Lecture #27 - 11/6/2007 1 Lecture 27 Lecture 27 Transposons...

Info iconThis preview shows pages 1–4. Sign up to view the full content.

View Full Document Right Arrow Icon

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
This is the end of the preview. Sign up to access the rest of the document.

Unformatted text preview: 11/6/2007 1 Lecture 27 Lecture 27 Transposons and Insertion Sequences Transposons and Insertion Sequences Recombinant DNA technologies Using recombinant DNA to create transgenic plants Insertion sequences and transposons IS elements and transposons are fragments of DNA that reside within chromosome or plasmid. They are able to identify a target DNA site on a chromosome, phage, or plasmid, and move from their original site to this new site. Sometimes called Genetic parasites, genetic weeds, selfish DNA. Some copy themselves in the process, so that a copy remains at the old site. IS elements: 500 1000 bases long. Contain just 1 gene encoding transposase, which causes transposition (e.g. IS2, see Fig. 9.29a). Found on the chromosomes of most or all bacteria. Ends have inverted repeats. Transposons 5000 10,000 bases long. Encode a transposase and 1 5 additional genes. Composite transposons (e.g. Tn5, Fig 9 29b) have IS sequences at their ends Fig 9.29b) have IS sequences at their ends. Tn5 has IS50 at either end, in inverted orientation. Tn5 also has 3 antibiotic resistance genes. Tn5 may have evolved by two copies of IS50 that flanked these genes. Implies that new composite transposons can evolve easily. Fig 10.28 11/6/2007 2 Fig 10.32 If transposon inserts within a gene, that gene will be disrupted (see Fig. 9.32). Also, if that gene lies within an operon, the transposon will block transcription of the downstream part of the operon. Recombinant DNA Technologies . Steps in Molecular Cloning 1) Isolate and fragment DNA to be studied. 2) Join DNA fragments to a cloning vector. 3) Introduce cloning vector with insert into a host organism. 4) Detect the desired clone against a background of other clones. 1. Isolation of fragmentation of DNA. DNA is easy to isolate, though fragmentation of very long fragments is a problem. Fragmentation is done using restriction endonucleases, enzymes that cut DNA at specific sites. Example: Bam HI cuts the sequence GGATCC. This sequence is called a palindrome. Bam HI enzyme binds as a dimer, each monomer cutting one strand V .................NNNNGGATCCNNNN.................. .................NNNNCCTAGGNNNN.................. ^ .................NNNNG 3' 5' GATCCNNNN................ .................NNNNCCTAG 5' 3' GNNNN................ The resulting ends have identical overhangs and are called sticky, important later. How frequent are BamHI sites? Answer: 1/4 x 1/4 x 1/4 x 1/4 x 1/4 x 1/4 = 1/4096; A Bam HI site appears on average once every 4096 nucleotides, or approximately 1000 times per bacterial chromosome. 11/6/2007 3 Many restriction enzymes have been isolated from bacteria. Bacteria protect themselves by modification systems, which modify the same site by methylation of a DNA base, making cutting impossible. The bacterium modifies itscutting impossible....
View Full Document

Page1 / 11

Lecture #27 - 11/6/2007 1 Lecture 27 Lecture 27 Transposons...

This preview shows document pages 1 - 4. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online