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  Section: Genetics » Sexuality and Recombination in Bacteria and Viruses
 
 
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Conjugation mapping through interrupted mapping

 
     
 
Content
Sexuality and Recombination in Bacteria and Viruses
Three Methods for Transfer of Genetic Material 
Sexual conjugation in bacteria 
Culture media and mutant strains
Discovery of gene transfer
Discovery of linkage in bacteria
Donor and recipient strains
- Mechanism of chromosome transfer
- Physical structures involved in chromosome transfer
Linkage maps in bacteria
Conjugation mapping through interrupted mapping
Circular linkage map
Linkage information from transformation
Recombination after gene transfer
High resolution mapping
Linear order of genes
Replication and recombination in viruses 
Replication of bacteriophages
Lysogenic bacteria
Transduction
Recombination in viruses
Circular genetic maps in viruses

Linkage Maps in Bacteria
Conjugation mapping through interrupted mapping
In conjugation experiments, it was observed that the number of genes transmitted from donor to recipient was directly proportional to the time interval for which conjugation was allowed. Based on this feature, Francois Jacob and Ellie Wollman developed a technique called 'interrupted mating technique' for mapping the bacterial chromosome. In this technique donor Hfr and recipient F" strains are mixed and allowed to conjugate for a short period of time. Then samples are removed at periodic intervals and subjected to violent agitation to break the conjugation tube. The length of donor chromosome transmitted could then be determined and mapped in terms of time units required for transfer. It is known that 8 minutes are needed for conjugation to begin and then chromo'some is transferred slowly in terms of time units (Fig. 12.13), one time unit being equal to one minute. The complete chromosome of E. coli is transferred in about 89 minutes (Fig. 12.14) and therefore the bacterial chromosome is 89 time units in length. Genes which are 2-3 time units apart can be precisely mapped by this method. Further mapping within the limits of 1-3 minutes is done by conventional recombination methods.

The transfer of genes in linear order from male bacterium (tagged with virus) to female; the length of transferred segment is dependent upon the time taken for transfer and the chromosome is, therefore, mapped in terms of time units.
Fig. 12.13. The transfer of genes in linear order from male bacterium (tagged with virus) to female; the length of transferred segment is dependent upon the time taken for transfer and the chromosome is, therefore, mapped in terms of time units.
 
Circular linkage map of Escherichia coli chromosome, the units of recombination shown as minutes required for transfer.
Fig. 12.14. Circular linkage map of Escherichia coli chromosome, the units of recombination shown as minutes required for transfer.

Circular linkage map
When linkage maps in bacteria were prepared using several Hfr strains, these maps differed with respect to the first and the last genes of the map. However, once we know the first gene transferred and the direction of transfer of subsequent genes, the order is predetermined. The only way these results could be explained, was through the assumption of a circular map, which cleaves at any point due to attachment of F factor to form a linear chromosome. This linear chromosome enters the recipient cell from the end away from the site of attachment of F factor. Experimental evidence was also available to prove that Hfr really results due to the insertion of F into the chromosome. Circular nature of linkage group was later confirmed by evidence for physical circular nature of bacterial chromosome.

Linkage information from transformation
Transformation is achieved through the uptake of naked DNA extracted from one strain of bacteria by another strain of bacteria. While extracting this DNA, some breakage into small pieces does take place. If two genes are closer to each other, they have better chance of being carried on the same piece of DNA, thus causing double transformation. On the other hand genes widely separated, will have a better chance of being carried on separate DNA segments and the frequency of double transformation will be the product of single transformation frequencies. Therefore a departure from product rule of probability for the observed frequency of double recombinants will prove close linkage. Further high resolution mapping will be done by a method given in a subsequent section.

Recombination after gene transfer
When a chromosome segment is transferred from a donor to a recipient strain (or through transduction or transformation), this transferred segment must be integrated into host genome by an exchange mechanism to produce a stable recombinant. The recipient cell at this stage is called a merozygote (partial diploid) which has the complete genome of F-, called endogenote and an incomplete genome called exogenote derived from F+ or Hfr. An even number of crossovers, rather than a single or odd number of crossovers, allows incorporation of a part of the genome from exogenote into the endogenote; one of the two products of exchange (i.e. a linear fragment) is usually lost (Fig. 12.15).
 
Recombination in bacteria after gene transfer (single and double crossovers give linear and ring chromosomes respectively).
Fig. 12.15. Recombination in bacteria after gene transfer (single and double crossovers give linear and ring chromosomes respectively).

 
     
 
 
     




     
 
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