1. DNA REPLICATION 3.4 CORE 3.4 CORE

2. 3.4.1 Explain DNA replication in terms of unwinding the double helix an separation of the strands by helicase, followed by the formation of the new complementary strands by DNA polymerase. Cells must prepare to divide by doubling the DNA content of the cell in a process called DNA replication. This process doubles the quantity of DNA and also ensures that there is an exact copy of each DNA molecule. DNA replicates during interphase of the cell cycle.

3. Molecules needed for Replication Among the variety of molecules present in the nucleoplasm are two types that are particularly important for the process of DNA replication; they are Enzymes needed for replication- these include helicase and group of enzymes collectively called DNA polymerase. Free nucleotides – these are nucleotides that are not yet bonded and are found floating freely in the nucleoplasm, some contain adenine some thymine, some cytosine and some guanine.

4. First step in Replication One of the early events of DNA replication is the separation of the double helix into two single strands. The double helix is held together by the hydrogen bonds between complementary base pairs. The enzyme helicase begins at a point in or at the end of a DNA molecule and moves one complementary base pair at a time, breaking the hydrogen bond so the double stranded DNA becomes two separate strands.

5. --- The first major step for the DNA Replication to take place is the breaking of hydrogen bonds between bases of the two antiparallel strands. The unwounding of the two strands is the starting point. The splitting happens in places of the chains which are rich in A-T. That is because there are only two bonds between Adenine and Thymine (there are three hydrogen bonds between Cytosine and Guanine). Helicase is the enzyme that splits the two strands. The initiation point where the splitting starts is called "origin of replication".The structure that is created is known as "Replication Fork".

6. Template strand formation The unpaired nucleotide on each of these single strand can now be used as a template to help create two double stranded DNA molecules identical to the original.

7. Formation of two Complementary Strands Once DNA has become unzipped, the nitrogenous bases on each of the single strands are unpaired. In the nucleoplasm, there are many free-floating nucleotides. These nucleotides are available to form complementary pairs with the single-stranded nucleotides of the unzipped molecule. This does not happen in a random fashion. A free nucleotide locates on one opened strand at one end and then a second nucleotide can come in to join the first. This will require that these nucleotides become covalently bonded together as they are beginning of a new strand. The formation of a covalent bond between two adjoining nucleotides is catalysed by one of the DNA polymerase enzymes.

8. --- A third nucleotide then joins the first two and the process continues in a repetitive way for many nucleotides. The other unzipped strand also acts as a template for the formation of another new strand. This strand forms in a similar fashion, but in the opposite direction to the first strand. One strand is replicating in the same direction as helicase is moving and the other strand is replicating in the opposite direction.

9. 3.4.2 Explain the significance of complementary base pairing in the conservation of the base sequence of DNA. The pattern of DNA replication ensures that two identical copies of DNA are produced from one. The area where replication has taken place, the two strands are absolutely identical to each other. This is because the original double- stranded molecule had complementary pairs of nucleotides and it was the complementary nucleotides that used the unzipped single- stranded areas as templates.

10. Replication is Semi onservative No DNA molecule is ever completely new. Every DNA molecule after replication consists of a strand that was ‘old’ now paired with a strand that is ‘new’. DNA replication is described as semiconservative process because half of pre- existing DNA molecule is always conserved. http://www.wiley.com/college/pratt/0471393878/student/a nimations/dna_replication/index.html http://www.wiley.com/college/pratt/0471393878/student/a nimations/dna_replication/index.html

11. Diagram of DNA Replication Refer to page number IB Biology by C.J Clegg Page Number 67

12. DNA Replication Video (Cartoon Video, a good one for review)

13. Replication To initiate DNA synthesis, a small segment of RNA, called an RNA primer complementary to the template DNA is synthesized by an unique RNA polymerase known as primase. It is to this primer, that DNA polymerase adds 5’ deoxyribonucleotides and extend the DNA. DNA polymerase can function only in 5’->3’ direction. While on the one strand, the DNA synthesis is continuous and in 5’-> 3’ direction, on the other strand, DNA is synthesized in small stretches resulting in discontinuous DNA synthesis. This happens in the opposite direction to the first strand. Such a process is also referred to as semi-discontinuous replication.

14. The short stretches of DNA, each primed by RNA are called Okazaki fragments named after the Japanese scientist who discovered them. RNA primers are then removed, and the gap is filled by DNA synthesis. Both steps are performed by DNA polymerase. These fragments are then sealed by the enzyme Ligase. The strand which supports the continuous DNA synthesis is the leading strand and the one which is replicated in short stretches is called the lagging strand.

15. Leading and Lagging Strand

16. Leading Strand The elongation process is different for the 5'- 3' and 3'-5' template. a)5'-3' Template: The 3'-5' proceeding daughter strand -that uses a 5'-3' template- is called leading strand because DNA Polymerase ä can "read" the template and continuously adds nucleotides (complementary to the nucleotides of the template, for example Adenine opposite to Thymine etc).

17. Lagging Strand b)3'-5'Template: The 3'-5' template cannot be "read" by DNA Polymerase ä. The replication of this template is complicated and the new strand is called lagging strand. In the lagging strand the RNA Primase adds more RNA Primers. DNA polymerase å reads the template and lengthens the bursts. The gap between two RNA primers is called "Okazaki Fragments". The RNA Primers are necessary for DNA Polymerase å to bind Nucleotides to the 3' end of them. The daughter strand is elongated with the binding of more DNA nucleotides.

18. ---- In the lagging strand the DNA Pol I -exonuclease- reads the fragments and removes the RNA Primers. The gaps are closed with the action of DNA Polymerase (adds complementary nucleotides to the gaps) and DNA Ligase (adds phosphate in the remaining gaps of the phosphate - sugar backbone). Each new double helix is consisted of one old and one new chain. This is what we call semiconservative replication. If you need a simpler answer for DNA replication you can make use of IB BIOLOGY By Andrew Allot (Oxford) Page number 16.

19. Discontinuous DNA Replication