➢ A few years after solution of the double

LESSON 2 helix, the

mechanism of semiconservative replication
DNA REPLICATION was demonstrated by Matthew Meselson and
Franklin Stahl using the technique of
equilibrium density centrifugation on a
cesium gradient.
DNA synthesis proceeds in the 5 to 3 direction,
➢ The order of nucleotides is maintained
DNA polymerase, the enzyme responsible for
because each strand of the parent double
polymerizing the nucleotides, uses a guide, or
helix is the template for a newly replicated
template, to determine which nucleotides to add.
strand.
The enzyme reads the template in the 3 to 5
direction. The resulting double strand, then, will ➢ Human DNA replicates about 50 bases
have a parent strand in one orientation and a newly per second. To get the job done, a human
synthesized strand oriented in the opposite chromosome replicates simultaneously at
orientation. hundreds of points along its length, and the
pieces join. A site where DNA is locally
• Each cell division cell must copy its entire opened resembling a fork, is called a
DNA. replication fork.
• So each daughter cell gets a complete copy
Note: Ligase comes from the Latin word
At first, some researchers suggested that DNA meaning “to tie”.
might replicate in any of three possible ways:
1. semiconservative, ➢ This replication process can be observed
2. conservative, with one double helix by ELECTRON MICROSCOPY as a
specifying creation of FORKED STRUCTURE OR
a second double helix, and REPLICATION FORK.
3. dispersive, with a double helix shattering ➢ OKAZAKI FRAGMENT- small pieces
into pieces that would join with newly of DNA about 1000 bases in length.
synthesized DNA pieces to form two
molecules. DNA REPLICATION PROCESS

SEMICONSERVATIVE REPLICATION 1. DNA replication occurs during

➢ Predicted by Watson and Crick; It is the __________ of the cell cycle.
key to maintain the sequence of the
nucleotides in DNA through new 2. When DNA replicates, it unwinds, breaks,
generations. builds a new nucleotide chain, and mends
➢ Every cell in a multicellular organism or Enzymes called helicases unwind and hold
in a clonal population of unicellular apart replicating DNA, enabling other
organisms carries the same genetic enzymes to guide the assembly of a new
information. It is important that this DNA strand.
information, in
the form of the DNA sequence, be 3. DNA replication begins when a helicase
transferred faithfully at breaks the hydrogen bonds that connect a
every cell division. The replication base pair. Binding
apparatus is designed to copy the DNA proteins hold the two strands apart. Another
strands in an orderly way with minimal enzyme, primase, then attracts
errors before each cell division. complementary RNA nucleotides to build a

MTE 124: MOLECULAR BIOLOGY and DIAGNOSTICS

Prepared by Liel Reyes, RMT & Karen Joy B. Tanguilan, RMT
 short piece of RNA, called an RNA primer, correct DNA bases. Yet another enzyme,
at the called an annealing helicase, rewinds any
start of each segment of DNA to be sections of the DNA molecule that remain
replicated. The RNA unwound.
primer is required because the major
replication enzyme, DNA polymerase 10. Finally, ligases seal the entire sugar-
(DNAP), can only add bases to an existing phosphate backbone.
nucleic acid strand.

4. The RNA primer attracts DNAP, which

brings in DNA nucleotides complementary
to the exposed bases on the parental strand;
this strand serves as a mold, or template.
New bases are added one at a time, starting
at the RNA primer.

5. The new DNA strand grows as hydrogen

bonds form between the complementary
bases. The nucleotides are abundant in cells
and are synthesized from dietary nutrients.

6. DNAP works directionally, adding new

nucleotides to the exposed 3 ′ end of the
sugar in the growing strand.
Overall, replication proceeds in a 5 ′ to 3 ′
direction, because this is the only chemical
configuration in which DNAP can add
bases.

7. Next, an enzyme called a ligase then seals

the sugar- phosphate backbones of the
pieces, building the new strand. These
pieces, up to 150 nucleotides long, are called
Okazaki fragments, after their discoverer.

8. DNA polymerase also “proofreads” as it

goes, excising mismatched bases and
inserting correct ones. It also removes the
RNA primer and replaces it with the correct
DNA bases. Yet another enzyme, called an
annealing helicase, rewinds any sections of
the DNA molecule that remain unwound.

9. DNA polymerase also “proofreads” as it

goes, excising mismatched bases and
inserting correct ones. It also removes the
RNA primer an replaces it with the

MTE 124: MOLECULAR BIOLOGY and DIAGNOSTICS

Prepared by Liel Reyes, RMT & Karen Joy B. Tanguilan, RMT