OF GENETIC INFORMATION
Microbiology, MBIM 650/720
explain the mechanisms of gene transfer in bacteria.
describe the nature of transposable genetic elements and
discuss the significance of gene transfer, transposable
genetic elements and plasmids.
OF GENETIC INFORMATION
E. coli (rod prokaryote) strains undergoing conjugation. One strain
has fimbriae © Dr
Dennis Kunkel, University of Hawaii. Used with permission
bacterial populations mutations are constantly arising due to errors
made during replication. If there is any selective advantage for a
particular mutation (e.g. antibiotic resistance), the mutant will
quickly become the major component of the population due to the rapid
growth rate of bacteria. In addition, since bacteria are haploid
organisms, even mutations that might normally be recessive will be
expressed. Thus, mutations in bacterial populations can pose a problem
in the treatment of bacterial infections. Not only are mutations a
problem, bacteria have mechanisms by which genes can be transferred to
other bacteria. Thus, a mutation arising in one cell can be passed on to
transfer in bacteria is unidirectional from a donor cell to a recipient
cell and the donor usually gives only a small part of its DNA to the
recipient. Thus, complete zygotes are not formed; rather, partial
zygotes (merozygotes) are formed.
genes are usually transferred to members of the same species but
occasionally transfer to other species can also occur. Figure 1
illustrates gene transfers that have been shown to occur between
different species of bacteria.
GENE TRANSFER MECHANISMS IN BACTERIA
is gene transfer resulting from the uptake by a recipient cell of
naked DNA from a donor cell. Certain bacteria (e.g. Bacillus,
Haemophilus, Neisseria, Pneumococcus) can take up DNA from the
environment and the DNA that is taken up can be incorporated into the
DNA size state - Double stranded DNA of at least 5 X
105 daltons works best. Thus, transformation is sensitive
to nucleases in the environment.
Competence of the recipient - Some bacteria are able
to take up DNA naturally. However, these bacteria only take up DNA a
particular time in their growth cycle when they produce a specific
protein called a competence factor. At this stage the bacteria are
said to be competent. Other bacteria are not able to take up DNA
naturally. However, in these bacteria competence can be induced in
vitro by treatment with chemicals (e.g. CaCl2).
Uptake of DNA - Uptake of DNA by Gram+ and Gram-
bacteria differs. In Gram + bacteria the DNA is taken up as a
single stranded molecule and the complementary strand is made in
the recipient. In contrast, Gram- bacteria take up double stranded
Legitimate/Homologous/General Recombination - After
the donor DNA is taken up, a reciprocal recombination event occurs
between the chromosome and the donor DNA. This recombination
requires homology between the donor DNA and the chromosome and
results in the substitution of DNA between the recipient and the
donor as illustrated in Figure 2.
Fig 2. General recombination. Donor DNA is
shown in red and recipient DNA in blue
requires the bacterial recombination genes (recA, B and C) and
homology between the DNA's involved. This type of recombination is
called legitimate or homologous or general recombination.
Because of the requirement for homology between the donor and host
DNA, only DNA from closely related bacteria would be expected to
successfully transform, although in rare instances gene transfer
between distantly related bacteria has been shown to occur.
Significance - Transformation occurs in nature and it can
lead to increased virulence. In addition transformation is widely
used in recombinant DNA technology.
is the transfer of genetic information from a donor to a recipient by
way of a bacteriophage. The phage coat protects the DNA in the
environment so that transduction, unlike transformation, is not
affected by nucleases in the environment. Not all phages can mediate
transduction. In most cases gene transfer is between members of the
same bacterial species. However, if a particular phage has a wide host
range then transfer between species can occur. The ability of a phage
to mediated transduction is related to the life cycle of the phage.
Generalized Transduction - Generalized transduction is
transduction in which potentially any bacterial gene from the donor
can be transferred to the recipient. The mechanism of generalized
transduction is illustrated in Figure 3.
