Transposon Pools


It is often useful to isolate a collection of transposon insertions at many different sites in the genome of a bacterium. A population of cells that each contains transposon insertions at random positions in the genome (a transposon "pool") can be used to select or screen for those cells with insertions in or near a particular gene. For a genome the size of E. coli or Salmonella with about 3000 nonessential genes, a transposon pool made up of approximately 15,000 random insertions will represent an insertion in most nonessential genes. About 1 in 3000 of these cells will contain a transposon insertion in any particular, nonessential gene, and about 1 in 100 of these cells will contain a transposon insertion within 1 centisome (i.e. 1 min or 1% of the chromosome length) of any particular gene.

A transposon pool can be made by any transposon delivery system. The following figure shows how a Tn10 pool could be constructed in Salmonella.

To isolate insertions in or near a gene, cells with transposon insertions are first selected by plating onto rich medium containing an antibiotic resistance expressed by the transposon. The resulting colonies can then be screened for insertion mutations in a gene or insertions near a gene.

An example of how to isolate insertions in a gene is shown below.

An example of how to isolate insertions near a gene is shown below.


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Please send comments, suggestions, or questions to smaloy@sciences.sdsu.edu
Last modified October 14, 2003