Download DNA to Protein

DNA to Protein
AP Biology
Monday, Dec. 9th
• Objective: Students will learn the sequence and
importance of the various experiments that lead to
the discovery of the structure and function of DNA.
– Go Over Test
– Morgan’s Flies Debate DNA vs. Protein
– The Answer
•
•
•
•
•
Griffith’s Experiment
Oswald Avery
Hershey, Chase
Chargaff
Rosalind Franklin; Watson and Crick
• Recycled DNA Model
Nucleotide
• Polynucleotide?
• “We are machines built by DNA whose purpose is
to make more copies of the same DNA...This is
exactly what we are for. We are machines for
propagating DNA, and the propagation of DNA is a
self-sustaining process.
It is every living object's sole
reason for living.”
-Richard Dawkins, The Selfish Gene
Watson and Crick
• “It has not escaped our notice that the specific
pairing we have postulated immediately
suggests a possible copying mechanism for
the genetic material”
Tuesday, Dec. 10th
• Objective: Students will understand the structure
of DNA and the preliminaries of DNA replication.
– Complete Recycled Model of DNA (Due at the end of
the period)
•
•
•
•
•
•
•
Nucleotide built correctly
Correct sequence in the backbone
Base pairing correctly identified
Double helix structure
5’ and 3’ ends correctly orientated in antiparallel directions
Highlight a nucleotide
5 rungs of the ladder
Wednesday, Dec. 11th
• Objective: Students will understand the
process of DNA replication.
– Discussion
• Explain the experiment that revealed how the
original strand of DNA was used in the
replication of DNA.
Figure 13.10
Parent cell
(a) Conservative
model
(b) Semiconservative
model
(c) Dispersive
model
First
replication
Second
replication
Figure 13.9-3
A
T
A
T
A
T
A
T
C
G
C
G
C
G
C
G
T
A
T
A
T
A
T
A
A
T
A
T
A
T
A
T
G
C
G
C
G
C
G
C
(a) Parental
molecule
(b) Separation of parental
strands into templates
(c) Formation of new
strands complementary
to template strands
• Describe the structure of a nucleoside
triphosphate and determine the only way that
it can be added to a growing strand (See
Diagram). Name the type of enzyme that
completes this addition.
• http://www.youtube.com/watch?v=OnuspQG
0Jd0
• Compare and Contrast the Leading strand with the lagging
strand.
• Create a diagram to describe the process of DNA
Replication. Use in your diagram the following structures:
–
–
–
–
–
–
–
–
Origins of Replication
Replication Fork
DNA Polymerase
DNA Ligase
Primase
Helicase
Topoisomerase
Single-strand binding protein
Figure 13.16b-1
3
1 Primase makes
RNA primer.
5
Template
strand
3
5
Figure 13.16b-2
3
1 Primase makes
RNA primer.
5
Template
strand
3
3
5
RNA primer
for fragment 1
2 DNA pol III
makes Okazaki
fragment 1.
5
3
5
Figure 13.16b-3
1 Primase makes
3
RNA primer.
5
Template
strand
3
3
5
RNA primer
for fragment 1
2 DNA pol III
makes Okazaki
fragment 1.
5
3
5
3
3 DNA pol III
detaches.
5
Okazaki
fragment 1
3
5
Figure 13.16c-1
5
3
RNA primer for fragment 2
Okazaki
4 DNA pol III
fragment 2
makes Okazaki
fragment 2.
3
5
Figure 13.16c-2
5
3
RNA primer for fragment 2
Okazaki
4 DNA pol III
fragment 2
makes Okazaki
fragment 2.
3
5
5
3
5 DNA pol I
replaces RNA
with DNA.
3
5
Figure 13.16c-3
5
3
RNA primer for fragment 2
Okazaki
4 DNA pol III
fragment 2
makes Okazaki
fragment 2.
3
5
5
5 DNA pol I
3
replaces RNA
with DNA.
3
5
6 DNA ligase forms
5
3
bonds between
DNA fragments.
