1. DNA and the Genome
Key Area 2a
Replication of DNA

2. Learning Intentions By the end of this topic you should be able to:
State 5 things that must be present for DNA replication
Describe the stages in DNA replication
Describe what a primer is and explain its role in DNA replication
Name 2 enzymes in involved in DNA replication
Explain the role of each enzyme in DNA replication
Describe the direction of replication on each DNA strand
Explain why the direction of DNA replication is always in this direction

3. You should already be able to:
Name the molecules in a DNA nucleotide and identify them in a diagram
Name the type of bond on the backbone of the DNA molecule
Give the names of the 4 DNA bases
Describe the base pairing rule for DNA bases
Describe the role of hydrogen bonds in the DNA structure
State the name of the coiled structure adopted by DNA

4. Why do cells need to copy their DNA?
Prior to cell division, DNA is replicated to ensure that new cells have the same number of chromosomes and to ensure that all cells have the same genes.

5. Importance of DNA Replication
DNA is unique as it can replicate and reproduce itself exactly
It is the molecule of inheritance and is passed from one cell to the next during growth and from parent to offspring during reproduction
DNA is essential for the continuation of life
The process of replication is complex and involves different enzymes including DNA POLYMERASE AND LIGASE

6. Semi- conservative Replication
As the two new molecules formed during replication each contain one strand from the original parent molecule replication is said to be ‘semi-conservative’.
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7. Nucleic Acids There are two types of nucleic acid you need to know:
• Deoxyribonucleic acid (DNA) which serves as a cellular database by storing an immense amount of information about all the polypeptides (proteins) a cell can potentially make.
• Ribonucleic acid (RNA) which occurs in several different forms (messenger RNA, ribosomal RNA, transfer RNA) and is needed to convert DNA information into polypeptide sequences (proteins)

8. Requirements for DNA Replication
For DNA to replicate the following are required:
DNA to act as the template; provides the code to copy
Primers to start replication process and allow attachment of DNA polymerase
Nucleotides all 4 types to produce the new DNA strands
Enzymes Helicase (unwinds DNA); DNA polymerase (pair up free nucleotides); Ligase (join Okazaki fragments
ATP to supply the energy for all of the above processes

9. DNA Replication Enzymes
The 2 main enzymes required for DNA replication are:
DNA Polymerase: adds DNA nucleotides (using complimentary base pairing) to the deoxyribose (3’) end of the new DNA strand that is forming. This means the enzyme can only elongate DNA in a 5’ 3’ direction
Ligase: joins fragments of DNA together

10. Primers
The enzyme responsible for adding nucleotides onto a chain is DNA polymerase but to work it can only add them to a pre-existing chain
DNA polymerase needs PRIMERS to start replication
A primer is a short strand of nucleotides which binds to the 3’ end of the template DNA strand allowing polymerase to add nucleotides

11. Specific primers bind to the 3’ end of the template DNAG T G A C 3’ 5’
STEP 1: DNA is unwound and hydrogen bonds between the bases break to form 2 template strands.
Specific primers bind to the 3’ end of the template DNA
DNA parental strand composed of two complementary strands 3’ 5’ 5’ A C T G T G A C 3’ 3’ 5’

12. A C T G T G A C A C A T C G G T
STEP 2: Free DNA nucleotides start to line up with complementary nucleotides T C A C G T A G

13. STEP 3: Sugar-phosphate bonds form. C T G T G A C A C T G T G A C
STEP 3: Sugar-phosphate bonds form.
Two DNA molecules identical to the parental molecule have been formed.

14. The leading strand
DNA polymerase can only add nucleotides in one direction (5’ to 3’)…
…therefore the leading strand is replicated continuously

15. After the DNA unwinds and thehydrogen bonds break, the DNA unzips.
A DNA primer (a short stretch of complementary DNA) attaches to the start of the template DNA.
DNA polymerase then attaches free nucleotides to the 3’ end of the primer.
This continuous process till leading strand is copied.

16. Replication of the leading strand of DNA
5’ end of DNA strand
Leading strand of replicated DNA
DNA polymerase enzyme
Direction of replication
Start of complementary strand of replicated DNA
Replication of the leading strand of DNA
3’ end of DNA strand
Primer

17. The lagging strand
As DNA polymerase can only add onto the 3’ end of a primer, the lagging strand is replicated in fragments and these are then joined together

18. Replication of the lagging strand of DNA
Many primers attach along the strand.
These are extended by the DNA polymerase.
The fragments (Okazaki fragments) are then joined by the enzyme ligase
This is a discontinuous process creating the lagging strand.
5’ end of DNA strand
3’ end of DNA strand
Primer
DNA polymerase
Ligase

19. Replicating Both Strands…
The two DNA strands are antiparallel so one strand runs from 3’ ‘ (called the LEADING strand) and the other from 5‘ ‘ (called the LAGGING strand
DNA polymerase can only synthesize new DNA in the 5‘ ' direction (directionality). This poses special problems for replicating double-stranded DNA.
The leading strand is continuously synthesised but the lagging strand has to be synthesised in fragments (Okazaki Fragments) and then these fragments joined together by the ligase

20. New leading strand DNA being synthesised Primer DNA Polymerase
Step 1: DNA unwinds forming a replication fork and a short primer bonds to the 3’ end of the parental DNA strand by hydrogen bonds between complementary bases
Step 2: DNA polymerase adds free nucleotides onto the primer and extends the newly synthesising DNA in a 5’ ’ direction (bases between new and parent strand held together by hydrogen bonds)
New leading strand DNA being synthesised
Primer
DNA Polymerase

21. New lagging strand DNA being synthesised forming an Okazaki
Step 3: a primer binds to the 5’ lagging parental DNA strand and polymerase extends new DNA in 5’ ’ direction forming an Okazaki Fragment (again, hydrogen bonds hold new and parental strands together)
New lagging strand DNA being synthesised forming an Okazaki
fragment
Primer

22. Step 4: The replication fork further unwinds and the leading strand continues to be continuously synthesised
Step 5: A second primer binds to the lagging strand and DNA polymerase extends a second Okazaki fragment (and over writes the first primer)
Step 6: Ligase enzyme will join the 2 Okazaki fragments together and the process is continued
Step 7: The 2 new double strands rewind producing the 2 daughter DNA molecules

24. Replication bubbles and forks

25. When copying a long chromosome many replication forks operate simultaneously to speed up the replication process.