MBB 222 Problem Set 5 ANSWER KEY.pdf

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Molec Biol & Biochem
MBB 222
Frederic Pio

Problem Set 5 MBB 222 – Molecular Biology and Biochemistry Fall 2013 First Name: _____________________________________________________ Last Name: _____________________________________________________ Student Number: __________________________________ Tutorial Day and Time: ___________________________________________ Total marks for Problem Set =41 marks Your score = _______ INSTRUCTIONS  Problem set is due on Friday, November 29 , 2013 in class.  Hand in your problem set to your Teaching Assistant for marking in class.  Write your answers in pen. Answers written in pencil will not be accepted.  Write CLEARLY! Zero marks will be given to questions with answers that are not legible!!  This problem set is worth 2% towards your final mark in the course. QUESTIONS 1. Which of the following statements about the polymerase chain reaction (PCR) is false? (2 marks) A. DNA amplified by PCR can be cloned. B. DNA amplification is linear in magnitude. C. Newly synthesized DNA must be heat-denatured before the next round of DNA synthesis begins. D. The boundaries of the amplified DNA segment are determined by the synthetic oligonucleotides used to prime DNA synthesis. E. The technique is sufficiently sensitive that DNA sequences can be amplified from a single animal or human hair. 2. Current estimates indicate that ________ % of the human genome is translated into protein. (2 marks) A. less than 0.5% B. roughly 1.5% C. roughly 10% D. roughly 25% E. more than 50% 3. Which of the following is not a characteristic of the -terminator in E. coli? (2 marks) A. The -terminator has an ATP-dependent RNA-DNA helicase activity. B. The -terminator migrates in the 3' to 5' direction along RNA. C. The -terminator causes RNA polymerase release at a CA-rich sequence near the end of the transcript. D. ATP is hydrolyzed by the -terminator during the termination process. E. All of the above are characteristics of the -terminator. 4. The 5'-terminal cap structure of eukaryotic mRNAs is a(n): (2 marks) A. 7-methylcytosine joined to the mRNA via a 2',3'-cyclic linkage. B. 7-methylguanosine joined to the mRNA via a 5'  5' triphosphate linkage. C. 7-methylguanosine joined to the mRNA via a 5'  3' diphosphate linkage. D. N -methyladenosine joined to the mRNA via a 5'  5' phosphodiester bond. 6 E. O -methylguanosine joined to the mRNA via a 5'  5' triphosphate linkage. 5. By increasing the spacing between sequence 1 and sequence 2 in the leader peptide of the trp operon of E. coli, attenuation compared to the normal sequence is: (2 marks) A. unchanged. B. always decreased. C. only decreased in the absence of Trp. D. increased. E. Cannot be determined from the information given. 6. Which one of the following statements about eukaryotic versus prokaryotic gene regulation is not correct? (2 marks) A. Access to eukaryotic promoters is restricted by the structure of chromatin. B. Most regulation is positive, involving activators rather than repressors. C. Larger and more multimeric proteins are involved in regulation of eukaryotic transcription. D. Transcription and translation are separated in both space and time. E. Strong promoters in eukaryotes are generally fully active in the absence of regulatory proteins. 7. Fill out the remaining boxes in the table below with the following information: the PCR cycling step, the temperature at which the cycling step occurs at (in some cases a temperature range may be required), and a brief description of what happens at cycling step (state in one sentence)? (0.5 mark each x 6 boxes = 3 marks in total) PCR cycling step Temperature Brief description of what happens during step o ~95 C ANNEALING ~72 C ANSWER: PCR cycling step Temperature Brief description of what occurs during step DENATURATION ~95 C Disrupts hydrogen bonds between complementary bases, yielding single-stranded DNA molecules o o Allow primers to complementary base pair at specified site ANNEALING ~50 C to 65 C on the single-stranded DNA template DNA polymerase synthesizes a new DNA strand by adding EXTENSION ~72 C dNTPs that are complementary to the template in 5' to 3' direction 8. What are three advantages of automated DNA sequencing method over manual DNA sequencing (be specific when mentioning each of the advantages) (1 mark for each correct advantage x 3 = 3 marks in total). ANSWER: (Any of the three are fine…1 mark each)  ddNTPs are labeled with specific fluorescent dye for each base (e.g., ddATP is labeled with the green dye, ddCTP is labeled with the blue dye, ddTTP is labeled with the red dye and ddGTP is labeled with the yellow dye) which permits sequencing in a single reaction, rather than four separate reactions for each ddNTP as in the manual DNA sequencing method. The fluorescently labeled DNA fragments will move across a path of a laser which allow each color dye to fluoresce at different wavelengths (and therefore correctly determine the position of each base)  Automated DNA sequencing equipment can eliminate the need for radioactive isotopes to label DNA, thereby reducing the volume of low-level radioactive waste generated…the fluorescent dye is not an environmentally hazardous chemical and has no special handling or disposal requirements.  Time is saved due to not having to perform autoradiography or associated tasks required for working with radioactive materials such as radiation survey
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