Figure 1. Bacterial transformation
Until the discovery of #TRANSFORMATION in the early 1940's it was assumed that bacteria lacked any means of exchanging genetic material between them. Within a few years two additional means of genetic exchange by bacteria, #CONJUGATION, or MATING, and #TRANSDUCTION, were discovered. All three of these methods result in the transfer of DNA from a DONOR bacterium to a RECIPIENT bacterium. The donor DNA is subsequently INTEGRATED by #RECOMBINATIONAL EXCHANGE into the genome of the recipient and becomes a functioning part of the recipient's DNA. If the donor DNA contains genes that differ in activity (ALLELIC genes) from those in the recipient, then the recipient assumes a different #PHENOTYPE; that of the donor.
FAQ is "Do these processes of DNA exchange occur naturally in nature?"
Answer: A clear "YES", however the frequency of these various exchanges are still being hotly discussed by microbiologists. What-ever-the-case in nature, in the research laboratory these techniques are employed EXTENSIVELY in genetic engineering experiments; in fact the genetic engineering revolution would not be possible without them. The easiest and, hence most commonly used means of transferring DNA between bacteria is TRANSFORMATION.
Although you have learned the details of these three processes of DNA exchange from the lectures, the following is a brief review of each:
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TRANSFORMATION: This process involves the ISOLATION of DNA from the donor strain. The CELL-FREE DNA is then mixed with #COMPETENT recipient cells which take up the donor DNA into their cytoplasm. The donor DNA then replaces some portion (genes) of the recipient's DNA by RECOMBINATION EXCHANGE.
 Figure 2. Bacterial conjugation. |
CONJUGATION: The donor (male) cell contains a #SEX PLASMID (FERTILITY PLASMID or F PLASMID) that contains genes which control the synthesis of #SEX PILI and of enzymes that induce DNA exchange. The F+ donor cell's sex pili attaches to a F- (female) cell and a CONJUGATION BRIDGE joining the two cytoplasms is formed. A series of events MOBILIZES the donor's sex plasmid so that a copy of it's DNA is transferred into the recipient cell (one way only). The donor DNA is converted to a double stranded form. The plasmid's genes are then EXPRESSED in the donor cell. Therefore, if the plasmid carries one or more dominant bacterial genes from the donor that are DIFFERENT (alleles) from those of the recipient, these donor-genes will give the recipient the PHENOTYPE of the donor. For example, if a recipient that is UNABLE to utilize the sugar lactose (LACTOSE MINUS) receives a plasmid carrying donor LACTOSE UTILIZING genes, the recipient will now be capable of using lactose (LACTOSE +). Furthermore, the donor genes may REPLACE the recipient's-genes by recombination exchange.
Figure 3. Bacterial transduction.
TRANSDUCTION: The donor cell is infected with a #BACTERIOPHAGE. During the replication process of the phage, FRAGMENTS of the shattered host's DNA are accidentally incorporated into the new phage protein coat producing a DEFECTIVE PHAGE. The rare defective phage can however infect another bacterial host, the recipient, and inject the donor's DNA into the recipient cell. Once in the recipient the donor DNA can then replace some portion of the recipient's DNA by recombination exchange. For a discussion on the nature of bacteriophage visit this site.
In each of these exchanges the exchange can only be detected if the recipient's genetic phenotype is CHANGED in some detectable way by the integration of a donor's gene that is an ALLELE of a recipient's gene.
Q. Of what possible importance is "bacterial genetic exchange or sex" in nature?
A. It is a way bacteria have of testing NEW COMBINATIONS of genes to see if a new combination will aid them in survival. It is also a ways of maintaining a large number of alleles within a population so that not every bacteria has to carry a huge load of extra genes (remember that bacteria are HAPLOID). We will see in this experiment how useful conjugation is in transferring an ability (GENE) lacking in one bacterium from another bacterium that has that ability (GENE). Conjugation is a major means of spreading resistant genes to sensitive bacterial populations.
Q. Why aren't all bacteria F+?
A. Plasmids represent an "extra load" of stuff a bacterium has to carry. When plasmids are not needed (when the genes they carry offer no survival advantage) they are frequently "dumped" . Consider what you carry in a back pack in the mountains; only that which is absolutely necessary, RIGHT?
Q. Why do microbiologists make such a big thing of DNA exchange by bacteria?
A. These are crucial research tools that allow scientists to isolate genes and to study their function. The discovery of these processes provided crucial techniques and information for studying molecular biology and developing genetic engineering.
- MacConkey agar is a DIFFERENTIAL MEDIUM. Cells which use lactose appear red on this medium, while cells that do not use lactose appear light pink or white. See pg. 12 in your ATLAS.
- The donor F+ cells are Str SENSITIVE, whereas the recipient F- cells are Str RESISTANT.
- The donor male contains a plasmid that carries the genes for utilizing LACTOSE +.
- The recipient female has lost the ability to use lactose (LACTOSE-).
SAMPLE QUESTIONS: You should be able to answer
these questions at the conclusion of this laboratory. Copyright © Dr. R. E. Hurlbert, 1999.
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