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Generalised Transduction

	Generalised Transduction Phage mediated transduction of a small segment from any region of the bacterial chromosome is called generalized genetic transduction. In 1952 Zinder and Lederberg discovered that bacteriophage particles could transfer bacterial genes from one bacterium to another. Earlier Lederberg and Tatum (1946) bad performed the now classical cross of two auxotrophic mutants of E. coli KI2 (See the section on Conjugation in this chapter)
Zinder and Lederberg initially began their experiments with the objective of discovering whether the E. coli type of genetic exchange also existed in Salmonella typhimurium. Various strains of auxotrophs were crossed on an amino acid free minimal medium in an attempt to find new prototrophic combinations. The LA22 strain was unable t"9 synthesize the amino acids phenylalanine and tryptophan (phe- trp-strain) but could synthesize methionine and histidine . The LA2 strain was unable to synthesize methionine and histidine( mec- his- strain) but could synthesize phmylalanine and tryptophan. Crossing of the LA22 and LA2 strains resulted in a wild--type prototroph which could synthesize all four amino acids (phe + trp + met + his + ). Each strain, was placed in one arm of a Davis V-tube
The two arms of the tube were separated by a sintered glass filter which was impervious to bacterial calls (and thus prevented conjugation), but allowed the free passage of the nutrient medium and parlic1es smaller than (J.1 fl. The culture medium was made to pass through the filter from one arm to the other by alternating suction and pressure. Thus the two auxotrophic strains, although physically separate, were grown in the same medium
Prototrophs appeared in the LA22 arm of the U-tube but not in the LA2 arm. A genetically active filterable agel1t(FA) was produced by LA2 that resulted in prototroph formation in LA22. When LA2 was grown separately from LA22 it did not produce an active FA. That the FA was not an ordinary transforming agent (DNA molecule) was shown by the facts that it was resistant to DNase treatment and was larger than DNA. The FA was soon shown to be the temperate Salmonella phage P22, which was carried in one of the parent strains as a prophage. This was confirmed when it was shown that the FA is destroyed by exposure to P22 anti serum
	P22 ordinarily exists in the lysogenic state in the LA22 strain of Salmonella. Occasionally it passes into the proliferative vegetative state and lyses the bacterial host cells. The phage particles now pass through the filter and infect the non lysogenic LA2 strain. They become associated with genetic material from LA2. Some of this material was wild-type for the mutant recessive genes carried by LA22. Recombination between LA2 material and the LA22 chromosome produces prototrophs. The phage particles have brought about transduction of the LA22 strain. The nonlysogenic LA2 does not carry a temperate phage, and hence can transfer material only in the presence of the lysogenic LA22. The transducting frequency is low, and only one in 105-107 cells undergo transduction.

Only a small segment of the bacterial chromosome material undergoes transduction by the phage particle. This is shown by the fact that a cross between a strain synthesizing tryptophan, galactose and xylulose and a lysogenic strain unable to synthesize these three compounds produces a prototroph that can synthesize only one of the compounds.

Two other temperate phages (PI and 363) were later discovered that could transfer markers from one E. coli strain to another. Here also transductions were limited to short sections of the bacterial chromosome. A cross between a donor wild type for threonine, leucine and sodium azide (thr+ leu+ azi+) and a recipient auxotroph (thr leu- azi-) mostly produced threonine prototrophs (thr + leu- azr). A small percentage were also prototrophic for leucine (thr+ leu+ azi-). No thr +leu+ azi+ prototrophs Were produced. It can, therefore, be concluded that the donor chromosome fragment containing the thr locus sometimes extended to the leu locus, but never extended from the thr to the azi locus. It will thus be seen that usually only a single marker locus is transduced, indicating that only a short segment of the bacterial DNA is involved.

It is only occasionally that a single phage will transduce several gene loci (multiple transduction). Linkage maps for short sequences of genes can be constructed by using multiple transductions. Generalized transduction resembles transformation in the respect that usually only a single marker locus is transduced. The difference between the two processes is that in transduction the DNA reaches the cell surrounded by the phage coat, while in transformation the DNA is naked. Transduction, therefore, has an advantage in mapping because it protects the DNA from the environment, and thus makes it more readily reproducible.

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