Gene Cloning and DNA Analysis: An Introduction, Sixth Edition ...

Chapter 2 Vectors for Gene Cloning: Plasmids and Bacteriophages 21
Figure 2.8
M13 phage attaches to
a pilus on an E.coli cell
and injects its DNA
New M13 phages are
continuously extruded
from an infected cell
The infection cycle of bacteriophage M13.
Figure 2.9
Capsid components
and assembly
M13 phage
Pilus
M13 DNA
M13 DNA replication
Daughter cells continue
to release M13 particles
b2 region
(non-essential) Integration
Infected cells continue
to grow and divide
and excision
Early regulation
DNA synthesis
M13 phages
Late regulation
Lysis of the host
AWB C D EFZUVGT H M LKI J int xis exo clll N cl cro OP Q SR
0 2 4 6 8 10 12 1416 18 20 22 24 26 28 30 32 34 36 38 40 42 44 46 4849kb
The λ genetic map, showing the positions of the important genes and the functions of the gene clusters.
The second role of the cos sites is rather different, and comes into play after the
prophage has excised from the host genome. At this stage a large number of new e DNA
molecules are produced by the rolling circle mechanism of replication (Figure 2.10c),
in which a continuous DNA strand is “rolled off” the template molecule. The result
is a catenane consisting of a series of linear e genomes joined together at the cos sites.
The role of the cos sites is now to act as recognition sequences for an endonuclease
that cleaves the catenane at the cos sites, producing individual e genomes. This endonuclease,
which is the product of gene A on the e DNA molecule, creates the singlestranded
sticky ends, and also acts in conjunction with other proteins to package each
e genome into a phage head structure. The cleavage and packaging processes recognize
just the cos sites and the DNA sequences to either side of them, so changing the
structure of the internal regions of the e genome, for example by inserting new genes,
has no effect on these events so long as the overall length of the e genome is not altered
too greatly.