Salty sweat due to

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Symptoms of cystic fibrosis
Salty sweat due to altered salt secretion in
sweat ducts
Mucus-clogged Airways Severe Respiratory
infections
Lung
Pancreas
Infertility in males due to clogged sex ducts
Testis
Cell lining ducts of the body
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• Cystic Fibrosis
• A single faulty protein is connected to the
  symptoms
• In 1989 the gene was mapped to chromosome 7

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Our Goals

• To determine the connection between the symptoms
  associated with cystic fibrosis and DNA How?
• Learn how DNA replicates
• Understand the genetic code and how the
  instructions in a gene are used to make a protein
  

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Central Dogma of Biology
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DNA and RNA Polymers of Nucleotides

• Nucleotide base sugar phosphate

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Fig 10.2
7
T
A single strand of DNA
A
C
Sugar- phosphate backbone
G
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• DNA vs. RNA Nucleotides
• Four nucleotides are found in DNA
• Differ in their nitrogenous bases
• Adenine (A), Thymine (T), Cytosine (C), Guanine
  (G)
• Sugar
• Four nucleotides are found in RNA
• Uracil instead of Thymine
• Adenine (A), Uracil (U), Cytosine (C), Guanine
  (G)
• Sugar

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• DNA is like a rope ladder twisted into a spiral

• DNA Structure
• Consists of 2 strands joined together by weak
  hydrogen bonds
• Rungs of the ladder are hydrogen bonded N-bases

Twist
Fig 10.4
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Base pairing in DNA
Figure 10.5
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What is a gene?

• The kind of proteins an organism makes helps to
  determine its phenotype

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The order of bases in a gene determines......

• The sequence of amino acids in the protein it
  codes for, which determines......
• the organism's phenotypethe physical and
  biochemical characteristics of an organism.

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DNA replication

• Questions to answer
• When during the cell cycle does it occur?
• What do we start with and end with?
• Where does it occur in a cell?
• Whats needed for it to occur?
• What is the sequence of events?
• Why is it said to be semi-conservative?
• What does proofreading mean?
• What does the proofreading?

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DNA Replication
Fig 10.6
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1
3
4
2

1. Parent molecule has 2 complementary strands of
   DNA
2. Enzymes initiate the breaking of the H-bonds,
   separating the double helix
3. Free nucleotides base pair to parent strands
   (A-T, G-C) using DNA polymerase enzyme
4. Each daughter strand consists of one parental
   strand and one new strand

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• DNA can be damaged eg. by ultraviolet light
• The enzymes (e.g. DNA polymerase) can repair the
  damage
• Is the damage always repaired? Consequences?

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The Flow of Genetic Information DNA to RNA to
Protein

• Transcription
• Translation ribosomes translate mRNA into
  proteina chain of amino acids

Fig 10.9
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• CF phenotype
• Genes determine which proteins a cell can make
• Proteins control phenotype
• e.g. CFTR Gene codes for CFTR protein

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CFTR Protein The cystic fibrosis transmembrane
regulator protein

• CFTR Protein
• Pumps chloride ions (salt) into cells lining
  ducts or the lungs
• What are the consequences when CFTR doesnt work?

Carbohydrate
Cytoplasm of cell lining duct or lungs
Chloride ions
CFTR Protein
Water
Inside of duct or Air sac in lungs
Water
Cell membrane
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The order of bases in a gene determines the order
of amino acids in the protein it codes for
Fig 10.10
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Why are proteins so important?

• Enzymes catalysts for nearly all chemical
  reactions in cells Determine what cells can make
  and digest
• Structural components
• Receptors on cell surface
• Hormones e.g. insulin, growth hormone, prolactin
• Transport e.g. hemoglobin, spindle fibers
• Immune system antibodies

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Transcription copying DNA into RNA

• Questions to answer
• 1. What do we start with and end with?
• 2. Where does transcription occur? When?
• 3. What is needed for transcription to occur?
• 4. What is the sequence of events?

