Difference between revisions of "Team:Austin UTexas/Results/Assemblies"
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<p>Assembly plasmids are assembled using a 1-5 bridge connector and part plasmids 6-8b in a single BsaI golden gate reaction. An assembled plasmid then contains an antibiotic resistance gene, origin of replication, origin of transfer, barcode sequence, and a part 1-5 bridge, which contains a colored reporter protein coding sequence. The same promoters, terminators, and origins of transfer are used within all assemblies. Each 1-5 bridge connector has a different barcode and reporter protein coding sequence making it easy to determine which plasmid each colony contains. This design eliminates as many variables as possible, allowing the origin of replication to be the single largest factor determining whether the bacteria sustain and express the plasmid. Using golden gate assembly, we were able to assemble these plasmids in just one reaction with the help of overhangs common to each part type.<P> | <p>Assembly plasmids are assembled using a 1-5 bridge connector and part plasmids 6-8b in a single BsaI golden gate reaction. An assembled plasmid then contains an antibiotic resistance gene, origin of replication, origin of transfer, barcode sequence, and a part 1-5 bridge, which contains a colored reporter protein coding sequence. The same promoters, terminators, and origins of transfer are used within all assemblies. Each 1-5 bridge connector has a different barcode and reporter protein coding sequence making it easy to determine which plasmid each colony contains. This design eliminates as many variables as possible, allowing the origin of replication to be the single largest factor determining whether the bacteria sustain and express the plasmid. Using golden gate assembly, we were able to assemble these plasmids in just one reaction with the help of overhangs common to each part type.<P> | ||
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| + | <img class="left" src= "https://static.igem.org/mediawiki/2018/d/d9/T--Austin_UTexas--AssemblyPlasmidParts.jpeg" width= "200" height= "300"   img class="center" src = "https://static.igem.org/mediawiki/2018/c/c1/T--Austin_UTexas--AssemblyPlasmid.jpeg" width= "200" height= "300" | ||
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<img class="resize" src = "https://static.igem.org/mediawiki/2018/b/b3/T--Austin_UTexas--AnnotatedCAMplate.jpg" width="300" height="300" > | <img class="resize" src = "https://static.igem.org/mediawiki/2018/b/b3/T--Austin_UTexas--AnnotatedCAMplate.jpg" width="300" height="300" > | ||
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<figcaption> | <figcaption> | ||
<b> Assembly plasmid BHR904. 901.1 Bridge connector (GFP), CAM 8b antibiotic resistance, p15A origin of replication, barcode 3, and origin of transfer </b> | <b> Assembly plasmid BHR904. 901.1 Bridge connector (GFP), CAM 8b antibiotic resistance, p15A origin of replication, barcode 3, and origin of transfer </b> | ||
Revision as of 06:16, 16 October 2018
Assembly Plasmids
Assembly Plasmids
Assembly plasmids are assembled using a 1-5 bridge connector and part plasmids 6-8b in a single BsaI golden gate reaction. An assembled plasmid then contains an antibiotic resistance gene, origin of replication, origin of transfer, barcode sequence, and a part 1-5 bridge, which contains a colored reporter protein coding sequence. The same promoters, terminators, and origins of transfer are used within all assemblies. Each 1-5 bridge connector has a different barcode and reporter protein coding sequence making it easy to determine which plasmid each colony contains. This design eliminates as many variables as possible, allowing the origin of replication to be the single largest factor determining whether the bacteria sustain and express the plasmid. Using golden gate assembly, we were able to assemble these plasmids in just one reaction with the help of overhangs common to each part type.
