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Bio A/H – Molecular Genetics Mistakes Happen!! Mutations are permanent changes in the DNA that can have a negative effect, a positive effect or no effect whatsoever. They can be caused by external environmental factors or simply when DNA polymerase makes a typo during replication. Since it is the DNA that is copied into RNA, this mistake will transfer to the RNA. Problems don’t usually arise, however, until a protein is made from the mutated RNA. DNA can replicate the mistake with out a problem and RNA can usually be transcribed without a problem. But when we translate the mistake we can wind up with a different protein or no protein at all. Since these are all happening in individual recipes called genes, all of these mutations are examples of Gene Mutations. We discussed larger chromosomal mutations in the last unit. What Kinds of Mutations Are There? (you are responsible for all of these) Point Mutation A point mutation is a simple change in which one base of the gene sequence is changed. A single base can be inserted, deleted or substituted. (although you will see it used synonymously with substitution) Example: Typing the word “Mog” or “Doog” when you wanted to type “Dog”. Frame-shift mutation In a frame shift mutation, one or more bases are inserted or deleted, the equivalent of adding or removing letters in a sentence. However since the RNA sequence is read three letters at a time, adding or removing one letter changes those groups of three, called the reading frame. That means, that each codon following the mutation is going to be read differently. Example: Remember there are no spaces separating codons in the DNA. So a sentence like “The cat ate the rat” really looks like “thecatatetherat.” We chose a reading frame to break up the words in threes. But let’s delete the “h” in the first word. This sentence becomes “Tec ata tet her at”. Each letter shifts over one and the sentence makes no sense. There are two mutations that cause Frame-shift mutations: Deletions and Insertions. If, however, we insert or delete 3 nucleotides (or 6, or 9) then we don’t shift the reading frame, we just add amino acids to our protein. This could still have dire consequences. (ex: The Bad Cat ate the rat) Other types of Mutations: - Nonsense Mutation: If a point mutation or frameshift leads to a pre-mature stop codon, the protein will end before it should. This protein will end up shortened, or truncated. This is called a nonsense mutation. - Silent Mutation: If a point mutation changes a codon to another codon that codes the same amino acid, then it will have no effect on the protein. This is called a silent mutation. Bio A/H – Molecular Genetics Directions: 1. For each type of mutation NEATLY put a line through the part of the original DNA sequence that you are mutating and replace the original sequence with your mutation. 2. NEATLY change the RNA to correspond to your mutated DNA 3. Write in the new Amino Acid sequence that is coded by your mutated DNA. You only need to rewrite the part(s) of the sequences that are affected by the mutation. 1. Substitution – In the sequence below, create a substitution mutation. Replace one nucleotide in the DNA with a different nucleotide Original DNA RNA Transcript: Original Amino Acids GCT ATG CTC GAT CCC TCG ATT CGA UAC GAG CUA GGG AGC UAA Arg Tyr Glu Leu Gly Ser STOP New Amino Acids 2. Deletion – Now create a single nucleotide deletion in the second codon of the DNA sequence. Original DNA RNA Transcript: Original Amino Acids GCT ATG CTC GAT CCC TCG ATT CGA UAC GAG CUA GGG AGC UAA Arg Tyr Glu Leu Gly Ser STOP New Amino Acids 3. Insertion – Now create a single nucleotide insertion in the first codon of your DNA. Original DNA RNA Transcript: Original Amino Acids GCT ATG CTC GAT CCC TCG ATT CGA UAC GAG CUA GGG AGC UAA Arg Tyr Glu Leu Gly Ser STOP New Amino Acids 4. Silent Mutations – time for a silent mutation. You’ll need to think about this one a little. Remember, what ever change you make, it must still code for the same amino acid. Start by looking at the codon charts to see which amino acids have multiple codons. Original DNA RNA Transcript: Original Amino Acids New Amino Acids GCT ATG CTC GAT CCC TCG ATT CGA UAC GAG CUA GGG AGC UAA Arg Tyr Glu Leu Gly Ser STOP Bio A/H – Molecular Genetics 5. Nonsense Mutations – last but not least, create a mutation in this sequence that changes the third codon in the DNA into a stop codon. Original DNA RNA Transcript: Original Amino Acids GCT ATG CTC GAT CCC TCG ATT CGA UAC GAG CUA GGG AGC UAA Arg Tyr Glu Leu Gly Ser STOP New Amino Acids Discussion Questions: 1. What types of mutation cause the biggest changes in the amino acid sequence? Explain. 2. Explain the evolutionary benefit to organisms having what is called a “redundant” code, or the fact that there are multiple codons that code for the same amino acid. 3. The protein that I did my research on is 481 amino acids long. How long is the RNA transcript for that protein? Careful……. 4. Let say that the protein above had a substitution that caused a nonsense mutation at base number 333. How many amino acids long would my protein be? 5. If there are multiple reading frames for every gene, how does the ribosome identify which reading frame is correct?