3
5
Overall direction of replication
Figure 13.17
Leading strand
template
Single-strand
binding proteins
Leading strand
Helicase
Overview
Origin of replication
Lagging
Leading strand
strand
Lagging strand
Leading strand
Overall directions
of replication
DNA pol III
5
3
3
Parental DNA
Lagging strand
template
Primer
5
3
Primase
5
DNA pol III
3
5
Lagging strand
DNA pol I
DNA ligase
3
5
DNA Replication
• http://www.youtube.com/watch?v=teV62zrm
2P0
Thursday, Dec. 12th
• Objective: Students will understand the
organization of DNA and chromosomes and
the modern use of DNA Technology.
– Bring Textbook and Reading guide Tomorrow.
– Label Flies – GRRRRRRRRRRRR!
– Discussion
• How has the process of DNA replication, in
eukaryotic cells, been linked to aging?
• Is DNA replication always perfect? How are
mistakes fixed?
Figure 13.19-3
5
3
3
5
Nuclease
5
3
3
5
DNA
polymerase
5
3
3
5
DNA ligase
5
3
3
5
On the White Board
• Diagram how DNA is packed into a Chromosome. Label
your diagram using the following terms:
–
–
–
–
–
–
–
–
–
–
DNA
Chromatin
Heterochromatin
Euchromatin
Histones
Nucleosome
Linker DNA
Chromosome
Positive Charge
Negative Charge
Figure 13.21
Chromatid
(700 nm)
DNA
double helix
(2 nm in diameter)
Nucleosome
(10 nm in diameter)
30-nm
fiber
Loops
H1
Histones
Histone tail
Scaffold
300-nm
fiber
Replicated chromosome
(1,400 nm)
How is this structure different in a prokaryotic cell?
• Would you eat food that has been genetically
modified (GMO)?
• Define Genetic Engineering, Gene Cloning,
Recombinant DNA.
• Identify possible uses for this technology
Figure 13.22
Bacterium
1 Gene inserted
into plasmid
Bacterial
chromosome
Cell containing gene
of interest
DNA of chromosome
(“foreign” DNA)
Plasmid
Recombinant
DNA (plasmid)
Gene of interest
2 Plasmid put into
bacterial cell
Recombinant
bacterium
3 Host cell grown in culture to form a clone of
cells containing the “cloned” gene of interest
Gene of
interest
Protein expressed
from gene of interest
Copies of gene
Protein harvested
4 Basic
Gene for pest resistance
inserted into plants
Gene used to alter bacteria
for cleaning up toxic waste
research
and various
applications
Human growth hormone
treats stunted growth
Protein dissolves blood clots
in heart attack therapy
• In five steps, layout the framework for
creating recombinant DNA and placing that
recombined DNA into a bacterial cell. Use all
relevant vocabulary.
Figure 13.23-1
Restriction site
5
3
GA AT TC
C T TA AG
DNA
3
5
1 Restriction enzyme cuts
the sugar-phosphate
backbones.
5
3
5
3
G
C
5
3
G
3
Sticky end
5
Figure 13.23-2
Restriction site
5
3
GA AT TC
C T TA AG
DNA
3
5
1 Restriction enzyme cuts
the sugar-phosphate
backbones.
5
5
3
G
C
3
G
5
3
3
Sticky end
5
5
3
2 DNA fragment added
G
from another molecule
cut by same enzyme.
Base pairing occurs.
5
3
3
5
3 5
3 5
G A AT T C
C T TA A G
G A AT T C
C T TA A G
5 3
5 3
3
One possible combination
5
Figure 13.23-3
Restriction site
5
3
GA AT TC
C T TA AG
DNA
3
5
1 Restriction enzyme cuts
the sugar-phosphate
backbones.
5
5
3
G
C
3
G
5
3
3
Sticky end
5
5
3
2 DNA fragment added
G
from another molecule
cut by same enzyme.
Base pairing occurs.
5
3
3
5
3 5
3 5
G A AT T C
C T TA A G
G A AT T C
C T TA A G
5 3
5 3
3
5
One possible combination
3 DNA ligase
seals the strands.
3
5
3
Recombinant DNA molecule
5
Gel Electrophoresis and PCR
Figure 13.24
Mixture of
DNA molecules of
different
sizes
Power
source
Cathode
Anode
Wells
Gel
(a) Negatively charged DNA molecules will move
toward the positive electrode.
Restriction fragments
(b) Shorter molecules are impeded less than
longer ones, so they move faster through the gel.
Figure 13.24a
Mixture of
DNA molecules of
different
sizes
Power
source
Cathode
Anode
Wells
Gel
(a) Negatively charged DNA molecules will move
toward the positive electrode.
