FA23 week 3 Problem Set

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Arizona State University *

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108

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Biology

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Apr 27, 2024

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docx

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BICD100 FA23 Week 3 Problem Set Question numbers are color coded to show which ones are based on Monday lecture material vs. Wednesday or Monday + Wednesday lecture material 1). The RNA sequence of the first six codons of a gene are: AUG AGA GAA UGG ACG AUG Use the codon table on textbook pg. 533 to answer the following questions. a. What amino acid sequence is encoded by these six codons? b. Which of the following could introduce a nonsense mutation that truncates this polypeptide? For each one you select (i.e. each one where your answer is yes), write the altered 18 nucleotide RNA sequence, show what was changed/added/deleted, and underline the new stop codon. i. Point mutation/substitution altering codon 3: ii. Point mutation/substitution altering codon 5: iii. Insertion of 3 nucleotides between codons 3 and 4: iv. Insertion of 1 nucleotide between codons 1 and 2: v. Deletion of the first 2 nucleotides of codon 3: c. Which of the mutations listed in part b, above, creates a frameshift? d. Which of the mutations listed in part b, above, are likely to be obtained via CRISPR-Cas9 genome editing of this sequence? e. What DNA repair mechanism would be involved in creating the mutation type(s) you identified in part d, above? 2). IL2RG encodes a subunit of a cytokine receptor that plays a role in the development of the immune system. Mutations in this gene cause approximately 30% of inherited immunodeficiency conditions (severe combined immunodeficiency disorder or SCID) in humans. The IL2RG gene is composed of 8 exons. Assume that all the mutations described below cause loss of gene/protein function. For each of the following pairs of mutations, indicate which one you think is more likely to be a null and which a leaky allele, or even a silent mutation if applicable (causing no disease). You will want to consult a codon table for some of these. Explain your reasoning for each answer in 1-2 sentences, including use of appropriate terminology such as silent, nonsense, missense, frameshift, SNP, indel, etc. a. Single nucleotide insertion in exon 1 vs. exon 7: cont’d 1

b. Deletion of two vs. three nucleotides in exon 4: c. Single nucleotide change in a codon in exon 1 from UAU to UAC vs. UAG: d. An inversion of exons 2-5 vs. an inversion of part of the cis-regulatory module that reduces but does not eliminate expression of this gene: 3). Which of the mutations described in question 2, if any, is/are likely to be the result of each of the following types of DNA damage or mutagen exposure? Refer to letters a-d used in Q2. a. a base mispairing event during DNA replication: b. UV irradiation: c. X-ray irradiation: 4). Which of the mutations described in question 2 (if any) could have been repaired via… a. mismatch repair: b. proofreading, i.e. the 3’ exonuclease activity of DNA polymerase: c. nucleotide excision repair: 5). A geneticist uses CRISPR-Cas9 editing to introduce mutations into a gene containing the following sequence in one of its exons. The CCG in red is a “PAM” sequence that is required for Cas9 cleavage, and the underlined portion is targeted for cleavage by the guide RNA: One of the mutant alleles obtained has a 4bp deletion as illustrated here: b. Draw the double-strand DNA molecule, containing the complete 28 nucleotide sequence shown at the top, after it was cut by Cas9 showing the overhangs at both ends of the cut that were trimmed off prior to NHEJ to create this 4bp deletion. Highlight the overhanging sequences that get trimmed off. 2