Chapter 6
Molecular
Biology of DNA
Replication and
Recombination
Jones and Bartlett Publishers © 2005
Replication of DNA by strand separation and copying of the 2
template strands using A-T and G-C base pairing (semiconservative replication)
Experimental proof of semi-conservative
replication of DNA
Use of a thymidine analog (BUdR) provides
cytological proof that DNA in chromosomes also
replicates semi-conservatively
Replication of a circular DNA molecule
through a qstructure
Replication can be uni- or bi-directional
During q-form replication, both parental DNA strands remain
intact. DNA replication begins at special sequences called “origins
of replication”. A single DNA molecule may have one (E. coli
chromosome) or many origins (a human chromosome can have
more than a thousand origins).
Replication of a circular DNA molecule
by a “rolling circle” mechanism
During rolling circle replication, one of the template DNA
strands is cut to create a primer 3’-OH end.
For every round of replication, the tail of the
rolling circle becomes one unit longer
One strand of
the rolling
circle grows
continuously
while the other
is made in
small pieces
(Okazaki
fragments).
Replication of a linear eukaryotic chromosome
Model of an E. coli DNA replication fork
showing the many proteins that play a role there
Prevention of knotting of DNA (as strands are
separated) by DNA gyrase working ahead of the
DNA replication fork
The components that are different in
RNA relative to DNA
Priming of DNA synthesis with and RNA segment
New DNA chains are initiated by
short RNA primers
Addition of a deoxynucleotide
to the 3’-OH end of a primer chain
A misinserted deoxynucleotide is excised by the
proofreading exonuclease function
of DNA polymerase
One side of the fork grows continuously (leading side)
while the other side grows by making small
DNA pieces (lagging side)
Sequence of events in the joining of adjacent
precursor fragments in eukaryotes
Addition of a dideoxynucleotide to the 3’-OH end
of a DNA chain terminates chain elongation
The chain termination method of DNA sequencing
DNA sequencing machines use
fluorescent dideoxynucleotides
A “shotgun sequencing” approach
for sequencing of a large DNA molecule
Analogs of Normal deoxynucleotides
useful as anti-AIDS agents
Excision repair of a mismatched base pair
Variations from normal 2:2 segregation (gene
conversion) can result from mismatch repair
Symmetric model of homologous recombination
showing some of the intermediate
steps and structures
Note the 2 nicks in
the same location
in the two duplexes
in step (B). This is
followed by strand
exchange and
Holliday junction
formation (C)
An asymmetric model of homologous
recombination
Recombination is
initiated by a cut in
only one of the 2
recombining duplexes
(A) followed by strand
invasion (B) and
Holliday junction
formation (C)
A recombining molecule showing the parental and the
recombinant duplexes joined at the Holliday junction
Resolution of a Holliday junction can
occur by 2 alternative cleavage pathways
Model for the recombinational repair
of a double strand break in DNA