Textbook Question
What are the advantages of using a restriction enzyme whose recognition site is relatively rare? When would you use such enzymes?
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Ch. 17 - Recombinant DNA Technology
Klug10th EditionEssentials of GeneticsISBN: 9780135588789
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Table of contents
- Ch. 1 - Introduction to Genetics16
- Ch. 2 - Mitosis and Meiosis28
- Ch. 3 - Mendelian Genetics37
- Ch. 4 - Modification of Mendelian Ratios53
- Ch. 5 - Sex Determination and Sex Chromosomes32
- Ch. 6 - Chromosome Mutations: Variation in Number and Arrangement37
- Ch. 7 - Linkage and Chromosome Mapping in Eukaryotes36
- Ch. 8 - Genetic Analysis and Mapping in Bacteria and Bacteriophages24
- Ch. 9 - DNA Structure and Analysis38
- Ch. 10 - DNA Replication29
- Ch. 11 - Chromosome Structure and DNA Sequence Organization22
- Ch. 12 - The Genetic Code and Transcription36
- Ch. 13 - Translation and Proteins27
- Ch. 14 - Gene Mutation, DNA Repair, and Transposition34
- Ch. 15 - Regulation of Gene Expression in Bacteria22
- Ch. 16 - Regulation of Gene Expression in Eukaryotes37
- Ch. 17 - Recombinant DNA Technology27
- Ch. 18 - Genomics, Bioinformatics, and Proteomics30
- Ch. 19 - The Genetics of Cancer21
- Ch. 20 - Quantitative Genetics and Multifactorial Traits37
- Ch. 21 - Population and Evolutionary Genetics45
Chapter 17, Problem 13
In a typical PCR reaction, describe what is happening in stages occurring at temperature ranges
(a) 92-26 °C
(b) 45-65 °C and
(c) 65-75 °C
Verified step by step guidance1
Identify the three main stages of a PCR cycle, each corresponding to a specific temperature range: denaturation, annealing, and extension.
For the temperature range 92-96 °C (noting the problem states 92-26 °C, which likely means 92 °C down to 26 °C, but typically denaturation occurs around 92-96 °C), explain that this is the denaturation step where the double-stranded DNA template is heated to separate into single strands by breaking hydrogen bonds.
For the temperature range 45-65 °C, describe the annealing step where the temperature is lowered to allow primers to bind or anneal to their complementary sequences on the single-stranded DNA templates.
For the temperature range 65-75 °C, explain the extension (or elongation) step where the DNA polymerase enzyme synthesizes new DNA strands by adding nucleotides to the primers, extending the DNA sequence complementary to the template strand.
Summarize that these three steps repeat cyclically to exponentially amplify the target DNA sequence during PCR.
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Key Concepts
Here are the essential concepts you must grasp in order to answer the question correctly.
Denaturation in PCR
Denaturation occurs at high temperatures (around 92-96 °C), where the double-stranded DNA melts into single strands by breaking hydrogen bonds between bases. This step is essential to separate the DNA template so primers can bind during the next stage.
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Traditional vs. Next-Gen
Annealing of Primers
Annealing happens at moderate temperatures (typically 45-65 °C), allowing primers to bind or hybridize to their complementary sequences on the single-stranded DNA. The exact temperature depends on primer sequence and length, ensuring specificity of binding.
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Extension or Elongation
Extension occurs at an optimal temperature for DNA polymerase activity (usually 65-75 °C), where the enzyme synthesizes a new DNA strand by adding nucleotides complementary to the template strand, extending from the primer.
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Related Practice
Textbook Question
In the context of recombinant DNA technology, of what use is a probe?
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Textbook Question
If you performed a PCR experiment starting with only one copy of double-stranded DNA, approximately how many DNA molecules would be present in the reaction tube after 15 cycles of amplification?
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Textbook Question
We usually think of enzymes as being most active at around 37°C, yet in PCR the DNA polymerase is subjected to multiple exposures of relatively high temperatures and seems to function appropriately at 65–75°C. What is special about the DNA polymerase typically used in PCR?
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Textbook Question
Traditional Sanger sequencing has largely been replaced in recent years by next-generation and third-generation sequencing approaches. Describe advantages of these sequencing methods over first-generation Sanger sequencing.
520
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Textbook Question
How is fluorescent in situ hybridization (FISH) used to produce a spectral karyotype?
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