Fig. 3 The mechanism of generalized transduction
Fig. 4 The mechanism of specialized transduction
that mediate generalized transduction generally breakdown host DNA
into smaller pieces and package their DNA into the phage particle
by a "head-full" mechanism. Occasionally one of the
pieces of host DNA is randomly packaged into a phage coat. Thus,
any donor gene can be potentially transferred but only enough DNA
as can fit into a phage head can be transferred. If a recipient
cell is infected by a phage that contains donor DNA, donor DNA
enters the recipient. In the recipient a generalized recombination
event can occur which substitutes the donor DNA and recipient DNA
(See Figure 2).
Specialized transduction - Specialized transduction is
transduction in which only certain donor genes can be transferred
to the recipient. Different phages may transfer different genes
but an individual phage can only transfer certain genes.
Specialized transduction is mediated by lysogenic or temperate
phage and the genes that get transferred will depend on where the
prophage has inserted in the chromosome. The mechanism of
specialized transduction is illustrated in Figure 4.
excision of the prophage, occasionally an error occurs where some
of the host DNA is excised with the phage DNA. Only host DNA on
either side of where the prophage has inserted can be transferred
(i.e. specialized transduction). After replication and
release of phage and infection of a recipient, lysogenization of
recipient can occur resulting in the stable transfer of donor
genes. The recipient will now have two copies of the gene(s) that
were transferred. Legitimate recombination between the donor and
recipient genes is also possible.
Significance - Lysogenic (phage) conversion occurs in nature
and is the source of virulent strains of bacteria.
of DNA from a donor to a recipient by direct physical contact between
the cells. In bacteria there are two mating types a donor (male) and a
recipient (female) and the direction of transfer of genetic material
is one way; DNA is transferred from a donor to a recipient.
types in bacteria
Donor - The ability of a bacterium to be a donor is a
consequence of the presence in the cell of an extra piece of DNA
called the F factor or fertility factor or sex
factor. The F factor is a circular piece of DNA that can replicate
autonomously in the cell; it is an independent replicon.
Extrachromosomal pieces of DNA that can replicate autonomously are
given the general name of plasmids. The F factor has genes on
it that are needed for its replication and for its ability to
transfer DNA to a recipient. One of the things the F factor codes
for is the ability to produce a sex pilus (F pilus) on the
surface of the bacterium. This pilus is important in the conjugation
process. The F factor is not the only plasmid that can mediated
conjugation but it is generally used as the model.
Recipient - The ability to act as a recipient is a
consequence of the lack of the F factor.
states of the F factor
Autonomous (F+) - In this state the F
factor carries only those genes necessary for its replication and
for DNA transfer. There are no chromosomal genes associated
with the F factor in F+ strains.
crosses of the type F+ X F- the F-
becomes F+ while F+ remains F+.
Thus, the F factor is infectious. In addition, there is only low
level transfer of chromosomal genes.
Integrated (Hfr) - In this state the F factor has
integrated into the bacterial chromosome via a recombination event
as illustrated in the Figure 5a
b Fig 5 . Physiological states of F
crosses of the type Hfr X F- the F- rarely
becomes Hfr and Hfr remains Hfr. In addition, there is a high
frequency of transfer of donor chromosomal genes.
Autonomous with chromosomal genes (F') - In this
state the F factor is autonomous but it now carries some
chromosomal genes. F' factors are produced by excision of the F
factor from an Hfr, as illustrated in Figure 5b. Occasionally,
when the F factor is excising from the Hfr chromosome, donor genes
on either side of the F factor can be excised with the F factor
generating an F'. F' factors are named depending on the
chromosomal genes that they carry.
crosses of the type F' X F- the F- becomes
F' while F' remains F'. In addition there is high frequency of
transfer of those chromosomal genes on the F' and low frequency
transfer of other donor chromosomal genes.
