Transcript S. DNA Replication

Construct a DNA model Using Base-pairing Rules.
Homework: 25 Pts. Due tomorrow to gain access to DNA Extraction Lab.
Template Strand
2 rings
3 bonds
1 ring
Directions:
Template Strand: _G_ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __
1) Answer Review Questions on back:
1. What is the basic subunit for DNA? Complementary Strand:___ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __
2. What are the three parts?
3. Which parts are the backbone?
4. Which nucleotides are purines?
Nucleotide Bank
5. Which nucleotides are pyrimidines?
6. How do you tell the difference between the two?
7. What type of bond holds together DNA strands?
2) Cut out the nucleotides from the Nucleotide Bank (right).
3) Match them up with the template strand above based
upon base-pairing rules then tape/glue beneath.
4) Determine the sequence (order of the string of
nucleotides read left to right) for both the template
strand and the new complimentary strand. Label the
sequence of the template and complimentary strand on
the lines with the nucleotide abbreviation.
Agenda:
O Discussion: DNA Replication
-------------Warm Up-----------------------------------------------1. What is DNA replication?
2. What is the goal of replication?
3. What stage of the cell cycle does replication occur in?
----------HW: -----------O Review your outlines on DNA & Replication (CH12).
O Re-read outlines on CH9: Cell cycle, Chromosomes and
Mitosis.
O If you were absent last week at all, you need to see me
towards the end of class.
OBJECTIVES
Restate what replications is and why it’s
important.
Illustrate the steps of replication in a drawing.
Describe the steps of DNA replication.
Compare the roles of DNA helicase, DNA
polymerase, and ligase.
Compare the process of DNA replication in
prokaryotes and in eukaryotes.
VOCABULARY
DNA replication
DNA helicase
DNA polymerase
DNA SYNTHESIS
These subunits are
comprised of a
phosphate group, a
ribose sugar, and
one of 4
nitrogenous bases.
In the last section you
learned what DNA is.
You should also know that
DNA is a large molecule
built of billions of subunits.
You should also recall that:
A bonds with T and
G bonds with C.
CELLS DON’T LAST FOREVER
Every cell in an organism has a
particular life span… they don’t
last forever.
The process is a cell is born, grows
and does its job, divides, and
creates two new daughter cells.
The life span is cyclical & with
every new generation the cells
that are formed carry out the
functions of the parent cell
because they are given each an
exact copy of the parent cell’s
DNA.
The cyclical life span of a cell is
called the CELL CYCLE.
THE CELL’S LIFE. THE CELL CYCLE
DNA SYNTHESIS = DNA DUPLICATION
Every time a cell goes through a
cycle it must duplicate its DNA
so that when it makes new
baby cells they both have the
exact same DNA.
The stage of the cycle that this
happens is in the “S” phase.
This is the stage we’re
focusing on.
In this section we are going
explore the process of DNA
synthesis, called DNA
replication.
This process is extremely
precise and an incredibly
important.
THE CELL CYCLE: THE PART
WE’RE ISOLATING.
From single-copy of
each chromosome
•To double-copy
WHAT’S DIFFERENT ABOUT THESE CELLS?
1st Gap Phase
DNA Synthesis Phase
REPLICATION
•These are identical strands of DNA
•There are two copies needed because each are destined for the 2 identical new cells
that are formed, called daughter cells.
•Identical DNA must be synthesized in order to achieve this.
•The process of DNA synthesis is REPLICATION.
HOW IS DNA REPLICATED?
Before it was concluded
how it happened
exactly, there were
several suggested ways
DNA was replicated.
Conservative
Semi-conservative
Dispersive
The Meselson-Stahl experiment:
•Showed that DNA is replicated semi-conservatively.
•This means, the new strands formed are half new
DNA, half old DNA
DNA REPLICATION: THE STEPS
The process of replicating DNA is broken down into three
major steps.
Your job is to know these 3 steps and be able to summarize
what happens in each.
The three steps are: (leave several spaces after each)
1.
2.
a.
b.
3.
Unwinding and Separating DNA Strands
Adding complimentary bases
Leading Strand:
Lagging Strand: RNA Primers attach
Formation of Two Identical DNA molecules
DNA REPLICATION
• Step 1:
• DNA helicases unwind the DNA
double helix.
• These proteins wedge themselves
between the two strands of the double
helix and break the hydrogen bonds
between the base pairs.
• Forms Replication Forks
• As the double helix unwinds, the two
complementary strands of DNA
separate from each other and form a Y
shape.
