Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Shiji Zhao
Date created: 2016-09-04 11:00:00
Date modified: 2016-10-14 08:28:15
Constitutive promotor J23105 and downstream CecropinXJ
| Types | DnaRegion |
| Roles | Device engineered_region |
| Sequences | BBa_K1919001_sequence (Version 1) |
Description
Biobrick BBa_K1919001 is a composite part, consisting of constitutive promotor J23105, RBS, TrxA tag, 6xHis tag, thrombin site, S tag, enterkinase site and the coding sequence of antimicrobial peptide CecropinXJ.Notes
Two steps were needed in order to obtain BBa_K1919001.Step 1, BBa_K1919000 was inserted into pET32a(+). The primers used for PCR are as following:
F: 5'-CCGGGATCCATGAACTTCGCTAAAATCCTGTCTT-3' (BamH I site was added)
R: 5'-CCGCTCGAGTTATTATTTACCGATAGCTTTA-3' (Xho I site was added)
BamH I/Xho I Double enzymatic cutting was used for both pET32a(+) and BBa_K1919000.
Step 2, RBS and 5 downstream tags as well as original CecropinXJ was inserted at the downstream of promotor J23105 with the same procedure as step 1. The primers used for PCR are as following:
F: 5'-GCTCTAGAGCGAAGGAGATATACATATGAGCG-3' (Xba I site was added)
R: 5'-TGCACTGCAGTGCAGACTAGTCGTTATTGCTCAGCGGTGG-3'(Pst I site was added)
Xba I/Pst I double enzymatic cutting was used for PCR product and Spe I/Pst I double enzymatic cutting was used for BBa_J23105. After enzymatic linkage, the construction of BBa_K1919001 was finished. Then this part was added to pSB1C3 backbone.
Source
The sequence of constitutive promotor J23105 comes from biobrick BBa_J23105. RBS and 5 downstream tags come from commercial plasmid pET32a(+). The sequence of CecropinXJ comes from biobrick BBa_K1919000.| Sequence Annotation | Location | Component / Role(s) |
| BBa_J23105 ATG start codon CecropinXJ coding sequence double TAA stop codon BBa_K1919000 RBS TrxA 6xHis Tag thrombin site S-Tag enterokinase site | 1,35 60,62 555,749 744,749 555,749 46,51 60,386 408,425 435,452 459,503 519,533 | feature/BioBrick engineered_region start_codon feature/start feature/cds CDS stop_codon feature/stop engineered_region feature/BioBrick ribosome_entry_site feature/rbs tag feature/tag tag feature/tag tag feature/tag feature/tag tag feature/tag tag |