Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Chris French
Date created: 2006-10-25 11:00:00
Date modified: 2015-08-31 04:08:46
Bacillus subtilis ars promoter and arsR gene plus E. coli lacZ
| Types | DnaRegion |
| Roles | engineered_region Reporter |
| Sequences | BBa_J33206_sequence (Version 1) |
Description
This part includes the arsenate-responsive ars promoter from Bacillus subtilis (Sato, T., and Kobayashi, Y. 1998. J. Bacteriol. 180, 1655-1661), together with the arsR gene encoding the negatively autoregulated repressor ArsR, plus the Escherichia coli lacZ' gene encoding the N-terminal 137 amino acid residues of LacZ, sufficient to complement the chromosomal lacZ-delta-M15 mutation carried by many common lab strains of E. coli. This construct is designed to give LacZ' expression in the presence of arsenic as arsenate or arsenite anion. In previous experiments using other reporter genes such as xylE (available as biobrick BBa_J33204), we have found that this promoter gives a more rapid and stronger response than the E. coli chromosomal ars promoter (available as biobrick BBa_J33201, and fused to lacZ' as BBa_J33203), but with higher background activity in the absence of arsenate. Our experiments with this particular construct showed no obvious inducibility of a pH response due to a high background expression level in the absence of arsenate; however, ONPG assays have not yet been done. This construct does not include a terminator, so another reporter gene could be added at the 3' end for further experiments.Notes
The B. subtilis ars promoter and arsR gene were amplified as a biobrick-like component, but unfortunately a base of the biobrick suffix was inadvertently omitted, so that this could not be submitted alone as a proper biobrick. The lacZ' component is similar to that submitted as BBa_J33202, but differs in including a larger amount of DNA, encoding 137 amino acid residues rather than 76. This was isolated as an unexpected minor product from the PCR reactions which gave BBa_J33202. It appears to complement the lacZ-delta-M15 mutation equally well. The two components were joined as biobricks to generate the construct given here.Source
The B. subtilis ars promoter and arsR gene were amplified from B. subtilis 168 genomic DNA using primers based on the published genome sequence. The E. coli lacZ' was amplified from E. coli BL21 genomic DNA using primers based on published sequence data (Genbank accession J01636, gi:146575).| Sequence Annotation | Location | Component / Role(s) |
| ArsR binding site -35 -10 rbs arsR fusion site rbs lacZ' | 98,115 118,123 142,147 172,178 187,504 505,510 510,516 524,934 | feature/operator operator feature/promoter promoter feature/promoter promoter feature/rbs ribosome_entry_site feature/cds CDS sequence_feature feature/misc ribosome_entry_site feature/rbs CDS feature/cds |