Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Alex Duryee, Grace Li
Date created: 2015-06-09 11:00:00
Date modified: 2015-11-11 04:24:59
rClone Tet Red: Device for GGA Cloning and Testing RBS elements and Riboswitches
| Types | DnaRegion |
| Roles | Device engineered_region |
| Sequences | BBa_J119388_sequence (Version 1) |
Description
rClone Tet Red allows users to clone and test new RBS elements and riboswitches without gel purification or other preparation of DNA. It is a destination vector for Golden Gate Assembly (GGA) using BsaI and ligase. A new RBS or riboswitch can be derived from synthetic oligos, PCR, or a plasmid clone. For proper assembly, the new insert must have the appropriate 4 nt sticky ends or be flanked by BsaI sites that produce the sticky ends. With reference to the top strand, the left site must be 5' CGAC 3' and the right site must be 5' GCGG 3'. BsaI always produces a 5' overhang sticky end. Successful GGA assembly replaces the reverse promoter driving GFP expression with the new RBS or riboswitch. Transcription will be initiated in the direction of the TetA-RFP fusion protein coding sequence. The level of expression of TetA-RFP fusion protein will depend on the efficiency of the newly cloned RBS or riboswitch.Notes
None.Source
PCR of J119385 and J119384 and BbsI GGA.| Sequence Annotation | Location | Component / Role(s) |
| P5 BsaI Left GFP Reverse B0030 RBS reverse P2 promoter reverse BsaI right Start codon TetA protein half Linker Sequence RFP protein half Stop codon | 1,36 42,47 55,774 781,792 803,848 849,854 862,864 862,2052 2053,2100 2101,2781 2779,2781 | promoter feature/promoter sequence_feature feature/dna CDS feature/cds feature/rbs ribosome_entry_site promoter feature/promoter feature/dna sequence_feature feature/start start_codon feature/cds CDS feature/cds CDS CDS feature/cds feature/stop stop_codon |