University of Pittsburgh at Bradford Science In Motion Biology Lab 030 DNA Molecular Modeling Introduction: Chromosomes are structures that are found in the nucleus of eukaryotic cells and are composed of long strands of deoxyribonucleic acid (DNA). In 1953 James Watson and F. Crick designed a model of this complex chemical structure. They described the model as a double helix or spiral, composed of nucleotides. A nucleotide is composed of a phosphate unit joined to deoxyribose, a five carbon sugar and a nitrogen containing base. The DNA molecule is a double strand of possibly thousands of nucleotides bonded by their bases. There are four bases found in DNA. Two are purines, either adenine or guanine. The other two are pyrimidines, either thymine or cytosine. These bases are represented by code letters A, G, T, and C. These are the alphabet used by ribosomes in the process of protein synthesis. When these bases bind to each other the Adenine always binds to thymine A-T and the guanine always binds to the cytosine G-C. During interphase of the cell cycle the DNA can build an exact copy of itself, the process of replication. This is the cells way of passing on its genetic code from one generation to the next. During replication, the DNA “unzips” its two halves, separating at the sight where the bases were bound together. New complimentary bases attach to the now free bases and form two new strands of DNA. Each new strand is made of nucleotides from the original strand and new nucleotides that are attached to the original strand. A similar process is used in the process of protein synthesis called transcription. This process allows for a specific portion of the DNA strand to be used to send messages to the ribosomes where protein synthesis can occur. The process of transcription is similar to replication in that the portion of DNA that is needed to send the message to the ribosomes separates and new bases attach. This molecule is called ribonucleic acid (RNA). In RNA, uracil replaces thymine, thus the base codes for RNA, which is a single strand, is A, G, U, and C. This RNA molecule is then transferred out of the nucleus to the ribosomes where it is used to participate in the production of proteins. Objective: This lab is designed to allow students to examine the structure of DNA and RNA by building their own model. Students will model DNA replication. Materials: 12 – three prong “deoxyribose” centers (black) 3 – “Adenine” straws (red) 3 – “Thymine” straws (blue) 3 – “Uracil” straws (White) 6 – Hydrogen bond centers (white) 1 – long strand (gray) Adapted from Hubbard Scientific Inc. 12 – two prong “phosphate” centers (red) 3 – “Guanine” straws (gray) 3 – “Cytosine” straws (Short green) 1 – four prong center 24 – connectors (yellow) 3 – long posts (green) Procedure: Part 1 – DNA structure 1. In order to construct your segment of the DNA model, you will need; 12 – black centers 10 – red centers 3 – red straws 3 – blue straws 3 – gray straws 3 – short green straws 6 – white centers 20 – yellow straws 1 – stand (one long gray straw, three green posts, and a four prong center). 2. Construct a backbone by attaching a deoxyribose (black center) to a phosphate (red center) with a yellow straw (figure 1). 3. Construct a chain composed of 6 deoxyribose (black centers) and five red centers with yellow straws to construct one backbone. 4. Attach one of the nitrogen bases (red, blue, green, or gray) to the open deoxyribose bond in any sequence you choose. After you are done you will have completed one side of the DNA model. 5. Repeat steps 2 and 3 to construct the other side of the DNA model (figure 2). Adapted from Hubbard Scientific Inc. 2 6. Use the six white bond centers which represent hydrogen bonds between bases, to attach the left side of the model, making sure to attach only a red straw (adenine) to a blue straw (thymine) and a gray straw (guanine) to green straws (cytosine). Note: you will have pieces left. 7. Place the constructed segment of the DNA model on the stand by passing the long gray straw through the holes in the white hydrogen centers. Attach the four-prong center to serve as a stand. Twist the DNA structure gently to form a double helix spiral (figure 3). Insert digital photo of model here. Adapted from Hubbard Scientific Inc. 3 8. Present your completed model on the front of your lab table or desk for the instructor to view and answer the following questions on a separate sheet of paper. a. What is the general structure of the DNA molecule? b. Name the two molecules which alternate to make the side portions or “backbone” of a DNA molecule. c. What is the name of the specific molecule to which each nucleotide is attached? d. Name the molecules or parts of a nucleotide, which join by a hydrogen bond to attach to the double strand of DNA. e. If there are four thymine bases on your model, how many adenine bases will there be? Explain why. f. List the molecules on the left side of your model, list the bases that should be on the right hand side, how did you know what would be on the right side? Part 2 – DNA replication Your DNA model represents only a short portion of DNA in a chromosome, which is usually composed, of thousands of nucleotides. Although your model is only a short portion, its replication is the same as that of an entire chromosome during mitosis and meiosis. 1. Find another group to work with. 2. “Unzip” your DNA chain, that is, separate one side of the DNA strand from the other by removing one set of nucleotides from the white centers. 3. Have the other group do the same with their model. 4. Observe your two separate strands. The white centers should now be removes from either side since they represented a hydrogen bond that no longer exists. 5. Begin to attach new nucleotides to the separated strands making sure the bases that attach are complimentary to each other, A to T and C to G. 6. Note that a replicating DNA strand does not take its new parts from an existing DNA strand. It would receive these new deoxyribose sugars, phosphate and nitrogen bases from the nucleoplasm. The new pieces are constructed in the cytoplasm of the cell and move into the nucleus of the cell where DNA replication takes place 7. Present your completed model on the front of your lab table or desk for the instructor to view and answer the following questions on a separate sheet of paper. a. If you were to open the entire molecule along the hydrogen bonds, what bases would the left side attach to? b. Would the two new DNA molecules contain the same base pairs? c. Would the two DNA molecules be exact copies of each other? Explain. Adapted from Hubbard Scientific Inc. 4 Part 3 – RNA structure and transcription DNA ensures the exact replication of chromosomes, which are genetic “codes” of instruction for the entire cell. These instructions are sent out of the nucleus of a cell by another nucleic acid, ribonucleic acid, or RNA. It is similar to DNA in that it is also composed of nucleotides, however deoxyribose and thymine are NOT found in RNA. Deoxyribose is replaced by ribose and thymine is replaced by uracil Other than those two replacements the RNA molecule contains the same materials as DNA. 1. Based on the information given in the previous paragraph, describe what a messenger RNA molecule would look like for the left side of your DNA molecule. What would an RNA for the left side look like? 2. Return the original pieces to the other team. 3. Replace all the thymine (blue straws) with uracil (white straws) on your strand of new DNA, this is now a strand of messenger RNA. 4. The structure of RNA is similar to DNA with several exceptions. Answer the following questions on a separate sheet of paper. a. A base sequence of Adenine, Adenine, Adenine (AAA) in mRNA would only join in what sequence of tRNA? Using the table below, answer questions d-h. Amino Acid Serine Proline Leucine Glutamine Phenylalanine Valine Lysine tRNA Code ACU GGG AAU GUU AAA CAA UUU d. What specific amino acid is brought to the mRNA by tRNA molecule with the following sequence? (AAU) e. If a protein molecule consists of the following amino acid sequence: leucine, glutamine, tyrosine, serine, and leucine what would the sequence of tRNA molecule be. f. A ribosome receives the following mRNA message: AAA, GUU, GAA, and CGA, what is the sequence of the tRNA that would join the mRNA. g. What is the function of mRNA? tRNA? h. How can a mutation occur? Adapted from Hubbard Scientific Inc. 5