Team:Imperial College London/M3/DamMethylation

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=[[Image:II09_Thumb_m3.png|40px]]<font size='5'><b>Module 3: Genome Deletion Overview</b></font>=
==Dam methylation==
==Dam methylation==
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[[Image:II09_meth_RE_balance.jpg| left| 300px]]
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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. <br>
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<br>
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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.<br>
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[[Image:II09_Dam action2.jpg| right|300px]]
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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. <br>
 +
<br>
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Dam methylases recognise the sequence GATC and transfer a methyl group to 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. <br>
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To protect against DNA destruction due to basal levels of restriction enzyme production, we have made use of the native E. coli [[Team:Imperial_College_London/M1/Dam| Dam methylase protection system]]. This methylates DNA.  Therefore, only high levels of restriction enzyme (ie. after thermal triggering) will cleave the DNA.<br>
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<html><a href="https://2009.igem.org/Team:Imperial_College_London/Dam"><img style="vertical-align:bottom;" width=90px align="left" src="http://i691.photobucket.com/albums/vv271/dk806/II09_Learnmore.png"></a></html><br><br>&nbsp; about the Dam methylation Defense Mechanism
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==References==
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[1] Alfred M. Pingoud, 2004, Restriction Endonucleases (Nucleic Acids and Molecular Biology), Springer pp10-40
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[2] Luke, McCallum and Halford, 1987, The EcoRV restriction endonuclease|Gene Amplification and Analysis pp183-205
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===Module 3 - Genome Deletion===
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<center><b>Module 3 - Genome Deletion </b></center>  
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<html><center><a href="https://2009.igem.org/Team:Imperial_College_London/M3"><img style="vertical-align:bottom;" width="20%" src="http://i691.photobucket.com/albums/vv271/dk806/II09_Homepageimage3.png"></a><a href="https://2009.igem.org/Team:Imperial_College_London/M3/RestrictionEnzymes"><img style="vertical-align:bottom;" width="20%" src="http://i691.photobucket.com/albums/vv271/dk806/II09_Drylabmainimage5.png"></a><a
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<td width="25%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/M3/Genetic"><b>Genetic Circuit</b></a></center></td>
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<td width="25%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/Temporal_Control/M3/Wetlab"><b>WetLab</b></a></center></td>
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<td width="20%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/M3"><b>Module 3 Overview</b></a></center></td>
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<td width="25%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/M3/Modelling"><b>Modelling</b></a></center></td>
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<td width="25%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/M3/Results"><b>Results</b></a></center></td>
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<td width="20%"><center><a href="/Team:Imperial_College_London/M3/RestrictionEnzymes"><b>Restriction Enzymes</b></a></center></td>
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<td width="20%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/M3/Genetic"><b>Genetic Circuit</b></a></center></td>
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<td width="20%"><center><a href="https://2009.igem.org/Team:Imperial_College_London/Temporal_Control/M3/Wetlab"><b>Wet Lab</b></a></center></td>
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<td width="20%"><center><a  
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href="https://2009.igem.org/Team:Imperial_College_London/M3/Modelling"><b>Modelling</b></a></center></td>
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Latest revision as of 01:53, 22 October 2009

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.

Dam methylases recognise the sequence GATC and transfer a methyl group to 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

[1] Alfred M. Pingoud, 2004, Restriction Endonucleases (Nucleic Acids and Molecular Biology), Springer pp10-40

[2] Luke, McCallum and Halford, 1987, The EcoRV restriction endonuclease|Gene Amplification and Analysis pp183-205



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