The ability to exchange genes within a population is a nearly universal
attribute of living things. Among prokaryotes, there is no known case that
genetic exchange is an obligatory step (as it often is among eukaryotes) in the
completion of an organism's life cycle. Rather, genetic exchange seems to be an
occasional process that occurs by three quite different mechanisms in various
prokaryotes. The three mechanisms of genetic exchange are transduction,
conjugation, and transformation.
In transduction, DNA is transferred from one prokaryotic cell to another as a
consequence of a rare formation of an aberrant phage viron in which some or all
of its normal complement of DNA is replaced by bacterial DNA (donor DNA). When
such a phage viron attaches to and introduces this DNA into another bacterial
cell (the recipient), genetic exchange is effected. With conjugation, genetic
exchange occurs between cells in direct contact with one another by a process
that is, in all known cases, encoded by plasmid-borne genes. Usually only the
plasmid itself is transferred from donor to recipient, but sometimes
chromosomal genes are transferred as well. In transformation, DNA is released
from cells into the surrounding medium, and recipient cells incorporate it into themselves from
this solution.
Transformation was the first mechanism of bacterial genetic exchange to be
discovered. In 1928, a classic experiment demonstrated that injection of mice
with an avirulent (not capable of causing disease) strain of Streptococcus
pneumoniae, together with heat-killed cells of a virulent (disease-causing)
strain-killed mice, while injection of either of these strains separately did not. These and subsequent experiments established that the
surviving cells were recombinant: they exhibited certain properties (including
virulence) that were typical of the killed cells and others that were typical
of the avirulent culture. Thus, a genetic exchange of the DNA dissolved in the
external medium had occurred between the dead cells and the live ones. At the
time, it was thought that a particular substance, a transforming principle,
caused the exchange to take place. Thus the word transformation came to be used to describe genetic exchange among
prokaryotes.
Cells that are in a state in which they can be transformed by DNA in their
environment are said to be competent. In a significant number of bacteria,
entry into the competent state is encoded by chromosomal genes and signaled by
certain environmental conditions. Such bacteria are said to be capable of
undergoing natural transformation. Many other bacteria do not become competent
under ordinary conditions but can be made competent by exposing them to a
variety of highly artificial treatments such as exposure to high
concentrations of divalent cations.
In this experiment, you will artificially transform E. coli cells. E. coli
cells do not posses a natural system for transformation. They are made
competent only following subjection of log-phase cells in culture to calcium chloride solution. Such remolded,
competent cells are now receptive to insertion of foreign DNA (contained in a
plasmid). The students will conduct a laboratory procedure in which they
transform wild-type E. coli by inserting a plasmid that contains the gene for
tetracycline resistance.
Teacher's Note: In another experiment ("Microbial Techniques in Recombinant DNA Technology" [88W8234]), students observed two
different strains of E. coli wild-type and a strain which contained plasmid
pBR322 which confers antibiotic resistance. The recommended time line for this
experiment covers a period of one week.
AE Partners Collection Index
