Journal: Molecular microbiology

Article Title: c-di-AMP modulates Listeria monocytogenes central metabolism to regulate growth, antibiotic resistance and osmoregulation.

doi: 10.1111/mmi.13622

Figure Lengend Snippet: Fig. 1. Growth of c-di-AMP-deficient mutants in rich medium and resistance to cefuroxime. A and B. Mutants constructed and grown overnight in LSM were serially diluted 10-fold in a 96-well plate with PBS then 5 ml of each dilution was spotted onto either LSM or BHI agar. Images were taken and CFU were enumerated after 48 hours of incubation at 37˚C. C. Immunoblot of DacA and P60 (loading control) proteins for the strains indicated, grown to mid-log in LSM at 37˚C. Data are representative of three independent experiments. D. Antibiotic sensitivity measured by disk diffusion of 125 mg of cefuroxime on LSM-agar for the indicated L. monocytogenes strains measured at 48 hours. (B and D) Data are mean 6 standard error of the mean (s.e.m) of at least three independent experiments.

Article Snippet: Antibiotics were used at the following concentrations: carbenicillin (100 lg ml21, Gold Biotechnology), streptomycin (200 lg ml21, Sigma), chloramphenicol (7.5 lg ml21 for L. monocytogenes and 10 lg ml21 for E. coli, Sigma), erythromycin (1 lg ml21, Sigma), kanamycin (15 mg ml21 in LSM, 30 mg ml21 in BHI, Gold Biotechnology) and tetracycline (2 lg ml21, Sigma).

Techniques: Construct, Incubation, Western Blot, Control, Diffusion-based Assay

Journal: Protein engineering, design & selection : PEDS

Article Title: Engineered DNA ligases with improved activities in vitro.

doi: 10.1093/protein/gzt024

Figure Lengend Snippet: Fig. 1. Gel-based screens for ligase activity. (A) Ligation of cohesive-ended PCR products (638/639 bp; lower band) to yield a product of 1277 bp (upper band). Lanes are numbered and loaded as follows: 1, molecular weight marker; 2, ligation catalyzed by Sso7d-ligase; 3, cTF-ligase; 4, ligase-cTF; 5, p50-ligase; 6, ligase-p50; 7, NFAT-ligase; 8, ligase-NFAT; 9, molecular weight marker; 10, PprA-ligase; 11, ligase-PprA; 12, Ku-ligase; 13, ligase-Ku; 14, T4 DNA ligase; 15, no ligase control; 16, [(HhH)2]2-ligase; 17, ligase-[(HhH)2]2; 18, molecular weight marker; 19, T4 DNA ligase. (B) Ligase-catalyzed joining of a blunt-ended 717-bp restriction fragment (lower band) to give a 1434-bp product (upper band). Lanes are numbered and loaded as described in (A). (C) Fraction of cohesive-ended substrate that was ligated to product, for selected ligases from (A) and (B). Data are the means (+SEM) of two independent experiments, conducted under identical conditions to the assay in (A). (D) Fraction of blunt-ended substrate that was ligated to product, for selected ligases from (A) and (B). Data are the means (+SEM) of two independent experiments, conducted under identical conditions to the assay in (B).

Article Snippet: D ow nloaded from https://academ ic.oup.com /peds/article/26/7/471/1477749 by guest on 16 June 2024 substrate ( 40 nM ligatable ends) in 10 ml of 1 New England Biolabs E.coli DNA Ligase Reaction Buffer (30 mM Tris-HCl, 4 mM MgCl2, 1 mM dithiothreitol, 26 mM NAD þ, 50 mg ml21 BSA, pH 8.0).

Techniques: Activity Assay, Ligation, Molecular Weight, Marker, Control