Describe a procedure that can be used to restrict or digest the TOL plasmid

Expert Solution

Hi!

First of all you need to have isolated the TOL plasmid from a Pseudomonas strain.

Here you can find a protocol to isolate TOL plasmids:

Isolation and characterization of spontaneously occurring TOL plasmid mutants of Pseudomonas putida HS1. D A Kunz, P J Chapman. Journal of Bacteriology Jun 1981, 146 (3) 952-964; DOI:

Then, you will have to know the sequence of the plasmid. For example this is the plasmid scheme of pKD1 TOL plasmid.

Reference: Yano H, Miyakoshi M, Ohshima K, et al. Complete nucleotide sequence of TOL plasmid pDK1 provides evidence for evolutionary history of IncP-7 catabolic plasmids [published correction appears in J Bacteriol. 2010 Oct;192(20):5558]. J Bacteriol. 2010;192(17):4337‐4347. doi:10.1128/JB.00359-10.

Genebank accesion number: AB434906.1

In blue are depicted the restriction enzymes that has 1 to 5 cutting sites in this plasmid.

So basically once you isolated the plasmid, you choose one of the enzymes and the perform the restriction reaction according to manufacturers instructions.

gladiator answered 2 years ago

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