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For this we used the HRP-mimicking DNAzyme which was described in 1998.<x-ref>Travascio1998</x-ref> It folds into a G-quadruplex that forms a complex with hemin and is then able to mimic the activity of a horse raddish peroxidase. Hence it is capable to produce reactive oxygen species (ROS) by the reduction of hydrogen peroxide to water. These ROS then leads to an activation of luminol which then becomes chemiluminescent. In several of our subprojects we exploit this function as an easy readout such as for the AptaBody.
 
For this we used the HRP-mimicking DNAzyme which was described in 1998.<x-ref>Travascio1998</x-ref> It folds into a G-quadruplex that forms a complex with hemin and is then able to mimic the activity of a horse raddish peroxidase. Hence it is capable to produce reactive oxygen species (ROS) by the reduction of hydrogen peroxide to water. These ROS then leads to an activation of luminol which then becomes chemiluminescent. In several of our subprojects we exploit this function as an easy readout such as for the AptaBody.
 
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Looking at the history of functional RNA in iGEM naturally occurring self-splicing introns were used by Team Waterloo 2011 to rejoin a split GFP posttranscriptionally. Riboswitches the natural relatives of aptamers have been used in several projects for example Cambridge 2012, Goettingen 2013 and also Bettencourt 2015. Hammerhead ribozymes have been discovered by several teams as handy tools (Peking 2011, Fudan 2012 and MIT 2011). DTU Denmark 2014 worked with the Spinach aptamer. Although functional RNA has been used in some iGEM projects DNA or DNAzymes have rarely been mentioned before. Our project this year aims to modularize and standardize all these and many more parts. With our new RFC 110 we simplify the work with RNA and thus want to encourage future teams to also dive into the world of RNA. By the addition and modularization of many DNA parts we want to stress to possibilities DNA possesses.
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By providing the community with tools for different applications, we contribute a foundational advance in the work with functional RNA. Not only is the system highly modular and therefore predisposed for standardization and usage in Synthetic Biology, the software developed by our team enables scientists to create aptamers and aptazymes <i>de novo</i>. Thereby we open up the world of functional nucleic acids to the entire Synthetic Biology community. Dive deeper now!
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Revision as of 10:45, 18 September 2015

Catalytic DNA – DNAzymes

SELEX revealed a variety of DNA aptamers and catalytic DNA known as DNAzymes (or Deoxribozymes). Breaker and Joyce selected the first DNAzyme, a RNA-cleaving DNA.breaker1994 In 1997 Santoro and Joyce describe the 7-18 and 10-23 DNAzyme santoro1997 which since then have been of special interested in DNAzyme research. Both recognize their substrate RNA by Watson-Crick base pairing and catalyze the attack of a 2’ hydroxyl group that leads to the formation of a 2’,3’-cyclic phosphate and leaves a 5’ hydroxyl RNA terminus.santoro1998 Their activity has also been engineered to be dependent on the presence of a small molecule by the addition of an aptamer to its substrate recognition site.Wanga2002Wangb2002 To verify the prediction of the software we fused the calculated aptamers to a DNAzyme and thus created a ligand dependent activity.

For this we used the HRP-mimicking DNAzyme which was described in 1998.Travascio1998 It folds into a G-quadruplex that forms a complex with hemin and is then able to mimic the activity of a horse raddish peroxidase. Hence it is capable to produce reactive oxygen species (ROS) by the reduction of hydrogen peroxide to water. These ROS then leads to an activation of luminol which then becomes chemiluminescent. In several of our subprojects we exploit this function as an easy readout such as for the AptaBody.