Journal:

Article Title: The Positive Regulator, TraJ, of the Escherichia coli F Plasmid Is Unstable in a cpxA * Background

doi: 10.1128/JB.184.20.5781-5788.2002

Figure Lengend Snippet: Strains and plasmids used in this study

Article Snippet: Standard genetic techniques were employed ( 46 ). table ft1 table-wrap mode="anchored" t5 TABLE 1. caption a7 Strain or plasmid Genotype or description a Source or reference Bacterial strains MC4100 F − araD139 Δ( argF-lac ) U169 rpsL150 (Str r ) relA1 flb5301 deoC1 ptsF25 rbsR 2 XK1200 Nal r F − lac Δ U124 Δ( nadA araG gal attL ) 34 TR8 MC4100 cpxA :: cam 42 TR20 MC4100 cpxA101 This work TR51 MC4100 cpxR :: spc 42 TR189 MC4100 cpxA101 zii ::Tn 10 λ RS88 [ degP-lacZ ] This work TR981 MC4100 cpxA101 zii ::Tn 10 λ RS88 [ degP-lacZ ] recA This work TR984 MC4100 cpxA101 zii ::Tn 10 λ RS88 [ degP-lacZ ] clpP lonA This work JMR201 MC4100 degP ::Tn 10 A. E. Rizzitello and T. J. Silhavy Plasmids pOX38-Km Km r F tra region, Rep FIA replicon 4 pOX38-Tc Tc r F tra region, Rep FIA replicon 1 pRS27 Tc r 9-kb partial Eco RI F fragmentin pSC101 50 pBAD24 Ap r P BAD cloning vector 16 pBADTraJ traJ coding region fused to P BAD This work pBADTraM traM coding region fused to P BAD This work pBADTraY traY coding region fused to P BAD This work pBR322 General cloning vector New England Biolabs pLD404 Ap r NlpE in pBR322 51 pMC874 Km r lacZ + Δ plac 3 pMCJ211 Km r traJ Ω lacZ FinP + traJ-lacZ reporter plasmid 57 pGEX-KG IPTG-inducible GST fusion expression Amersham Pharmacia Biotech pLJ5-13 Ap r T7 Φ 10-finP fusion in pUC19 19 pSLF20 finP F derivative 24 Open in a separate window a Str r , streptomycin resistant; Nal r , nalidixic acid resistant; Km r , kanamycin resistant; Tc r , tetracycline resistant; Ap r , ampicillin resistant.

Techniques: Plasmid Preparation, Clone Assay, Expressing

Journal: Journal of Bacteriology

Article Title: The MerR-Like Transcriptional Regulator BrlR Contributes to Pseudomonas aeruginosa Biofilm Tolerance

doi: 10.1128/JB.00765-12

Figure Lengend Snippet: Bacterial strains and plasmids used in this study

Article Snippet: Where necessary, Escherichia coli cultures were grown in LB broth in the absence or presence of 1 mM isopropyl-β- d -thiogalactopyranoside (IPTG). table ft1 table-wrap mode="anchored" t5 caption a7 Strain or plasmid Relevant genotype or description Source or reference Escherichia coli DH5α F − φ80d lacZ ΔM15 Δ( lacZYA - argF ) U169 recA1 endA1 hsdR17 (r K − m K + ) phoA supE44 thi - 1 gyrA96 relA1 tonA Invitrogen Corp. BL21 F − ompT hsdS B (r B − m B − ) gal dcm rne131 (DE3) Invitrogen Corp. P. aeruginosa PAO1 PAO1 Wild type B. H. Holloway PAO1/pJN105 PAO1 bearing empty pJN105 vector; Gm r 55 PAO1/pMJT1 PAO1 bearing empty pMJT1 vector; Carb r This study PAO1/pJN- brlR Arabinose-inducible expression of brlR ; Gm r This study PAO1/pMJT- brlR -V5/6×His Arabinose-inducible expression of V5/6×His-tagged brlR ; Carb r This study Δ brlR mutant PAO1 Δ brlR (PA4878) This study Δ brlR /pJN- brlR mutant Arabinose-inducible expression of brlR ; Gm r This study Δ brlR /pMJT1 mutant Δ brlR harboring empty pMJT1, vector control; Carb r This study Δ brlR /pMJT- brlR -N mutant Arabinose-inducible expression of N-terminal DNA binding domain of BrlR (BrlR-N) in Δ brlR ; Carb r This study CF1-2 Isolate from CF patient 53 CF1-8 Isolate from CF patient 53 CF1-8/pJN- brlR Arabinose-inducible expression of brlR in CF1-8; Gm r This study CF1-8 Δ brlR CF1-8 harboring Δ brlR (PA4878), Gm r This study CF1-13 Isolate from CF patient 53 CF1-13 Δ brlR CF1-13 harboring Δ brlR (PA4878); Gm r This study PAO1 attB :: lacZ pminiCTX- lacZ conjugated into PAO1, vector control; Tet r This study PAO1 attB ::PbrlR- lacZ pminiCTX-PbrlR- lacZ conjugated into PAO1; Tet r This study Plasmids pCR2.1-TOPO TA cloning vector; Km r Ap r Invitrogen Corp. pRK2013 Helper plasmid for triparental mating; mob tra Km r 25 pEX18Gm Gene replacement vector; pUC18 MCS oriT sacB Gm r 26 pJN105 Arabinose-inducible gene expression vector; pBRR-1 MCS araC -P BAD Gm r 50 pMJT1 Arabinose-inducible gene expression vector; pUCP18 MCS araC -P BAD Amp/Carb r 35 pET101D Vector for directional cloning and high level V5/6×His fusion protein expression; Amp r Invitrogen Corp. pminiCTX- lacZ attB site-specific integration vector; Tet r 10 pET- brlR -V5/6×His brlR cloned into pET101D This study pJN- brlR brlR cloned into pJN105 This study pMJT- brlR -V5/6×His brlR -V5/6×His cloned into pMJT1 This study pMJT- brlR -N N-terminal DNA binding domain of BrlR cloned into pMJT1 This study pminiCTX-PbrlR- lacZ brlR promoter reporter construct in mini-CTX- lacZ using PbrlR-lacZF/R primers; Tet r This study Open in a separate window Bacterial strains and plasmids used in this study

