Microbial Genetics (Micr340) Lecture 12 Transposition

Nonhomologous recombination No requirement for two DNAs being of the same or similar nucleotide sequences Needs enzymes that recognize specific regions in DNA Mechanisms include: transposition phage integration and excision resolution of cointegrates

Transposition Transposons – DNA elements that can hop (transpose) from one place in DNA to another Movement by a transposon is called transposition, catalyzed by enzymes called transposases Transposons usually encode their own transposases Transposons are known to exist in all organisms on earth

History of Transposons Transposons (jumping genes) were first discovered in Maize (corn) by Barbara McClintock. Transposons were later found in bacteria, flies and humans. This discovery led to the awarding of Nobel Prize to McClintock in 1983.

Transposition Many transposons are essentially cut out of one DNA and inserted into another Other transposons are copied and then inserted elsewhere Donor DNA and target DNA

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Structure of Bacterial Transposons All contain repeats at their ends, usually inverted repeats (IR) Presence of short direct repeats in the target DNA that bracket the transposon The sites of insertion are different among target DNAs

Types of Bacterial Transposons Smallest bacterial transposons are called insertion sequence elements (IS elements); they only encode transposase enzymes Composite transposons – formed by two IS elements of the same type, bracketing other genes Noncomposite transposons are bracketed by two short inverted repeats, with multiple resistance markers.

Structure of a Bacterial transposon

IS Elements and Composite Transposons

Composite transposons

Noncomposite transposons

Assay of transposition

Transposon Mutagenesis A gene that has been marked with a transposon is relatively easy to map Genes marked by transposons are very easy to be cloned Transposon mutagenesis to identify essential genes (related to research of my lab).

Properties of transposons for mutagenesis Transpose at a fairly high frequency Not very selective in target sequence Carry selectable marker (antibiotic resistance genes) Broad host range for transposition

Tn5 mutagenesis

Cloning genes mutated by transposition

Isolating random gene fusions