Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Binglin Liu
Date created: 2016-10-11 11:00:00
Date modified: 2016-10-12 11:31:43
RBS and terephthalic acid(TPA) transporter
| Types | DnaRegion |
| Roles | Translational_Unit engineered_region |
| Sequences | BBa_K2013004_sequence (Version 1) |
Description
The sequence in this part is one of sequences encoding TPA transporter. Because of it,TPA will be transported from extracellular to intracellular.In addition，We carried out codon optimization on this part.Experimental Validation
This part is validated through Four ways: amplification, PCR, Enzyme cutting and Sequence.
Amplification
Enzyme:Q5
Primer-F:5′GAATTCGCGGCCGCTTCTAGAGTACTAGAGTCACACAAGAAGGTACTAGATGAAG-3′
Primer-R:5′- CGCTACTAGTATTATTATTGGCCAAAAATGGTCGG-3′
Results
PCR
Enzyme:Taq
Primer-F:5′-CCACCTGACGTCTAAGAAAC-3′
Primer-R:5′-GTATTACCGCCTTTGAGTGA-3′
Results
Double digestion
After the assembly ,the plasmid was transferred into the Competent E. coli top10. After culturing overnight in LB,we minipreped the plasmid for double digestion .The first cutting procedure was performed with EcoRI and NcoI restriction endonuclease. The second cutting procedure was performed with EcoRV and PstI restriction endonuclease.
The plasmid was cutted in a 25μL system at 37 ℃ for 1 hours. The Electrophoresis was performed on a 1％ Agarose glu.
Results