Journal: Genome Biology

Article Title: NT-seq: a chemical-based sequencing method for genomic methylome profiling

doi: 10.1186/s13059-022-02689-9

Figure Lengend Snippet: NT-seq simultaneously detects three types of methylation motifs in bacteria. a A to G frequency at known 6mA sites (G6mATC, A6mACN 6 GTGC, and GC6mACN 6 GTT) and unmethylated A sites in E. coli MG1655 genome from native and PCR amplified DNA. b C to T frequency at known 4mC sites (T4mCTTC and 4mCCGG) and unmethylated C sites in H. pylori JP26 genome from native and PCR amplified DNA. c–e Negative log normalized A to G ratio of different 6mA motifs in E. coli WT strain MG1655 ( a ), hsdM deleted strain ( b ), and dam/dcm/hsdR mutated strain ( c ). 6mA position was underlined. f Negative log normalized C to T ratio of different 4mC motifs in H. pylori JP26. g Negative log normalized C to T ratio of 4mC sites identified by SMRT-seq in E. coli strain MG1655. h Negative log normalized C to T ratio of different 5mC motifs in H. pylori JP26. Only motifs with sequencing depth larger than 25× were considered for violin plots. Statistic test were performed by two-sided Mann-Whitney-Wilcoxon (MWW) test with Bonferroni correction (ns: P > 1.0e-3; ****: P ≤ 1.0e−6)

Article Snippet: WT E. coli strain MG1655 and hsdM deletion K12 strain were kindly provided by Dr. Susan M Rosenberg’s lab (Baylor College of Medicine). dam/dcm mutant E. coli strain JM110 was purchased from Addgene (Bacterial strain #49763).

Techniques: Methylation, Amplification, Sequencing, MANN-WHITNEY

Journal: iScience

Article Title: Biosensor and chemo-enzymatic one-pot cascade applications to detect and transform PET-derived terephthalic acid in living cells

doi: 10.1016/j.isci.2022.104326

Figure Lengend Snippet:

Article Snippet: E. coli K-12 MG1655 RARE , Prof. K.L.J. Prather ( ) , Addgene Bacterial strain #61440.

Techniques: Recombinant, Purification, Variant Assay, Mutagenesis, Plasmid Preparation, Software

Journal: mBio

Article Title: lon Deletion Impairs Persister Cell Resuscitation in Escherichia coli

doi: 10.1128/mbio.02187-21

Figure Lengend Snippet: Lon overexpression rescues colony formation in OFX-treated Δ lon cells. (A and B) WT E. coli MG1655 (A) and Δ lon E. coli MG1655 (B) containing the sulA-gfp reporter plasmid (pMSs201-P sulA -gfp ) were grown to stationary phase and diluted 100-fold in fresh LB medium. Diluted cells were then treated with 5 μg/mL OFX for 6 h. After treatment, phase-contrast and fluorescence (GFP) micrographs of cells were obtained. Images at 0 ( t = 0) and 6 h ( t = 6 h) from a representative replicate are shown, but similar data were obtained from four biological replicates ( n = 4). Scale bar, 25 μm. (C and D) E. coli cells (WT and Δ lon ) containing an inducible lon overexpression plasmid (pUA66- lon ) or empty vector control (pUA66-EV) were treated with OFX (5 μg/mL) for 6 h and transferred to LB agar plates supplemented with isopropyl β- d -1-thiogalactopyranoside (IPTG) at the indicated concentrations. CFU were measured for each strain before and after treatment. n = 4. (E) WT and Δ lon E. coli containing pUA66- lon were cultured with 1 mM IPTG before and/or during OFX treatment. After treatment, cells were transferred to agar plates with or without 1 mM IPTG. CFU were measured for each strain before and after treatment. n = 4. Statistical significance for pairwise comparisons was assessed using one-way analysis of variance (ANOVA) with Dunnett’s post hoc test. * * , P < 0.01; ** * , P < 0.001; *** * , P < 0.0001; ns, nonsignificant.

Article Snippet: E. coli MG1655 and the pQE-80L and pUA66 plasmids were obtained from Mark P. Brynildsen at Princeton University, and pXY027 was a gift from Jie Xiao (plasmid no. 98915; Addgene, Watertown, MA, USA) ( ).

