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Chapter 13
This document summarizes key aspects of DNA structure and function. It describes DNA as a double helix with complementary base pairing between adenine and thymine and cytosine and guanine. DNA stores genetic information, replicates semiconservatively using enzymes like DNA polymerase, and its sequence can be amplified via the polymerase chain reaction using thermal cycling. DNA replication proceeds from primer sites on the leading strand continuously but in fragments on the lagging strand.
Chapter 13
- 1. Chapter 13 DNA andIts Role in Heredity
- 2. Review of DNAStructure Four key features of DNA structure: • It is a double-stranded helix of uniform diameter • It is right-handed • It is antiparallel • Outer edges of nitrogenous bases are exposed in the major and minor grooves
- 3. Review of DNAStructure Complementary base pairing: • Adenine (A) pairs with thymine (T) by two hydrogen bonds • Cytosine (C) pairs with guanine (G) by three hydrogen bonds • Every base pair consists of one purine and one pyrimidine
- 4. Review of DNAStructure Two Copies of the same sequence, just reverse compliments
- 5. Four Important Functionsof DNA 1) Stores genetic information 2) Is susceptible to mutation 3) Is precisely replicated in cell division 4) Is expressed as the phenotype
- 6. DNA Replication Three possiblereplication patterns: • Semiconservative: Parent serves as a template and new molecules have one old and one new strand • Conservative: Original helix only serves as a template • Dispersive: Parent fragments serve as templates, assembling old and new parts into molecules
- 7. Semi-Conservative Replication Two steps in DNA replication: • The double helix is unwound, making two template strands • New nucleotides are added to the new strand at the 3′ end and joined by phosphodiester linkages. Sequence is determined by complementary base pairing
- 8. Strands Grow fromthe 3’ End
- 9. How does ReplicationInitiate A large protein complex called the “replication complex” interacts with the template strands. All chromosomes have a region called origin of replication (ori). Proteins in the replication complex bind to a DNA sequence in ori.
- 10. Main Components inDNA Replication Primase synthesizes RNA primers to start replication DNA polymerase (I and III) adds nucleotides to the 3′ end. DNA helicase uses energy from ATP hydrolysis to unwind the DNA. Single-strand binding proteins keep the strands from getting back together. DNA ligase “glues” together any gaps in the newly synthesized sequence
- 11. So how doesit work? Helicase will unwind the DNA Strand SS-Binding Proteins keep the strands apart Primase hops on and lays down a 10-20bp RNA primer DNA polymerase III recognizes the primers and continues to extend the growing strand by reading the complimentary base pairs DNA polymerase I hopes on and replaces RNA primer with DNA and proofreads the new sequence DNA Ligase then glues together any gaps
- 12. DNA Polymerase • ProofReading Ability • Can NOT start without a primer or existing template sequence
- 13. Not all DNAStrands are Equal • Leading Strand “Easy Replication” • Lagging Strand is slower and more difficult • Replication Fork
- 14. Leading Strand • Primeris created on the 3’ end of the existing sequence • Replication occurs following the Replication fork until it falls off the other end
- 15. Lagging Strand • Primeris created near the replication fork • The strand is then replicated moving away from the Replication fork • Forms Okazaki Fragments
- 16. After Replication… • DNApol I replaces all RNA primers with DNA • DNA ligase glues all Okazaki Fragments together • DNA pol can check for errors in sequencing (proofreading)
- 17. PCR Copies of DNAsequences can be made by the polymerase chain reaction (PCR) technique. PCR is a cyclical process: • DNA fragments are denatured by heating • Primers, plus dNTPs and DNA polymerase are added • New DNA strands are synthesized
- 18.
- 19. PCR PCR results inmany copies of the DNA fragment—referred to as amplifying the sequence. The base sequence at the 3′ end of the DNA fragment must be known. Complementary primers, about 15–30 bases long, are made in the laboratory.
- 20. PCR An initial problemwith PCR was its temperature requirements. The heat needed to denature the DNA destroyed most DNA polymerases. A DNA polymerase that does not denature at high temperatures (90 °C) was taken from a hot springs bacterium, Thermus aquaticus.
- 21. Steps in atypical PCR Cycle 1) Denature 2) Annealing of the Primers 3) Elongation of new sequence DNA Pol Leads to exponential amplification of your target…