MARINE BIOTECHNOLOGY & BIOINFORMATICS FOR TEACHERS MOSS LANDING MARINE LABS NSF ITEST GRANT TEACHER LESSON PLAN FOR CLASSROOM USE TREE OF LIFE Designed by Treena Joi tjoi@psvd.net Background The purpose of this lesson is to bridge the concepts of genetics and evolution for Middle or High School level students and provide a hands-on experience with phylogenetic trees. Given a group of organisms students will estimate their relatedness and make a branching phylogenetic tree by hand. In the computer lab, students will be provided with single letter amino acid protein sequences of the same or similar organisms and use the multi-sequence online program ClustalW (http://clustalw.genome.jp/) to create a genetically based phylogenetic tree. Other organisms of interest are also included so students can predict the evolutionary relationship by making a tree using several vertebrate and invertebrate species before they use the online tool. This activity will introduce students to the field of bioinformatics by utilizing the ClustalW program for a practical application. The sequences will be provided for students in this activity but connections can be made to the biotechnology techniques employed to obtain sequences by purifying DNA. Description of Audience: This biotechnology/bioinformatics activity is designed for use by Middle or High School students studying in Grades 7-12. State Standards: This biotechnology/bioinformatics activity fulfills the following State of California Science Standards: Grade 7 1. Genetics 2.e 2. Evolution: 3.d 3. Investigation & Experimentation: 7.a,b,c Grades 9-12 1. Genetics: 4.d,e, 5.b 2. Ecology: 6.g 3. Investigation & Experimentation: 1.a,d,k National Standards: This biotechnology/bioinformatics activity fulfills the following National Science Standards: Grades 5-8 1. Content Standard A: Science as Inquiry 2. Content Standard C: Life Science 3. Content Standard E: Science and Technology Grades 9-12 1. Content Standard A: Science as Inquiry 2. Content Standard C: Life Science 3. Content Standard E: Science and Technology Copyright 2006, MBB – NSF ITEST Program, p. 1 STEM Connection: Bioinformatics careers are tied to this activity, such as contributing data to or developing software for biological analysis such as ClustalW. Such jobs could include Biologists, web programmers and database programmers. Biotechnology careers with reference to the source of the DNA sequences used may entail jobs in Paleontology, Comparative Anatomy, Science Teaching, Ecology and any of a number specialty fields such as Entomology, Mycology or Immunology. Technology Integration: N.A. Goals(s): The goal of this lesson is to: Make predictions Use an online database as an introduction to the field of bioinformatics Relate differences at the genetic/protein level to differences between organisms Learning Objective(s) Upon completion of this lesson, students will be able to conduct multiple-sequence analysis using an online database and produce a phylogenetic tree comparing several organisms: Properly navigate a computer to utilize an online database Conduct a multiple-sequence query using a database to produce a phylogenetic tree Relate the trees to evolutionary trends and hypothesize on the selective forces involved Understand the rationale behind modern Linnean classification of organisms Purpose/Rationale This lesson follows both the genetics & evolution units late in the school year. The evolution unit follows the genetics unit and this lesson is a culminating activity connecting both of those units as well as comparative anatomy and organ systems. The evolution unit is based on the Lawrence Hall of Science GEMS unit, “Life Through Time.” That unit has been modified to include several dissections of model organisms interspersed between the GEMS lessons. The rationale for this lesson is to connect the genetics and evolution units and associated concepts of proteins and inheritance with the dissections. It is hoped that by incorporating comparative anatomy and dissecting model organisms during an evolution unit the “Tree of Life” lesson will review material covered during the entire year. Comparing protein sequences to generate a phylogentetic tree will be a culminating activity to tie together prior studies of anatomy/organ systems and cell biology as well as making a direct link between genetics and evolution. CA state science standards addressed in this lesson include topics under the genetics, evolution and investigating/experimentation headings. National standards covered by this lesson include Life Science, Science as Inquiry, and Science and Technology threads. Materials/Resources In order to complete this lesson, the following materials are needed: Computer Lab or computers adequate for students working in groups Internet access http://align.genome.jp/ Student directions Teacher background notes Optional Power Point template for student presentation Electronic version of DNA or protein sequences of organisms for alignment on Clustal web site o To paste in, align, and generate tree Optional electronic versions of a different DNA or protein sequence from same organisms o (to compare resulting tree) Copyright 2006, MBB – NSF ITEST Program, p. 2 Prior Teacher Preparation Teachers may use the files provided for this lesson or modify the genes or organisms by following the notes provided in supplemental background materials. Directions are provided so that teachers or advanced students can research bioinformatics more in depth. In order to customize proteins or organisms in brief: See supplemental background files to ease navigating the necessary databases Decide on organisms to compare Use NCBI to obtain DNA or protein sequences (same gene) for each o http://www.ncbi.nlm.nih.gov/ Enter species name in search box and select gene, then search Create text file in FASTA format of the sequences o FASTA format sample: >Euglena_viridis GDAERGKKLFESRAGQCHSSQKGVNSTGPALYGVYGRTSGTVPGYAYSNANKNAAI VWEDESLNKFLENPKKYVPGTKMAFAGIKAKKDRLDIIAYMKTLKD (“>” symbol required with species and/or gene identification followed with one return and a text string of DNA bases or single letter amino acid code; must be file type .txt) Make file available to students Practice making the tree yourself and practice navigating the NCBI site to obtain sequences 3-Step Procedure #1 Introduction Introduce lesson by connecting the organisms used in the comparison to prior studies in evolution and genetics. Review the conventions involved in making a phylogenetic tree where branch length is directly related to how long ago an organism evolved relative to the others on the tree. Major differences between organisms require a new branch in the tree. Ask students what features might be used to