Explorer

Comparative Proteomics Kit I:Protein Profiler Module

Catalog Number

166-2700EDU

explorer.bio-rad.com

Note: Kit contains temperature-sensitive reagents. Open

immediately and see individual components for storage

temperature.

For Technical Support Call Your Local Bio-Rad Office or in the U.S. Call 1-800-4BIORAD (1-800-424-6723)

Duplication of any part of this documentpermitted for classroom use only.

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Dear Educator:

In 2001 the Human Genome Project completed sequencing all human genes. Far fromclosing a book, this body of work opened up a whole new field, proteomics, which asks afar more important question what do our genes do? Genes encode proteins and proteinsdetermine an organism's form, function and phenotype and as such are the raw materials

of natural selection. Proteomics is the study of the function, structure and interaction ofproteins with each other and their environment. This Protein Profiler kit moves beyond DNAand allows students to employ one of the most widely used techniques in biotechnologyresearch-protein gel electrophoresis.

This kit allows your students to explore evolution at the molecular level using the centralframework of biology. Students generate protein profiles from the muscles of distantly andclosely related species of fish and compare and contrast these profiles to test the hypothesisthat protein profiles are indicators of genetic and evolutionary relatedness.DNA RNA Protein Trait.

Changes in proteins reflect changes in the gene pool. Muscle protein mostly consists ofactin and myosin, but numerous other proteins also make up muscle tissue. While actinand myosin are highly conserved across all animal species, the other muscle proteinsexhibit more variation even among closely related species. Variations between organisms'protein profiles reflect physiological adaptations to different environments, but they originateas chance DNA mutations. Such random mutation events, if favorable, persist through thenatural selection process and contribute to the evolution of species with new specializedfunctions:Mutation Variation Specialization Speciation Evolution

This is an open-ended inquiry-based kit. Students make predictions about their results inpre-lab activities using Internet databases or published phylogenetic information. Theygenerate novel results and apply their results directly to the problem of solving evolutionaryrelationships. By constructing cladograms (phylogenetic trees) based on their data studentsdecide whether their results support or refute their own predictions and published phylogeneticdata.

Protein electrophoresis is principally performed using vertical electrophoresis apparatus

and polyacrylamide gels; however, horizontal electrophoresis apparatus and agarose gelsmay be employed using a specifically designed protocol included in the kit curriculum. TheProtein Profiler kit is a stand alone activity. However, it was developed in conjunction withthe Comparative Proteomics Kit II: The Western Blot Module which takes this activity to thenext level and enables students to use the proteomics techniques of Western blotting andimmunodetection, to specifically identify a single muscle protein, myosin, out of the hundredsof muscle proteins in the muscle proteome.

This kit guides students through the thought processes involved in a laboratory-basedscientific investigation. Students are asked: Can molecular data reveal similarities anddifferences among species? In their analyses, students compare similarities and differencesin their fish protein profiles, create evolutionary trees (cladograms) from their own gelelectrophoresis data, and compare their results to published evolution data. Students areasked: Do the phylogenetic data agree? Does molecular evidence support or refute the

theory of evolution? Why or why not? What explanations can you suggest?This curriculum was developed in collaboration with Dr. Kristi DeCourcy of the FralinBiotechnology Center at Virginia Tech. We'd like to thank Dr. DeCourcy for her invaluableguidance and contribution to this curriculum.

Ron MardigianDivision Marketing ManagerBiotechnology Explorer Program

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Create context. Reinforce learning. Stay current.New scientific discoveries and technologies

create more content for you to teach,but not more time. Biotechnology

Explorer kits help you teach more

effectively by integrating multiplecore content subjects into a

single lab. Connect conceptswith techniques and put

them into context withreal-world scenarios.

ProteinProfiler

Kit

Environmentaland Health

Science

Symbiosis Interdependence of organisms Protein profiles and evolutionary

relationships Using protein profiling to create

cladograms

Cell biology Muscle structure and function Adaptions to environment

Interpretation of pedigrees Morphological vs. genetic

classification

Use of sophisticatedtechniques to generate

protein fingerprints Use of experimental controls Interpretations of experimental

results

Central dogma:DNA > RNA > protein > trait

DNA structure and function Genetic vs. morphological classification Phylogenetic relationships

Chemical properties of proteins Amino acid sequences Protein structure (1, 2, 3, 4) Protein extraction techniques Gel electrophoresis of proteins

Molecular variation Evolution of adaptive traits Environment and natural

selections Molecular basis of organisms

form and function

ScientificInquiry

Chemistryof Life

Heredityand Molecular

Biology

Structureand Functionof Organisms

EvolutionaryBiology

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Table of Contents

Overview

Kit Summary ....................................................................................................................1

Kit Inventory Checklist ......................................................................................................3Syllabus Alignment ..........................................................................................................5

Instructors Manual ..........................................................................................................6

Background................................................................................................................6

Pre-Lab Activity........................................................................................................26

Instructors Advance Preparation Guide..........................................................................28

Lesson 1: Protein Extraction From Muscle ..............................................................28

Lesson 2: Electrophoresis Gel Loading, Running, and Staining ............................30

Lesson 3: Destain and Dry Gels ..............................................................................32

Post-Lab Activity ......................................................................................................33

Quick Guide ..................................................................................................................42

Student Manual ..............................................................................................................46

Background..............................................................................................................47

Pre-Lab Activity........................................................................................................52

Alternate Pre-Lab Activity ........................................................................................56

Lesson 1: Introduction to Protein Electrophoresis and SDS-PAGE ..........................60

Protein Extraction From Muscle ..............................................................64

Lesson 2: Electrophoresis Gel Loading, Running, and Staining ............................67

Lesson 3: Destai