DNA REPLICATION
Step 2:
New nucleotides are added
by the enzyme DNA
Polymerase according to
the base-pairing rules.
Proteins called DNA
polymerases catalyze
the formation of the new
complimentary DNA
molecule by moving
along each strand in a 3’
to 5’ direction.
They add nucleotides to a
new daughter
compliment in a 5’ to 3’
direction only!
DNA REPLICATION
You know that DNA is anti-parallel
because of the bi-directionality
of DNA.
One side goes 3’ to 5’.
The other 5’ to 3’.
DNA polymerase can only add
nucleotides in a 5’ to 3’
direction.
This means the new nucleotides
can only be added to the 3’ end
of the existing chain.
One side (the top pictured here)
has its new strand continuously
synthesized as helicase
unwinds more DNA. This is the
LEADING STRAND.
The other side, the LAGGING
STRAND (on bottom) is
discontinuous replication
because it the bases are
oriented in the wrong direction.
DNA REPLICATION
Step 2: Leading strand
Primers attach.
There exists small premade sequences of RNA
called primers. These bind to
complementary regions of
the original DNA once it’s
separated.
• DNA Polymerase attach
free nucleotides to the 3’
end of the primer.
• These serve as tethers
from which replication can
proceed from the 3’ end.
• On the leading strand there
is one primer and the newly
forming DNA follows the
replication fork as the new
DNA is synthesized.
•
•
•
DNA REPLICATION
•
•
•
•
•
Step 2: Lagging strand
The lagging strand
creates a particular
problem for replication.
Since DNA can only be
synthesized from the 3’
end of the primers, the
lagging strand would be
left incomplete.
Multiple primers attach
to the lagging strand as
the replication fork
moves forward, creating
what are known as
“Okasaki Fragments”
These are usually 12000 nucleotides long.
DNA REPLICATION
•
•
•
Step 2: Lagging strand
If you notice, the Okazaki Fragments are not joined
together.
A special enzyme, call ligase, joins Okasaki fragments
together to form one continuous molecule.
Ligase
Ligase
DNA REPLICATION, CONTINUED
Step 3:
The process completes when
all the original bases have
been paired with a new
complementary nucleotide.
Each double-stranded DNA
helix is made of one new
strand of DNA and one
original strand of DNA.
DNA REPLICATION
Click to animate the image.
REPLICATION PROTEINS…QUALITY CONTROL
• Replication involves many proteins that form a machine-like
complex of moving parts.
• These proteins play a key role in making sure that the process
is flawless.
• DNA Helicase unwinds DNA carefully so it doesn’t get torn.
• Ligase is a protein that ensures proper bonding of growing
Okasaki fragments.
• DNA polymerase adds complimentary nucleotides.
• DNA polymerase also has a “proofreading” function.
• During DNA replication, errors sometime occur and the wrong
nucleotide is added to the new strand.
• If a mismatch occurs, the DNA polymerase has the amazing
ability of being able to backtrack, remove the incorrect
nucleotide, and replace it with the correct one.
• This decreases the chances of the wrong DNA being made,
lessening our chances of CANCER!
VISUAL CONCEPT: DNA REPLICATION
REPLICATION V TRANSCRIPTION
You may notice that replication has similarities to
TRANSCRIPTION
Replication is not the same process as Transcription, however.
In transcription, a new molecule of RNA is made from the DNA.
In DNA replication, a new molecule of DNA is made from the
DNA.
SUMMARY
What is DNA replication?
Why does it happen?
Know the steps…
Where is…
a. Leading Strand
b. Lagging Strand
c. Okazaki fragments
d. DNA Ligase
e. Replication Fork
f. DNA Polymerase
g. All 3’ & 5’ ends.
h. Where helicase should be
i. Where the replication fork is.
Item
Points
Step1: DNA unwinds
1
DNA Helicase
2
Correct DNA Sequence
2
Direction Replication is proceeding.
1
Step 2: New DNA forming
1
Leading Strand
1
Lagging Strand
1
Replication Fork
1
RNA Primer
1
Okasaki Fragment
1
DNA Polymerase
2
Free Nucleotides
1
Step 3: New DNA complete
1
5’ end all DNA
2
3’ end all DNA
2
New DNA (color blue)
1
Old DNA (color red)
1
DNA Ligase
2
Exact Copies
1
Total
25
DNA Replication Drawing Scoring Rubric
• You are responsible for understanding the
process of replication and being able to
identify the 3 steps as well as identifying
all the key components involved.