determine how the organisms in question are related; make a predictive phylogenetic tree (optional: Let students identify the shared characteristics where branches are joined and then write the advanced character at the end of the branch). Review the role of genes in determining phenotypes and then the role of proteins. Students should be introduced to the term bioinformatics in this lesson and be able to understand the source of biological data in online databases. Teacher PowerPoint is an option to introduce this material and the task and may be one option for sharing results. Students may work in pairs of groups on the computers, depending on what is available. Students should make a predictive phylogenetic tree individually and compare the computer generated tree to theirs individually, in groups, or during a class discussion, depending on time available. Lesson rubric is designed so that students reflect on the lesson and share the trees they generate in a collaborative group PowerPoint presentation. #2 Exploration Students will predict evolutionary relatedness between several organisms of interest based on their intuition. Students will then use an online program to compare protein sequences provided to them for the organisms of interest. Students will be asked: Copyright 2006, MBB – NSF ITEST Program, p. 3 How closely do you think the organisms listed are related? Make a tree/cladogram to share your results. o Did your tree differ from the tree generated using the protein sequence and ClustalW program? Why or why not? o Would a different protein provide different results? Why or why not? o For advanced students, use the cytochrome b data and include the grasshopper sequence (from mitochondria). Most likely this will yield a tree where the insect is more closely related to humans than other vertebrates. Ask how this is possible and then go to NCBI using the accession number to read in detail the source of the sequences. o Are mitochondrial and genomic DNA and proteins different? How can this be an evolutionary advantage? How can scientists use these differences to conduct research and answer questions? Use the worksheet for students to record predictions, take notes, record the ClustalW tree, and answer questions or use science notebooks to respond to similar prompts. Students will revise their reflection on their original prediction by recording the tree generated online on the worksheet or in a science notebook. A class discussion may help students articulate their thoughts better. A PowerPoint presentation aligned to the worksheet is provided to allow students to collaborate as a group to present their results and ideas in front of the class. Conclude by asking students how this activity relates to their studies of cell biology, genetics, and evolution. A final assessment could be to have students write in their own words how DNA and proteins contribute to differences between organisms. Ask students what jobs may be involved with using or creating the data or software used in this activity. o #3 Application Students can predict and then see their position in the phylogenetic tree by including Homo sapiens in the sequences to be compared. Have students brainstorm how this and similar genetic databases might be of use socially, not just in a research field; bring up forensic analysis, and genetic disease screening. Students can be asked to generate their own list of organism that they want to determine evolutionary relatedness and if time allows or for AP biology, students may search NCBI to compare other organisms of interest. Guiding questions for student driven comparisons may include determining which terrestrial mammals are most closely related to marine mammals. Comparing amino acid/protein sequences from the same organisms may show different results due to the redundancy of codons; seemingly closely related organisms can be selected for this follow up; student should recognize that seemingly major differences in DNA sequences can yield similar if not identical proteins. Students should select an organism of interest to research in greater depth, describing its place in the Linnean classifcation scheme, from Kingdom to species. For more in depth research students could determine approximately which era or period it evolved, what its closest ancestors are thought to be, and describe the organisms life cycle and general ecological role. Career Connection: Bioinformatics careers are closely tied to this activity. Students can be shown how a research position may involve contributing data to a database such as those collected on the NCBI site. Students can also be made aware of the need for software engineers to develop software for biological analysis such as ClustalW, the databases used, or the protein visualization software. Such jobs could include biologists, biotechnicians, academic or private sector researchers, web programmers and database programmers. Biotechnology careers with reference to the source of the DNA sequences used may entail jobs in Paleontology, Comparative Anatomy, Science Teaching, Ecology and any of a number specialty fields such as Entomology, Mycology, and Marine Biology. Copyright 2006, MBB – NSF ITEST Program, p. 4 Assessment Observe and question students’ initial handmade phylogenetic trees to establish they are incorporating branch difference in length and number to express the desired degree of relatedness. Observe students use of the online database to produce a tree. Students will print out or sketch a copy to a notebook showing their final computer generated tree next to their predicted tree. Design a worksheet, journal recording, test, quiz, or performance-based activity for students to demonstrate what they have learned. For students having difficulty with the material, select fewer more distantly related organisms and create a special file that is simpler, has less data and shows obvious evolutionary branch differences – remove the emphasis from the branch length in discussions with these students. This lesson can be modified for additional activities as a class or as differentiation for individual students in several ways. Connections to the central dogma can be made by using an additional database to find the DNA sequences that code for the amino acid sequences used. Students could back translate one cytochrome c protein to mRNA and DNA by using a key to the single letter amino acid codes: o Advanced students would discover that more than one codon is possible for most amino acids. o Discuss the advantages of redundancy in translation. o Calculate the possible genetic variation for one protein sequence as a math extension. Teachers’ Self Evaluation • N.A. Copyright 2006, MBB – NSF ITEST Program, p. 5