Fig. 6. Mechanism of F+ x F- crosses
F+ X F- crosses (Figure 6)
Fig. 7 Mechanism of Hfr x F- crosses
Pair formation - The tip of the sex pilus comes in contact with
the recipient and a conjugation bridge is formed between
the two cells. It is through this bridge that the DNA will pass
from the donor to the recipient. Thus, the DNA is protected from
environmental nucleases. The mating pairs can be separated by
shear forces and conjugation can be interrupted. Consequently,
the mating pairs remain associated for only a short time.
DNA transfer - The plasmid DNA is nicked at a specific site
called the origin of transfer and is replicated by a
rolling circle mechanism. A single strand of DNA passes through
the conjugation bridge and enters the recipient where the second
strand is replicated.
This process explains the characteristics of F+ X F-
crosses. The recipient becomes F+, the donor remains
F+ and there is low frequency of transfer of donor
chromosomal genes. Indeed, as depicted in Figure 7 there is no
transfer of donor chromosomal genes. In practice however, there
is a low level of transfer of donor chromosomal genes in such
Hfr X F- crosses (Figure 7)
DNA transfer - The DNA is nicked at the origin of transfer and
is replicated by a rolling circle mechanism. But the DNA that is
transferred first is the chromosome. Depending upon where in the
chromosome the F factor has integrated and in what orientation,
different chromosomal genes will be transferred at different
times. However, the relative order and distances of the genes
will always remain the same. Only when the entire
chromosome is transferred will the F factor be transferred.
Since shearing forces separate the mating pairs it is rare that
the entire chromosome will be transferred. Thus, the recipient
does not receive the F factor in a Hfr X F- cross.
Legitimate recombination - Recombination between the transferred
DNA and the chromosome results in the exchange of genetic
material between the donor and recipient.
This mechanism explains the characteristics of Hfr X F-
crosses. The recipient remains F-, the donor remains Hfr and
there is a high frequency of transfer of donor chromosomal
F' X F- crosses (Figure 8)
Fig. 8. The mechanism of F" x F- crosses
DNA transfer - This process is similar to F+ X F-
crosses. However, since the F' has some chromosomal genes on it
these will also be transferred.
Homologous recombination is not necessary although it may occur.
This mechanism explains the characteristics of F' X F-
crosses. The F- becomes F', the F' remains F' and the is high
frequency transfer of donor genes on the F' but low frequency
transfer of other donor chromosomal genes.
Significance - Among the Gram negative bacteria this is the
major way that bacterial genes are transferred. Transfer can occur
between different species of bacteria. Transfer of multiple
antibiotic resistance by conjugation has become a major problem in
the treatment of certain bacterial diseases. Since the recipient
cell becomes a donor after transfer of a plasmid it is easy to see
why an antibiotic resistance gene carried on a plasmid can quickly
convert a sensitive population of cells to a resistant one.
positive bacteria also have plasmids that carry multiple antibiotic
resistance genes, in some cases these plasmids are transferred by
conjugation while in others they are transferred by transduction.
The mechanism of conjugation in Gram + bacteria is different than
that for Gram -. In Gram + bacteria the donor makes an adhesive
material which causes aggregation with the recipient and the DNA is
TRANSPOSABLE GENETIC ELEMENTS
Transposable Genetic Elements
genetic elements are segments of DNA that have the capacity to move
from one location to another (i.e. jumping genes).
Properties of Transposable Genetic Elements
Random movement - Transposable genetic elements can move from
any DNA molecule to any DNA other molecule or even to another
location on the same molecule. The movement is not totally random;
there are preferred sites in a DNA molecule at which the
transposable genetic element will insert.
Not capable of self replication - The transposable genetic
elements do not exist autonomously (exception - some transposable
phages) and thus, to be replicated they must be a part of some other
Transposition mediated by site-specific recombination -
Transposition requires little or no homology between the current
location and the new site. The transposition event is mediated by a transposase
coded for by the transposable genetic element. Recombination that
does not require homology between the recombining molecules is
called site-specific or illegitimate or nonhomologous
Transposition can be accompanied by duplication - In many
instances transposition of the transposable genetic element results
in removal of the element from the original site and insertion at a
new site. However, in some cases the transposition event is
accompanied by the duplication of the transposable genetic element.