Figure 13.25
Technique
5
3
Target sequence
Genomic DNA
1 Denaturation
3
5
5
3
3
5
2 Annealing
Cycle 1
yields 2 molecules
Primers
3 Extension
New
nucleotides
Cycle 2
yields 4 molecules
Cycle 3
yields 8 molecules;
2 molecules
(in white boxes)
match target sequence
• What if we could bring the passenger pigeon
or the woolly mamoth back from extinction,
should we?
Monday, Dec. 16th
• Objective: Students will understand the
connection of genes and proteins, and the
physical process of creating genes from
proteins.
– Discussion
DNA
RNA
Protein
Phenotype
• Label each cell as
prokaryotic or
Eukaryotic.
• Label each structure
and each process –
processes are in the
boxes.
Figure 14.4
Nuclear
envelope
TRANSCRIPTION
RNA PROCESSING
DNA
Pre-mRNA
mRNA
DNA
TRANSCRIPTION
mRNA
Ribosome
TRANSLATION
Ribosome
TRANSLATION
Polypeptide
Polypeptide
(a) Bacterial cell
(b) Eukaryotic cell
Figure 14.5
DNA
template
strand
3
5
A
C
C
A
A
A
C C
G
A
G
T
T
G
G
T
T
T
G G
C
T
C
A
5
3
TRANSCRIPTION
U G
mRNA
G
U U
U G
G
C
U
C
3
5
Codon
TRANSLATION
Protein
A
Trp
Amino acid
Phe
Gly
Ser
Second mRNA base
A
C
U
UUU
U
UUC
First mRNA base (5 end of codon)
UUA
C
Leu
UAU
UCC
UCA
UAC
Ser
UGU
Tyr
UGC
U
Cys
C
UAA Stop UGA Stop A
UCG
UAG Stop UGG Trp
G
CUU
CCU
CAU
U
CUC
Leu
CCC
CCA
Pro
CAC
CAA
CUG
CCG
CAG
AUU
ACU
AAU
AUC IIe
ACC
AUA
ACA
AUG
Met or
start
GUU
G
UCU
UUG
CUA
A
Phe
G
GUC
GUA
GUG
AAC
Thr
ACG
AAG
GCU
GAU
GCC
Val
AAA
GCA
GCG
GAC
Ala
GAA
GAG
CGU
His
Gln
CGC
CGA
C
Arg
CGG
AGU
Asn
Lys
Asp
Glu
AGC
AGA
A
G
Ser
Arg
U
C
A
AGG
G
GGU
U
C
GGC
GGA
GGG
Gly
A
G
Third mRNA base (3 end of codon)
Figure 14.6
3’-ATCGTATATTTTATGTACCATCAGTTTTGCATGCAATGCTTTATTTTTTTTTT-5’
Coding Segment:
5’(cap) GAUGGUAGACAAAACGUACGUUACGAAAUAAAAAAAAAA -3’
Figure 14.13
Small RNAs
Spliceosome
5
Pre-mRNA
Exon 2
Exon 1
Intron
Spliceosome
components
mRNA
5
Exon 1
Exon 2
Cut-out
intron
Tuesday, Dec. 17th
• Objective: Students will understand the how
the yolk becomes a chicken in an egg.
– Discussion
• Write the following DNA sequence on your white board:
– 3’-ATCGTATATTTTATGTACCATCAGTTTTGCATGCAATGCTTTATTTTTTTTTT-5’
• Initiation: On your DNA strand recreate the transcription initiation
complex. Include transcription factors, promoter region, start
point, TATA Box, RNA polymerase.
• Elongation: Transcribe your DNA Strand
• Termination: How is transcription terminated?
• Now that you have a piece of pre mRNA on your white board,
process the pre mRNA so that it is ready to leave the nucleus. 5’
cap, 3’ tail, Introns, Exons, UTR, Splicesome.
• What is the evolutionary significance of RNA processing.?
•
• DNA
3’-ATCGTATATTTTATGTACCATCAGTTTTGCATGCAATGCTTTATTTTTTTTTT-5’
• http://www.youtube.com/watch?v=u9dhO0iC
Lww
What would happen to the reading frame if the following occurred:
Base is substituted
Missense vs. Nonsense
Base inserted or deleted
Frameshift mutation
Wednesday, Dec. 18th
•Count Flies
1.
