Difference between revisions of "Team:Warwick/Protocols"
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<h4>Experiment 1: Testing the binding of specifically designed DNA strands to glass slides</h4> | <h4>Experiment 1: Testing the binding of specifically designed DNA strands to glass slides</h4> | ||
| − | <br>Glass slides were prepared <a href="https://static.igem.org/mediawiki/2015/f/f8/WarwickGlassSlideProtocol.pdf">Glass Slide Preperation Protocol</a> by being cleaned and functionalised (with HCl and GOPTS respectively). | + | |
| − | <br>Specifically designed oligonucleotides containing zinc finger binding domains were introduced to the slides. | + | <ul> |
| − | <br>These oligonucleotides comprise of a general adaptor strand, a specific short strand and a specific long strand. | + | <br><li>Glass slides were prepared <a href="https://static.igem.org/mediawiki/2015/f/f8/WarwickGlassSlideProtocol.pdf">Glass Slide Preperation Protocol</a> by being cleaned and functionalised (with HCl and GOPTS respectively).</li> |
| − | <br>Terminal amine groups within the oligonucleotides bind (by a nucleophilic addition reaction) to the epoxy group of GOPTS, sticking the DNA to the glass slides. | + | <br><li>Specifically designed oligonucleotides containing zinc finger binding domains were introduced to the slides.</li> |
| − | <br>The presence of an EcoR1 cut site in the oligonucleotide allows us to have an extra level of control in our experiments. Although the zinc finger proteins will stay attached to their binding domains, cutting the oligonucleotide at this site allows cells to become ‘unstuck’ from the slides. | + | <br><li>These oligonucleotides comprise of a general adaptor strand, a specific short strand and a specific long strand.</li> |
| + | <br><li>Terminal amine groups within the oligonucleotides bind (by a nucleophilic addition reaction) to the epoxy group of GOPTS, sticking the DNA to the glass slides.</li> | ||
| + | <br><li>The presence of an EcoR1 cut site in the oligonucleotide allows us to have an extra level of control in our experiments. Although the zinc finger proteins will stay attached to their binding domains, cutting the oligonucleotide at this site allows cells to become ‘unstuck’ from the slides.</li> | ||
<br>Expected results: | <br>Expected results: | ||
| − | <br>Slides treated with GOPTS should show fluorescence. | + | <br><li>Slides treated with GOPTS should show fluorescence.</li> |
| − | <br>Untreated cells should show no fluorescence. | + | <br><li>Untreated cells should show no fluorescence.</li> |
<br>This is a control to ensure that our zinc finger binding domains can be bound to glass slides. | <br>This is a control to ensure that our zinc finger binding domains can be bound to glass slides. | ||
| − | + | </ul> | |
Revision as of 14:29, 17 September 2015