Techniques: Over Expression, Plasmid Preparation, Fluorescence, Cell Culture

Journal: mBio

Article Title: lon Deletion Impairs Persister Cell Resuscitation in Escherichia coli

doi: 10.1128/mbio.02187-21

Figure Lengend Snippet: Lon overexpression rescues the colony-forming ability of Δ lon cells exposed to UV. (A) WT and Δ lon E. coli containing pMSs201-P sulA -gfp were grown to stationary phase, diluted 100-fold in fresh LB medium, and exposed to UV for the indicated times. After exposure, cells were cultured in a shaker for 2 h, and phase-contrast and fluorescence (GFP) micrographs of cells were obtained. Images from a representative replicate are shown, but similar data were obtained for four biological replicates. n = 4. Scale bar, 25 μm. (B) WT and Δ lon E. coli were exposed to UV as described in panel A and spotted onto LB agar plates to enumerate CFU. n = 4. (C to E) Cells containing the inducible lon overexpression plasmid (pUA66- lon ) or empty vector (pUA66-EV) control were exposed to UV for the indicated times and transferred to agar plates supplemented with IPTG at indicated concentrations to enumerate CFU. n = 4. Statistical significance for pairwise comparisons was assessed using one-way ANOVA with Dunnett’s post hoc test. ** * , P < 0.001; *** * , P < 0.0001; ns, nonsignificant.

Article Snippet: E. coli MG1655 and the pQE-80L and pUA66 plasmids were obtained from Mark P. Brynildsen at Princeton University, and pXY027 was a gift from Jie Xiao (plasmid no. 98915; Addgene, Watertown, MA, USA) ( ).

Techniques: Over Expression, Cell Culture, Fluorescence, Plasmid Preparation

Journal: mBio

Article Title: lon Deletion Impairs Persister Cell Resuscitation in Escherichia coli

doi: 10.1128/mbio.02187-21

Figure Lengend Snippet: Starvation in PBS solution rescues Δ lon persisters. (A) WT and Δ lon E. coli with or without the inducible lon overexpression plasmid (pUA66- lon ) or empty vector control (pUA66-EV) were grown to stationary phase and diluted 100-fold in fresh LB medium. Diluted cells were then treated with OFX for 6 h. After treatment, cells were washed, serially diluted, and spotted onto LB agar plates with or without IPTG (1 mM). Plates were incubated for 48 h, and images were captured at indicated time points. A representative replicate is shown, but similar data were obtained for four biological replicates. n = 4. (B) WT and Δ lon E. coli with or without pUA66- lon or pUA66-EV were treated with OFX for 6 h, transferred to sterile PBS solution with or without 1 mM IPTG, and incubated at 37°C in a shaker for 7 days. Samples were collected at the indicated time points and spotted onto agar plates to enumerate surviving cells. n = 4. Statistical significance for pairwise comparison was assessed using one-way ANOVA with Dunnett’s post hoc test. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, nonsignificant.

Article Snippet: E. coli MG1655 and the pQE-80L and pUA66 plasmids were obtained from Mark P. Brynildsen at Princeton University, and pXY027 was a gift from Jie Xiao (plasmid no. 98915; Addgene, Watertown, MA, USA) ( ).

Techniques: Over Expression, Plasmid Preparation, Incubation

Journal: Synthetic and Systems Biotechnology

Article Title: A linear DNA template-based framework for site-specific unnatural amino acid incorporation

doi: 10.1016/j.synbio.2021.07.003

Figure Lengend Snippet: Screening of 14 cell extracts. In this experiment, pPaF was selected for the UNAA embedding screening of 14 kinds of cell extracts. A total of 14 extracts were selected, including K-12 series (K-12, K-12 ΔtnaA, K-12 ΔtnaAΔsdaB), commercial series (BL21 (DE3), BL21 ΔserB, Rosetta (DE3), Rosetta-gami B (DE3), Origami, Origami B), rEc series (rEc. 13, rEc. 13. ΔA), EcAR7 series (EcAR7 ΔA ΔSer), C321 series (C321, C321. ΔA).

Article Snippet: A total of 14 E. coli extracts were selected in this study, including K-12 series (K-12 [ ] (Addgene #61440), K-12 ΔtnaA [ ], K-12 ΔtnaAΔsdaB [ ]), commercial series (BL21 (DE3) (Biomed, China), BL21 ΔserB [ ] (Addgene #34929), Rosetta (DE3), Rosetta-gami B (DE3), Origami, Origami B) (Biomed, China), rEc series (rEc.

Techniques: