Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Chris French
Date created: 2006-10-26 11:00:00
Date modified: 2015-08-31 04:08:46
lac promoter and lacZ
| Types | DnaRegion |
| Roles | Reporter engineered_region |
| Sequences | BBa_J33207_sequence (Version 1) |
Description
This part (submitted in pSB1A2) consists of the lac promoter and lacZ' gene, encoding the N-terminal 76 amino acid residues of LacZ, sufficient to complement the lacZ-delta-M15 mutation for blue-white selection on Xgal plates. A SacI site has been introduced at the 3' end, overlapping the XbaI site of the Biobrick prefix. This is designed to be used as a cloning vector for making new biobricks. PCR primers can be designed with a SacI site in one primer and an SpeI site in the other. This removes the necessity for an excessively long non-complementary tail on one primer bearing either the full biobrick prefix or suffix. The PCR product can then be digested with SacI and SpeI for insertion into this plasmid, replacing the Plac-lacZ' cassette. Recombinant plasmids will then be white on IPTG/Xgal plates, whereas any that still contain the original insert will be blue. We have used this strategy to prepare several biobricks, including BBa_J33204, which contains the xylE gene encoding catechol-2,3-dioxygenase. (For making biobricks that contain lacZ', BBa_J33204 can be used in the same way; in this case, clones with plasmids that still contain xylE will turn yellow on addition of a drop of 10 mM catechol.)Notes
Note that the SacI site overlaps the SpeI site. The Biobrick prefix ends ...TCTAGAG. When this is added to the CTC at the start of the sequence shown here, the SacI site, GAGCTC, is generated.Source
The DNA was amplified from E. coli BL21 genomic DNA using primers based on published sequence (Genbank accession J01636, gi:146575). The annotation shown here is based on that associated with this Genbank entry. The sequence shown here is derived by sequencing the construct.| Sequence Annotation | Location | Component / Role(s) |
| SacI CAP binding site LacI binding site rbs lacZ' -10 -35 | 1,3 248,285 332,352 359,362 370,600 320,325 297,302 | sequence_feature feature/misc operator feature/operator feature/operator operator ribosome_entry_site feature/rbs feature/cds CDS feature/promoter promoter promoter feature/promoter |