Autolytic E. coli Cell Lines
For easy purification of recombinant proteins and plasmid DNA
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Legend: brief treatment of plasmid-containing NTC-DH5-AL cells (with EDTA and Triton X100) partially disrupts outer and inner bacterial cell membranes, while chromosomally encoded lysozyme punctures the peptidoglycan cell wall, creating a molecular sieve, whereby plasmid DNA leaks from the cells, thus preventing viscous lysate, and leaving bacterial chromosomal DNA behind in the cell. |
• Simplified purification of recombinant proteins and plasmids
• Stable, chromosomally integrated genes
• Heat or IPTG inducible lysozyme
• Derivatives of well-known strains: DH5a (plasmid production); or BL21 (recombinant proteins)
• Can be used with standard antibiotic resistance markers
Description:
Nature Technology Corporation (NTC) has developed autolytic (AL) Escherichia coli host strains that express a chromosomal, integrated lysozyme gene (λR) to permeabilize the bacterial cell wall. Expression of the endolysin is induced during growth by either a heat inducible (ALHEAT) or IPTG inducible (ALIPTG) promoter. The endolysin remains in the cytoplasm, where it is separated from its peptidoglycan substrate in the cell wall; hence the cells remain alive and intact and can be harvested by the usual methods. Cell lysis can then be performed using a freeze thaw cycle or with Triton X-100/EDTA lysis solutions.
NTC’s autolytic cell lines are derivatives of the popular DH5α and BL21 cells lines.
These cell lines can be used for autolytic purification of plasmid or protein. No reduction of growth rate, viability, plasmid yield or integrity, is observed. NTC3012 cells can be used in an acidic autolytic plasmid DNA extraction procedure to selectively extract plasmid DNA without releasing bulk genomic DNA and cell debris.
Features:
• Simple, regulated E. coli autolysis using chromosomally integrated stable cell lines
• Heat or IPTG-inducible phage λR endolysin
• Heat inducible λR (tightly repressed at 30ºC, induced at 42ºC)
• IPTG inducible λR (tightly repressed in glucose media, induced with IPTG)
• Tight regulation prior to induction
• High-level Endolysin production post-induction using optimized zwf- λR cistron
• Derivatives of established strains for plasmid (DH5α) or protein (BL21) production
• Streamlines and simplifies plasmid or protein extraction protocols
• Compatible with a variety of plasmid selection markers including ampR, kanR, chlorR