• Your assignment is to draw all 3 steps in
the process of replication in one drawing
(template provided).
• You must show each stage of replication
in the illustration, labeling all the steps,
proteins, landmarks, and & showing what
DNA is where according to the rubric.
• This is worth 25 points.
Replication Enzymes
DNA Polymerase
Helicase
Ligase
OBJECTIVES
Primary
Compare the process of DNA replication in prokaryotes
and in eukaryotes.
Identify the features of prokaryotic and Eukaryotic
replication on an illustration.
Secondary
Compare the number of nucleotides replicated in
eukaryotes and prokaryotes by calculating the rates of
replication.
PROKARYOTIC AND EUKARYOTIC REPLICATION
All cells have chromosomes, but eukaryotes and
prokaryotes replicate their chromosomes
differently.
The main difference between prokaryote and
eukaryote replication is how many start sites each
have.
The start sites then regulate how replication
proceeds in each organism type.
PROKARYOTIC DNA REPLICATION
Recall the structure of prokaryotic DNA.
Does anyone remember what the structure is?
Prokaryotic cells usually have a single
chromosome which is a closed loop
attached to the inner cell membrane.
Replication in prokaryotes begins at a single
site along the loop. This site is called the
origin of replication.
PROKARYOTIC REPLICATION
Two replication forks begin at the origin of
replication.
Replication occurs in opposite directions
until the forks meet on the opposite side
of the loop.
The result is two identical loops of DNA.
EUKARYOTIC REPLICATION
Eukaryotic cells often have several chromosomes which are
linear and contain both DNA and protein.
 How many chromosomes do humans have?
 What are the proteins called that hold DNA in nucleosomes?
Eukaryotic replication starts at many sites along the
chromosome.
This process allows eukaryotic cells to replicate their DNA
faster than prokaryotes.
EUKARYOTIC REPLICATION
Two distinct replication forks form at each start site and
replication occurs in opposite directions.
This process forms replication “bubbles” along the DNA molecule.
Replication bubbles continue to get larger as more of the DNA is
copied.
The replication bubbles keep growing until they join together with
other bubbles and complete replication.
Replication is complete when two identical complementary
strands of DNA is formed.
EUKARYOTIC REPLICATION
Original DNA
Replication bubbles
Replication bubbles joining
Original + new DNA
New + original DNA
PROKARYOTIC AND EUKARYOTIC REPLICATION
Even the smallest eukaryotic chromosomes are often 10 times
the size of a prokaryotic chromosome. Eukaryotic
chromosomes are so long that it would take 33 days to
replicate a typical human chromosome if there were only
one origin of replication.
As such, evolution has allowed Human chromosomes to
replicate in about 100 sections that are 100,000
nucleotides long, each section with its own starting point.
 Remember, the 46 human chromosomes laid end to end would measure
≈ 2m. Bacterial chromosomes measure ≈ 0.25cm!
Because eukaryotic cells have multiple replication forks
working at the same time, an entire human chromosome
can be replicated much faster, in only about 8 hours.
Bacteria replicate their small genomes in minutes…
IN-CLASS ACTIVITY: REPLICATION COMPARISON
•Label the letters in your notes.
•Save this paper for a follow-up activity that will be turned in at the end of
class.
•Word-bank: Original DNA (x2), New DNA (x2), Replication Forks (x2),
Replication bubble
•What is the difference between prokaryotic replication & eukaryotic
replication?
•Why are they different?
A
C
E
B
G
F
D
SUMMARY
In DNA replication, the DNA molecule unwinds, and the two
sides split. Then, new bases are added to each side until
two identical sequences result.
The replication of DNA involves many proteins that form a
machinelike complex of moving parts.
In prokaryotic cells, replication starts at a single site. In
eukaryotic cells, replication starts at many sites along the
chromosome.
IN-CLASS EXERCISE/HOMEWORK
Each student is responsible for understanding the process of
replication and being able to identify the 3 steps. In
addition, all must be able to identify all the key proteins &
molecules involved.
Your assignment is to draw all 3 steps in the process of
replication in one drawing. You must show the 3 stages of
replication in one illustration, labeling all the steps,
proteins, landmarks, and & showing what DNA is where.
This is worth 25 points.
GET A HAND-OUT… THIS IS HOMEWORK
REFLECTION
Discuss with your groups & summarize in your notes
what you learned today.
1. What are the steps of eukaryotic replication?
2. How can you tell the difference between prokaryotic &
eukaryotic replication?
3. What is the end product of replication?
SUMMARY… THE MORE COMPLEX VERSION