LacZ-alpha (pUC19 like)
This part encodes the alpha fragment of the LacZ gene derived from the pUC19 cloning vector.
Usage and Biology
The LacZ-alpha fragment is used as a reporter for cloning experiments performed in E. coli strains that have the deletion of the LacZ-alpha, but bear the LacZ-omega part of the LacZ gene. The concept is known as the alpha-complementation: a functional ß-galactosidase is restored when the two individual non-functional fragments of the LacZ gene (alpha and omega) are co-expressed. The alpha-complementation is detected using X-gal (5-bromo-4-chloro-3-indoyl-β-d-galactopyranoside), a soluble colorless compound that is a substrate of ß-galactosidase and yields a blue product when cleaved.
LacZ-alpha fragment is much smaller then the LacZ-omega part and thus can be easily included in a plasmid. It is used to determine if a transformed bacterial colony has received a recombinant plasmid (i.e. +insert) or not. For this the cloning experiment should be designed in such that the LacZ-alpha is disrupted in the recombinant plasmid and functional in the non recombinant plasmid.
Many derivates of the LacZ-alpha fragment were developed to be included in various cloning vectors. This was based on an experimental observation that "as long as the N-terminal ∼41 amino acid residues are present, the length of the α-complementing peptide is not important". However, the intensity of the blue colour may be different between the LacZ-alpha fragment variants.
This part encodes the alpha fragment of the LacZ gene derived from the pUC19 cloning vector. We choose this version based on the experimental observation that the E. coli colonies transformed with pUC19 vector have a deep blue colour on X-gal containing media.
The pUC19 DNA sequence encoding LacZ-alpha was modified with the help of JCAT in order to remove the numerous cloning site present especially in the MCS of pUC19.
Our team used this version of LacZ-alpha to build the Universal Biosensing Chassis (BBa_K2448023).
Sequence and Features
- 10COMPATIBLE WITH RFC
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- 1000COMPATIBLE WITH RFC