2.
3.
4.
Total Number
Expected offspring
Count and categorize
Data Analysis and write up tomorrow.
Thursday, Dec. 19th
• Objective: Students will understand how to
statistically analyze their fly data.
– Discussion
– Work Time
•
Chi Square Test
Dihybrid Crosses have 4 possible phenotypes
4 – 1 = 3 degrees of freedom
If the calculated chi square value is less than 7.82 you except your hypothesis
(normal dihybrid cross) unlinked autosomal. Null hypothesis accepted your
data meets the expected.
If the calculated chi square value is greater than 7.82 you must reject your
hypothesis.
Probability
0.05
Degrees of
Freedom
1
2
3.84
5.99
3
7.82
4
5
9.49
11.1
13.2
15.1
18.5
20.5
(= or less
than)
0.01
6.64
9.21
.001
10.8
13.8
11.3
16.3
Thursday, Jan. 10th
• Objective: Students will understand the steps
I protein synthesis
– Task Card
– Discussion
Friday, Jan. 11th
• Objective: Students will understand the
process of protein synthesis.
– Discussion
– Video.
Translation:
Give the function of the following terms in translation:
Codon, tRNA, anticodon, aminoacyl-tRNA synthetase, ribosome, a site, p site, e
site, Translocation,
Using your mRNA Strand diagram the process of Translation.
What would happen to the reading frame if the following occurred:
Base is substituted
Missense vs. Nonsense
Base inserted or deleted
Frameshift mutation
5’(cap) CAUGGUAGACAAAACGUACGUUACGAAAUAAAAAAAAAA -3’
Monday, Jan. 14th
• Objective: Students will understand the
process of protein synthesis.
– 4 square
– Video Translation
4 square
• Define Transcription.
– Name 1 Enzymes involved in transcription and its
functions.
– Use the following terms in a sentence:
• Promoter, transcription factors, RNA polymerase, terminator
• What is meant by the term RNA processing?
– What is the difference between an intron and a exon?
– What is the evolutionary significance of RNA processing?
4 square
• Write a haiku that describes the process of translation. (57-5).
• Draw a ribosome. Label: Large subunit, Small subunit, Asite, P-site, E-Site
– What is the function of a ribosome
• Draw a model of tRNA. Label anticodon, and amino acid.
– How did the amino acid and tRNA molecule come together.
• Draw and mRNA molecule attached to a ribosome include
the following:
– Growing polypeptide that is 4 amino acids long.
– tRNA in the A site matching your mRNA with an amino acid.
– What the tRNA would look like in the E site.
• Write 2 sentences that describe the process of translation.
Mutations
What would happen to the reading frame if the following occurred:
Base is substituted
Missense vs. Nonsense
Base inserted or deleted
Frameshift mutation
Wednesday , Jan. 8th
• Objective: Students will understand the
fundamental differences in gene regulation in
Prokaryotes vs. Eukaryotes.
– Fly Lab Due: Monday Jan. 13th
– No Quiz this Friday – Next Test – Final Exam –
Adding chapter on viruses.
– Go Over Quiz
– Discussion: Prokaryotes vs. Eukaryotes.
Thursday , Jan. 9th
• Objective: Students will understand the
fundamental differences in gene regulation in
Prokaryotes vs. Eukaryotes.
– Fly Lab Due: Monday Jan. 13th
– Discussion: Prokaryotes vs. Eukaryotes.
Friday, Jan. 10th
• Objective: Students will understand how
eukaryotic cells regulate protein synthesis and
connect this understanding to developmental
biology.
– Discussion
• Review the steps
in protein
synthesis
• Where can we
regulate gene
expression?
• Can you make a
hypothesis on
how each step
could be
regulated?
• Define chromatin,
histones, and
nucleosomes.
• How could DNA
packaging relate to
gene regulation?
• Histone Acetylation (COCH3)
– Nuetralizes charges on
histones tails..
– Releases nucleosome
binding.
– Makes the DNA available
for transcription
• Alternate histone
acetylation
• Methylation – opposite
affect
• Alternate patterns are
passed on in a way that
is not coded in
nucleotide sequence
(Epigenetic inheritance)
• General Transcription factors – common to most genes
• Turn on and Off: Control elements up stream from
coding DNA (gene)
• Enhancers: Non: Coding regions of DNA that initiate
transcription when activated.
• Activators: Proteins that bind to enhancers to activate gene
expression
• Repressors: Opposite activators.
Compare the roles of general and specific transcription factors in
regulating gene expression.
Suppose you compared the nucleotide sequences of the distal control
elements in the enhancers of three genes that are expressed only in
muscle cells. What would you expect to find? Why?
DNA sequences can act as “tape measures of evolution”. Scientist
analyzing the human genome sequence were surprised to find that
some of the regions of the human genome that are most highly
conserved (similar to comparable regions in other species) don’t
code for proteins. Propose a possible explanation for this
observation.
• http://www.youtube.com/watch?v=vizWoobt_Q
Coordinated Control
Alternate RNA Splicing
• mRNA degradation
– Enzymatic Shortening of Poly-A-tail (length)
– Initiates removal of 5’ cap
– Nuclease enzymes degrade mRNA
• miRNA – will bind to mRNA and degrade
molecule.
Initiation of Translation
• Regulatory proteins bind to specific sequences
within the 5’ UTR’s
• Activation or inactivation of protein factors
needed for initiation complex.
Protein Activation and Degradation
• Protein Must be activated – generally phosphorylated.
• Degradation: Marked with Ubiquitin and the taken
apart by proteasome
• Why would activation control of certain proteins be
important?
Monday, January 13th
• Objective: Students will understand the
connection of stems cells to embryonic
development to gene regulation.
– Collect Fly Lab
– Assignment: Read Chapter on Viruses: Chapter 17
– Reading: The Great Debate Over Stem Cell Research
•
•
•
•
What was the debate?
What are the two sides?
What do we need to know to be fully informed?
Preliminary opinion.
Where do Embryonic Stem Cells Come
From?
• http://www.youtube.com/watch?v=UgT5rUQ
9EmQ
How does this information inform the debate on
stem cells?
Adult Stem Cells
• Stems Cells = Pluripotent
– Can give rise to all specialized cells within an
organism.
– Why stem cells? What hasn’t happened yet?
– How do the cells know what to become?
Cytoplasmic Determinants: Not
homogenous (What are they?)
Induction
• Cells are
influenced
by signals
received by
nearby cells.
How cells differentiate
Using what we already know.
Example of Cell Differentiation in Muscle Cells.
Figure: 16.4 (pg. 314)
Discuss and explain.
Figure 16.4-1
Nucleus
Master regulatory
gene myoD
Other muscle-specific genes
DNA
Embryonic
precursor cell
OFF
OFF
Figure 16.4-2
Nucleus
Master regulatory
gene myoD
Other muscle-specific genes
DNA
Embryonic
precursor cell
Myoblast
(determined)
OFF
OFF
mRNA
OFF
MyoD protein
(transcription
factor)
Figure 16.4-3
Nucleus
Master regulatory
gene myoD
Other muscle-specific genes
DNA
Embryonic
precursor cell
Myoblast
(determined)
OFF
OFF
mRNA
OFF
MyoD protein
(transcription
factor)
mRNA
MyoD
Part of a muscle fiber
(fully differentiated cell)
mRNA
Another
transcription
factor
mRNA
mRNA
Myosin, other
muscle proteins,
and cell cycle–
blocking proteins
The potential of Cloning
Dolly the
sheep.
• What have been the problems? Why?
• How would this ability change the debate on
stem cell research?
• Do we use embryonic stem cells or not?
Cancer
• Based on what you know, what do you think
needs to happen to initiate cancerous growth
in cells.
– Make 2 specific hypothesis.
Cancer and Viruses
• What we know:
–
–
–
–
Viruses play a role in approx. 15% of cancers
Tumor Viruses – viral DNA into host DNA
Viral DNA initiates tumor growth (oncogenes)
In terms of viral survival why would a virus want to do
this?
– Viruses linked to: Lymphoma’s, cervical cancers, leukemia
• Proto-oncogenes – code for proteins that stimulate
normal cell growth and division.
• How do proto–oncogenes become oncogenes?
Proto-oncogene to a oncogene? Increase amount or activity
(Promote)
–
–
–
–
Translocation to a new more active promoter
Gene Amplification
Mutation in promoter or enhacer
Mutation that changes protein activity
• Tumor Suppressor Genes:
– Mutations that decrease normal activity – contribute to
cancer.
