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A process by which a chromosome is formed from the DNA derived from two different parental cells. Three major processes lead to the formation of recombinant chromosomes in bacteria:
1. transformation,
2. transduction and
3. conjugation.
These processes differ from eucaryotic cells in that a true fusion cell (zygote) is not formed; instead, part of the genetic material of the donor (D) cell, called an exogenote, is transferred to a recipient (R) cell, called an endogenote. The partial zygote is called a merozygote.
The host cell is protected from foreign DNA by restriction and modification (R-M) enzyme systems. The R-M enzyme system is composed of:
a. endonuclease (restriction enzyme) - functions to degrade DNA.b. methylase (modification enzyme) - functions to protect DNA by causing
the attachment of methyl groups at specific sites on DNA. Apparently
these are the same sites that are recognized and attacked by the
endonuclease. DNA modified by the methylase of one R-M system is
protected from the endonuclease of that system, but is still
susceptible to a restriction enzyme with a different specificity.
a. breakage of the parental (D and R) molecules.b. conservation of base-pair sequence. Involves the complementary
pairing of bases, in a proper alignment of parental molecules,
between single-stranded regions of the parental double helixes.
c. reunion of DNA
8.4.1Transformation
Hotchkiss defined transformation as a modification of a hereditary property of one bacterial strain through the action of DNA from a related but different organism.
a. All mutant loci (markers) of the R cell are capable of beingtransformed; however, rarely more than one marker is transformed.
This is due to an artifact resulting from technical procedures used
in preparing donor DNA.
b. The competent state of the R cell is important in the process. The competent state is a physiological state that fluctuates during the cell cycle and is related to the ability of the R cell to absorb DNA and express a phenotypic characteristic.
c. Double stranded DNA is required. In the penetration process, one strand of donor DNA is degraded. Fragments with mol wts of less than 5 X 105 may not be taken up.
d. The competent state is suggested to be part of an active transport system since the transformation process can be inhibited by metabolic and protein inhibitors such as DNP, azide, and chloramphenicol.
e. Examples of bacteria participating in transformation: Hemophilus, Diplococcus, Bacillus and certain species of Streptococcus.
f. Phenotypic characters transformed: capsular antigens, antibiotic and drug resistant factors, inducible enzyme synthesis and colonial morphology.
8.4.1.1 Kind of Transformation
1. Autogenic Transformation: The R cell is the transformed cell exhibiting the phenotypic character from the D cell from which DNA had been taken. This is a result of segregation of alleles. Alleles are two or more forms of a gene controlling the same characteristic and occupying the same genetical location.
2. Allogenic transformation: the R cell exhibits a phenotypic marker not present in the D or R cell. This phenomenon is a result of supplementation and complimentation of alleles.
Example:
R Cell (capsule; type II) treated with DNA from D cell
(capsule; type IV). Results: capsule; type VIII.
Explanation: The R cell contained a part of the metabolic pathway for capsule VIII. The D cell contained the part that was missing.
Practical Applications:
The employment of interspecific transformation to establish a species concept among bacteria. Genetic studies on streptococci and the pneumococci suggest that these two entities could be considered a variety rather that a distinct species in that genetic crossing has been observed.
The following is an example of an application of transformation in the establishment of genetic relationships among bacteria:
Tube A Tube B Tube C Tube D H. influenzae H. aegyptius H. parainfluenzae H. suis
H. influenzae H. influenzae H. influenzae H. influenzae
A single strand of DNA from Hemophilus influenzae is labeled with H3T. Single strands of DNA from H. aegyptius, H. parainfluenzae and H. suis are labeled with 32PP. Labeled H. influenzae is mixed with the other labeled species and subjected to density gradient centrifugation. The location of the labels determines the degree of hybridization. Tube A is the control. See diagram. Results indicate that H. suis is more distantly related along the phylogenetic scale to H. influenzae as indicated by a low or no degree of hybridization.
8.4.2Transduction:
A process in which a fragment of host genome, other than the genetic material specific for the transmitting phage per se, is carried by phage particles from one bacterial cell to another.
8.4.2.1 Language
1. Prophage: phage-specific genetic material.2. Lysogenic bacterium: Processes and transmits through its progeny the ability to produce phage.
3. Virulent phage: produces a lytic infection
4. Temperate phage: May produce lysogenic strains of bacteria as well as produce lytic infections.
5. gal+ - beta galactosidase gene
6. bio+ - biotin gene
8.4.2.2 Transductive Process
Phage infection of donor bacterium incorporation of genetic fragments into the transducing phage Lysis of donor bacterial cell injection of genetic material into recipient cell synapsis, cross-over and replication integration of foreign DNA into a new recombinant chromosome and segregation into a progeny cell.
8.4.2.3 Forms of Transduction:
Specialized Transduction:
E. coli lysogenic for phage lambda will carry the prophage inserted into the chromosome at a site between the
gal+ and bio+ loci. induction the prophage detaches infrequently, pairing occurs between the 2 different sites and the recombinant event generates a circle of DNA consisting of a major fraction of the phage and a segment of chromosomal DNA from either side but not both of the attachment sites.
If the excise DNA includes the gal+ gene.
The phage is called: dg (Lambda bio+ gene defective).
The total amount of DNA packaged in a phage particle is fixed. If
gal+ or bio+ are included, a corresponding amount of lambda DNA must be missing. This explains why transducing phages are always defective for some phage functions. The induction of lambda-lysogenic gal+ culture produces a lysate containing a small number of lambda dg phages. Infect nonlysogenic gal bacteria. Lambda dg DNA becomes integrated into the chromosome. Each bacterial cell retains its gal locus and thus becomes gal+/gal- partial diploid.
Generalized Transduction:
The transducing phage contains only a small amount of phage DNA. Thus a lysate will produce two types of particles: a majority type that contains only phage DNA and a minority type that contains mainly host DNA
8.4.2.4 Phage Conversion
Certain properties of bacterial cells are controlled by phagenes. These properties are manifested only by lysogenic strains.
Example:
Corynebacterium diphtheria produces an exotoxin in the lysogenic state.