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An RNA Nucleotide
Phosphate
Base (Uracil, U)
Sugar ribose
This oxygen is absent in deoxyribose
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RNA

• Base pairing
• Single stranded
• Sugar ribose (one more oxygen than deoxyribose)

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Stages of Transcription
Fig 10.13b
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Fig 10.13a
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Transcription

1. Initiation - RNA polymerase enzyme binds to the
   promotor (section of DNA indicating start of a
   gene)
2. Elongation RNA polymerase catalyzes base
   pairing on the template strand (U-A, G-C)
3. Termination RNA polymerase reaches the stop
   sequence and the new mRNA is released.
4. mRNA processing non-coding regions of the mRNA
   are removed and the mRNA leaves the nucleus.

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Fig. 10.14 Step 4 mRNA processing
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Translation Ribosomes reading mRNA to produce a
polypeptide

• Questions to answer
• What do we start with and end with?
• 2. Where does translation occur?
• 3. What is needed for translation to occur?
• What is the sequence of events?
• What are the roles of mRNA, ribosomes, start
  codon, tRNA, anticodons, stop codon?

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How do ribosomes read the code?
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The genetic code Fig 10.11 Codon 3 letter
section of mRNA that codes for one amino acid
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Transfer RNA tRNA
Fig 10.15

• tRNA
• Matches amino acids with codons in mRNA using
  anticodons

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A portion of an mRNA molecule attached to a tRNA
Codon on mRNA
mRNA
Each Codon specifies a specific tRNAamino
acid complex
Amino acid
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Stages of translation
1. Initiation mRNA start codon binds to tRNA
anticodon Ribosome binds to both
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2. Elongation

• tRNA brings specific AAs to the ribosome as mRNA
  passes through the ribosome (codon anticodon
  recognition)

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(No Transcript)
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3. Termination Ribosome reads an mRNA stop
codon (no tRNA with anticodon). mRNA and protein
detach from the ribosome
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Explaining the symptoms of CF

• Mucus build-up in the lungs
• Lung infections (e.g. pneumonia)
• Male sterility (blocked vas deferens)
• Salty sweat
• Trouble digesting food (blocked pancreatic duct)

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Explaining the symptoms of CF

• In CF, the faulty CFTR protein never makes it to
  cell membrane

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Understanding Cystic Fibrosis at the Cellular
Level

• How does CFTR protein get from where its
  produced to its home in the cell membrane?
• Where is the CFTR protein produced?
• Where does it go for modification?
• How does it get there?
• How does the modified CFTR protein get to the
  plasma membrane?
• The defective CFTR protein is recognized at the
  ER as defective
• Where is the defective CFTR protein sent?

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CF symptoms may be mild or severe
CFTR Gene
Several hundred different mutations are
associated with CF
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Whats a Mutation?

• Any change in the nucleotide sequence of DNA
• Types of Mutations
• Mutations may Result from
• Random errors in DNA replication
• Viruses
• Chemicals/toxins (cigarette smoke)
• Radiation (e.g. U.V. light, X-rays)

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DF508 deletion the most common cause of cystic
fibrosis
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Mutation responsible for Sickle Cell Anemia
nucleotide substitution
Glu
Val
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3 Types of Mutations Base Substitutions,
Insertions or Deletions

• Base substitutions
• May result in changes in the amino acid sequence
  in a protein, or
• May be silent (have no effect) Why?

Substitute C for U
Met
Lys
Phe
Gly
Ala
AACGGUUAUCCU
Asn Gly Tyr Pro
Met
Phe
Ser
Ala
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• Types of Mutations Base Insertions and
  deletions

• Changes the reading frame of the genetic message

mRNA
Protein
Met
Lys
Phe
Gly
Ala
(b) Nucleotide deletion
Met
Lys
Leu
Ala
His
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• Although mutations are often harmful

• They are the source of the rich diversity of
  genes in the living world
• They contribute to the process of evolution by
  natural selection