e coli top10  (Thermo Fisher)

Journal: Antibiotics

Article Title: Development of New Antimicrobial Peptides by Directional Selection

doi: 10.3390/antibiotics14111120

Figure Lengend Snippet: Schematic overview of the experimental approach. A library of expression plasmids encoding random mutants of well-characterized antimicrobial peptides (Hm-AMP2, melittin and cecropin) was generated by random mutagenesis with partially degenerate oligonucleotides. The plasmid library was transformed into E. coli , followed by induction of recombinant AMP expression. Clones with the strongest growth inhibition were selected for subsequent rounds of mutagenesis and screening. The most promising variants were synthesized and tested for their properties.

Article Snippet: The strain used for recombinant plasmid construction was E. coli TOP10 (F − mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 nupG recA1 araD139 Δ(ara-leu)7697 galE15 galK16 rpsL(Str^R) endA1 λ − ) (Invitrogen, Carlsbad, CA, USA).

Techniques: Expressing, Generated, Mutagenesis, Plasmid Preparation, Transformation Assay, Recombinant, Clone Assay, Inhibition, Synthesized

Journal: Antibiotics

Article Title: Development of New Antimicrobial Peptides by Directional Selection

doi: 10.3390/antibiotics14111120

Figure Lengend Snippet: Activity of synthetic peptides melittin ( A ), cecropin ( B ), and Hm-AMP2 ( C ) in relation to introduced mutations. In the alignment on the left, shades of blue indicate amino acid sequence similarities. On the right, antimicrobial activity against E. coli and B. subtilis , cytotoxicity toward Expi293F cells, and the calculated therapeutic index based on peptide activities are shown. MEL—melittin, CECR—cecropin, AMP2—Hm-AMP2 peptide. AMP’s cytotoxicity was measured as the IC 90 , the concentration that inhibits 90% of cell growth. NA (not applicable) indicates that peptide showed no activity within tested concentration range. Minimal inhibitory concentration (MIC) and IC 90 are given in µM. The therapeutic index (TI) was defined as the lowest IC 90 divided by the highest MIC.

Article Snippet: The strain used for recombinant plasmid construction was E. coli TOP10 (F − mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 nupG recA1 araD139 Δ(ara-leu)7697 galE15 galK16 rpsL(Str^R) endA1 λ − ) (Invitrogen, Carlsbad, CA, USA).

Techniques: Activity Assay, Sequencing, Concentration Assay