• Prevent accumulation of cancer causing mutations
• Control of adhesion of cells to each other
• Cell Signaling pathway that inhibit cell cycle.
• Two Examples:
– Ras proto-oncogene: 30% of human cancers
– Normal function vs. Mutation
• p53 tumor supressing gene
– Normal Vs. Mutation?
• http://www.youtube.com/watch?v=2jCYocd13
Lw
• Sequence of events in colon cancer:
– Multiple mutations and steps to develop
cancerous cells
– Telemorase?
Tuesday, Jan 15th
• Objective: Students will relate the processes
of DNA replication and Protein synthesis to
viruses and viral reproduction.
– Task Card
– Discussion
27 deaths in
Minnesota
FLU
•
•
•
•
What is it?
What is its Structure?
How does it work?
http://www.youtube.com/watch?v=cE0qdqoB
Fa8
Viruses
• Draw a typical virus label its structures?
– DNA or RNA
– Capsid, protein coat
– Animal - Viral envelopes derived from cell membrane
of host
• Diagram and explain a simplified reproductive cycle
of a virus.
• How is a virus different than a cell? Is a virus alive?
Wednesday, Jan 16th
• Objective: Students will be able to describe
the life cycle of various viruses
– Discussion
– Read
• Compare and Contrast the lytic and lysogenic
life cycle of bateriophages?
• In evolutionary terms which is the more
advantagous cycle?
• How is the viral envelope created in animal viruses?
• HIV is a retrovirus, why do they call it a
retrovirus? Why are retroviruses so hard to
cure?
Monday, Jan. 21st
• Objective: Students will understand basic mutations that occur in
the process of protein synthesis and investigate gene technology.
– Understand basic principles:
• DNA Cloning
• Polymerase Chain Reaction
• Gel Electrophoresis
– Test
Monday, Nov. 28th
• Objective: Students will understand the
importance, purpose, and process of DNA
Replication.
– Check Flies
– Partner Q & A to the Group
Q & A DNA Replication
1. Describe the structure of a nucleoside
triphosphate and determine the only way that it
can be added to a growing strand (See Diagram).
Name the type of enzyme that completes this
addition.
Q & A DNA Replication
4. Compare and Contrast the Leading strand with
the lagging strand.
5. Use the diagram to describe the process of DNA
Replication. Use in your description the
following structures or terms:
•
•
•
•
•
•
•
•
Origins of Replication
Replication Fork
DNA Polymerase
DNA Ligase
Primase
Helicase
Topoisomerase
Single-strand binding protein
Q&A
• How does DNA replication possibly contribute
to the aging process?
Tuesday, Nov. 29th
• Objective: Students will understand the process of
transcription and RNA processing.
• What is happening here? Stop, Think, Label, Describe.
Wednesday, Nov. 30th
• Objective: Students will understand the
process of RNA Splicing and Translation.
– Task Card
– Discussion
What happens next?
Build a working model of the
translation process.
• Must include:
– Ribosome
• Large and small subunit
• A, P and E site
– Outcome must be a
polypeptide 6 amino
acid long.
– Appropriate tRNA and
Amino Acids
– Read mRNA 5’ to 3’
– Codons and Antiocodons
– Moveable parts.
– What did I miss?
Thursday, Dec. 1st
• Objective: Students will understand basic mutations that occur in
the process of protein synthesis and investigate gene technology.
– Present working model of translation
– Review:
• Point Mutations
• Base-pair substitutions
– Missense vs. Nonsense
• Frameshift Mutations
– Insertions vs. Deletions
– Investigate and Understand basic principles:
• DNA Cloning
• Polymerase Chain Reaction
• Gel Electrophoresis
Test Starts Tomorrow
Wednesday, Dec. 10th
• Objective complete the fly lab.
– Count flies and complete lab.
Fruit Flies – Wild Type
• Red Eyes
• Brown Body color
• Regular Wings
Male vs. Female
Male
Female
P – Vestigal Females X Sepia Males
Apterous Females x Wild Males
Apterous Female x Sepia Male
White Eyed Females X Wild Males
Vestigial Female X Ebony Male
Vestigial Female X Sepia Male