Team:Imperial College London/M3/DamMethylation

From 2009.igem.org

Revision as of 23:31, 11 October 2009 by Kun (Talk | contribs)

Contents

II09 Thumb m3.pngModule 3: Genome Deletion Overview

Dam methylation

II09 meth RE balance.jpg

Restriction enzymes often come together with methylation enzymes to form a restriction-modification system. This well-known combination prevents the genome of the cell from being cleaved by its own restriction enzymes.

There is a strong asymmetry between the function of restriction enzymes and methylases. Restriction enzymes can cause just one cleavage, which if unrepaired, kills the cell. However, to effectively protect the cell, methylases need to methylate all the recognition sites. There is a fine balance that exists naturally between rstriction enzymes and methylases which can be easily disrupted. This is why endogenous promoters are often preferred.

II09 Dam action2.jpg
Methylation as a protection device against resriction enzymes is well documented, and has been proven to work. In our system, to protect against DNA destruction due to basal levels of restriction enzyme production, we have made use of the native E. coli Dam methylase protection system. The Dam system is chosen as both DpnII and TaqI enzyme activity can be blocked by Dam methylation.
II09 Dam action2.jpg


Dam methylases recognise the sequence GATC and methylate the Adenine base. This prevents the restriction enzymes from recognising the sequence and cleaving it. Therefore, only high levels of restriction enzyme (ie. after thermal triggering) will cleave the DNA.


References

Template:FormatRef
Template:FormatRef


Mr. Gene   Geneart   Clontech   Giant Microbes