Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Phoebe Parrish
Date created: 2013-06-27 11:00:00
Date modified: 2015-08-31 04:08:23
eCDM8 and Tetracycline Resistance Riboswitch into pSB1A8
| Types | DnaRegion |
| Roles | Other sequence_feature |
| Sequences | BBa_J100111_sequence (Version 1) |
Description
High promoter/RBS combination with eCDM8 (part J100101) plus promoter/riboswitch/tetA gene construct (J119140), ligated into pSB1A8 (J119043). This part was constructed using iPCR and PCR to amplify the desired sequences and add BsaI sites, and Golden Gate Assembly to ligate the sequences together. eCDM8 demethylates caffeine to produce theophylline, which in turn binds to the riboswitch, allowing the translation of tetA mRNA, producing a protein that gives cells with this plasmid tetracycline resistance. Therefore cell growth can be used as a reporter of theophylline production.Notes
This construct was built using iPCR primers to amplify the promoter/RBS/eCDM8 construct (J100101) on pSB1A8 and add to BsaI sites to the end of the amplified sequence. This amplification removed the stuffer and the XbaI site from pSB1A8. PCR primers were used to amplify the promoter/riboswitch/tetA construct (J119140) and to add BsaI sites to the end of this sequence. GGA was then used to ligate these two amplified parts together.Source
All parts taken from the Registry.| Sequence Annotation | Location | Component / Role(s) |
| eCDM8 P5 Promoter (J119031) BD2 Bicistronic Translational Junction (J119025) Double Stop T5 promoter tetA gene Stop Riboswitch | 122,3274 1,36 37,88 3269,3274 3273,3323 3430,4625 4623,4625 3394,3452 | sequence_feature feature/dna promoter feature/promoter feature/rbs ribosome_entry_site stop_codon feature/stop promoter feature/promoter sequence_feature feature/dna stop_codon feature/stop feature/misc sequence_feature |