Description:
pKYB1 is an E. coli expression vector for use with the IMPACT™ Kit (NEB #E6901) (1,2). It is designed for in-frame insertion of a target gene into the polylinker, upstream of the Sce VMA intein/chitin binding domain (55 kDa)(1,3). pKYB1 carries the kanamycin resistance gene (Kn) from Tn 903. This double stranded vector is 8,393 bp in length.
Source:
pKYB1 is isolated from an E. coli strain (r- m-) by a standard plasmid purification procedure.
Advantages:
• A pBR322 derivative.
• The NdeI site in the polylinker contains an ATG sequence for translation initiation.
• Use of the SapI site allows for cloning of the target protein adjacent to the intein, resulting in purification of the target protein without any additional amino acids at its C-terminus.
• Unique sites are in bold. NheI and NruI are not unique.
• Expression of the fusion gene is under the stringent control of the T7 promoter (4) and is regulated by IPTG due to the presence of a lac I gene.
• Origin of DNA replication from bacteriophage M13, which allows for the production of single-stranded DNA by helper phage superinfection of cells bearing the plasmid (M13K07 Helper Phage, NEB #N0315).
• Kanamycin resistant.
•Compatible restriction sites for subcloning a fusion gene from other IMPACT vectors.
• A wide range of E. coli host strains: T7 Express Competent E.coli (High Efficiency) (NEB #C2566) or BL21(DE3) Competent E. coli (NEB #C2527) and derivatives.
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