A dual expression plasmid with protegrin-1 compatible with both prokaryotic and mammalian systems
Year: 2026
Type: article
Abstract: Antimicrobial peptide (AMP)-based positive selection is a powerful strategy for rapid gene cloning. A notable example is the use of the Escherichia coli ( E. coli ) CcdB toxin to eliminate non-recombinant plasmids. Building on this, dual prokaryotic/mammalian expression vectors like pgMAX and pgMAX-II minimize subcloning by allowing direct expression after a single ligation. However, combining selection modules (e.g., CcdB and Microcin B17) may affect recovery, particularly when their mechanisms both target DNA gyrase. In this study, we evaluated the feasibility of cloning and expressing the porcine AMP protegrin-1 (PG-1) using an AMP-assisted selection framework. PG-1 is an 18-residue, cysteine-rich cathelicidin that adopts a disulfide bond–stabilized β-hairpin structure. It exhibits potent antibacterial activity by permeabilizing anionic lipid bilayers through pore formation. We demonstrate that protegrin-1 functions as an effective selectable marker for plasmid recombination within the pgMAX-II system, achieving high cloning efficiency. This novel framework enables simple, high-efficiency subcloning with dual-expression capability in both E. coli and mammalian cells.
Source: MethodsX
Institution: Hirosaki University
Language: English
Subfield: Microbiology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $1,260
OpenAlex ID: W7160778141