One copy remains at the original site and the other is transposed to
the new site.
Types of Transposable Genetic Elements
Insertion sequences (IS)- Insertion sequences are
transposable genetic elements that carry no known genes except those
that are required for transposition.
Nomenclature - Insertion sequences are given the
designation IS followed by a number. e.g. IS1
Fig. 9. Structure of transposable genetic elements
Structure (Figure 9)
sequences are small stretches of DNA that have at their ends
repeated sequences, which are involved in transposition. In
between the terminal repeated sequences there are genes involved
in transposition and sequences that can control the expression of
the genes but no other nonessential genes are present.
Mutation - The introduction of an insertion sequence into a
bacterial gene will result in the inactivation of the gene.
Plasmid insertion into chromosomes - The sites at which plasmids
insert into the bacterial chromosome are at or near insertion
sequence in the chromosome.
Phase Variation - The flagellar antigens are one of the main
antigens to which the immune response is directed in our attempt
to fight off a bacterial infection. In Salmonella there are two
genes which code for two antigenically different flagellar
antigens. The expression of these genes is regulated by an
insertion sequences. In one orientation one of the genes is
active while in the other orientation the other flagellar gene
is active. Thus, Salmonella can change their flagella in
response to the immune systems' attack. Phase variation is not
unique to Salmonella flagellar antigens. It is also seen with
other bacterial surface antigens. Also the mechanism of phase
variation may differ in different species of bacteria (e.g.
Transposons (Tn) - Transposons are transposable genetic elements
that carry one or more other genes in addition to those which are
essential for transposition.
Nomenclature - Transposons are given the designation
Tn followed by a number.
Structure - The structure of a transposon is similar
to that of an insertion sequence. The extra genes are located
between the terminal repeated sequences. In some instances
(composite transposons) the terminal repeated sequences are
actually insertion sequences. (See Figure 10).
Fig. 10. Transposon structure
Importance - Many antibiotic resistance genes are
located on transposons. Since transposons can jump from one DNA
molecule to another, these antibiotic resistance transposons are a
major factor in the development of plasmids which can confer
multiple drug resistance on a bacterium harboring such a plasmid.
These multiple drug resistance plasmids have become a major
medical problem because the indiscriminate use of antibiotics have
provided a selective advantage for bacteria harboring these
Definition - Plasmids are extrachromosomal genetic elements
capable of autonomous replication. An episome is a
plasmid that can integrate into the bacterial chromosome.
Conjugative plasmids - Conjugative plasmids are those
that mediated conjugation. These plasmids are usually large and have
all the genes necessary for autonomous replication and for transfer
of DNA to a recipient (e.g. genes for sex pilus).
Nonconjugative plasmids - Nonconjugative plasmids are
those that cannot mediate conjugation. They are usually smaller than
conjugative plasmids and they lack one or more of the genes needed
for transfer of DNA. A nonconjugative plasmid can be transferred by
conjugation if the cell also harbors a conjugative plasmid.
plasmid (F factor)
Fig. 11. R plasmid structure
Bacteriocinogenic plasmids - These plasmids have
genes which code for substances that kill other bacteria. These
substances are called bacteriocins or colicins.
Resistance plasmids 7 factors) - These
plasmids carry antibiotic resistance genes.
Origin - The origin of the R factors is not known. It is likely
that they evolved for other purposes and the advent of the
antibiotic age provided a selective advantage for their
Structure - R plasmids are conjugative plasmids in which the
genes for replication and transfer are located on one part of
the R factor and the resistance genes are located on another
part as illustrated in Figure 11.
(Resistance Transfer Factor) - carries the transfer genes.
determinant - carries the resistance genes. The resistance genes
are often parts of transposons.
of action of resistance genes
Modification (detoxification) of antibiotic - e.g.
Alteration of target site - e.g. Streptomycin
Alteration of uptake - Tetracycline resistance
Replacement of sensitive pathway - e.g. new folic acid
pathway for resistance to sulfa drugs