Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Laura van Smeden
Date created: 2015-09-02 11:00:00
Date modified: 2015-09-18 05:40:03
Outer Membrane Protein X (OmpX) with BsoBI-linker and NanoLuc
| Types | DnaRegion |
| Roles | Coding CDS |
| Sequences | BBa_K1761002_sequence (Version 1) |
Description
OmpX is an outer membrane protein with the C- and N-termini in the intracellulair domain. To be able to use OmpX as a scaffold, a non-natural amino acid needs to be introduced. This can be done by implementing the amber stop codon TAG in one of the loops of OmpX via a mutation. With a specific tRNA an azide-functionalized amino acid can be built in, which can be used for the SPAAC click chemistry reaction with DBCO functionalized groups.The BsoBI linker is a 213 bp long flexible GGSGGS linker. Using the restriction enzyme BsoBI, the linker can become 45 bp shorter.
NanoLuc is a small sized and bright luciferase. Under the addition of furimazine, the NanoLuc luciferase generates a bioluminescece emmission spectrum with a maximum around 460 nm.
Notes
The sequence is redesigned considering codon optimalization. Also restriction sites for biobricking and classical cloning are deleted. From the BsoBI linker, the coding sequecne for GGSGGS is redesigned causing that only two BsoBI restriction sites remain.Source
OmpX is a outer membrane protein naturally occuring in wildtype E.coli.The BsoBI linker is inspired by the article "Quantitative Understanding of the Energy Transfer between Fluorescent Proteins Connected via Flexible Peptide Liners" by Toon H. Evers et all from 29 August 2006.
NanoLuc naturally occurs in the organism Oplophorus gracilirostris and was optimalized by the company Promega.
| Sequence Annotation | Location | Component / Role(s) |
| BBa_K1761000 BsoBI linker NanoLuc | 1,513 514,726 727,1239 | engineered_region feature/BioBrick feature/dna sequence_feature feature/protein CDS |