Techniques: Plasmid Preparation, Expressing, Mutagenesis, Control, Binding Assay, TA Cloning, Gene Expression, Cloning, Clone Assay, Construct

Journal: bioRxiv

Article Title: Biocatalytic Formation of Novel Polyesters with para -Hydroxyphenyl groups in the Backbone – Engineering Cupriavidus necator for production of high-performance materials from CO 2 and electricity

doi: 10.1101/2021.12.12.472320

Figure Lengend Snippet: 1 H-NMR spectra of P (3HB- co -4HB) produced by C. necator H16 Δ phaC1 pCM66T_P BAD - pct540 - phaC1437 (A) as well as P (3HB- co -4HB- co -6HC) (B), P (3HB- co -2H4PheB) (C), P (3HB- co -PheLA) (D), P (3HB- co -MA) (E), and P (3HB- co -PA) (F) produced by C. necator H16 Δ phaC1 pCM66T_P BAD - hadA - phaC1437 . Areas are indicative of the ratios for 3HB to non-natural repeat units. (A): In addition to the P3HB sextet resonance at 5.25 ppm, a triplet resonance at 4.11 ppm shows that the C4-methylene of the 4HB repeat is present, indicating incorporation of 4HB into the polymer. The large fraction of P4HB causes multiple P3HB peaks because of different possible orders in the polymer (e.g., -3HB-P3HB-P4HB-, -P4HB-P3HB-P4HB-, -P4HB-P3HB-P3HB-, -P3HB-P4HB-P3HB-). (B): In addition to the P3HB sextet resonance at 5.25 ppm and P4HB triplet resonance at 4.11 ppm, a triplet resonance at 4.07 ppm shows that the C6-methylene of the 6HC repeat is present, indicating incorporation of 6HC into the polymer. (C): In addition to the P3HB sextet resonance at 5.25 ppm, a triplet resonance at 5.13 ppm shows that the C2-methine of the 2H4PheB repeat is present. The broad aromatic multiplet resonance at 7.21 ppm is also indicative of P2H4PheB. (D): The P3HB sextet resonance at 5.25 ppm overlaps with the methine PPheLA resonance, so the methylene doublet resonances of 3.15 & 3.04 ppm and 2.58 & 2.48 ppm were used to show the presence of P3HB and PPheLA respectively. The broad aromatic resonance at 7.21 ppm is also indicative of PPheLA. (E): In addition to the P3HB sextet resonance at 5.25 ppm, a multiplet resonance at 5.12 ppm shows that the C2-methine of the MA repeat is present. The broad aromatic multiplet resonance at 7.28 ppm is also indicative of MA. (F): In addition to the P3HB sextet resonance at 5.25 ppm, two doublet resonances at 7.19 and 6.98 ppm show that the aromatic PA repeat is present, indicating incorporation of the para -hydroxyphenyl unit into the polymer.

Article Snippet: The plasmids pCM66T_P BAD - pct540 - phaC1437 and pCM66T_P BAD - hadA - phaC1437 were constructed by GenScript, cloning the synthetic operons containing pct540 & phaC1437 and hadA & phaC1437 , respectively, into pCM66T_P BAD -RFP (cf. supplementary material 2).

Techniques: Produced, Polymer

Journal: bioRxiv

Article Title: Biocatalytic Formation of Novel Polyesters with para -Hydroxyphenyl groups in the Backbone – Engineering Cupriavidus necator for production of high-performance materials from CO 2 and electricity

doi: 10.1101/2021.12.12.472320

Figure Lengend Snippet: Molecular weight distribution (M n and M W ) for PHA obtained from wild-type C. necator H16, and engineered C. necator H16 (Δ phaC1 pCM66T_P BAD - hadA - phaC1437 , non-induced and fully induced, respectively).

Article Snippet: The plasmids pCM66T_P BAD - pct540 - phaC1437 and pCM66T_P BAD - hadA - phaC1437 were constructed by GenScript, cloning the synthetic operons containing pct540 & phaC1437 and hadA & phaC1437 , respectively, into pCM66T_P BAD -RFP (cf. supplementary material 2).

Techniques: Molecular Weight

Journal: bioRxiv

Article Title: Biocatalytic Formation of Novel Polyesters with para -Hydroxyphenyl groups in the Backbone – Engineering Cupriavidus necator for production of high-performance materials from CO 2 and electricity

doi: 10.1101/2021.12.12.472320

Figure Lengend Snippet: Cultivation-profile (growth by means of OD) of representative experiment with transgenic C. necator H16 Δ phaC1 pCM66T_P BAD - hadA - phaC1437 in bio-electrochemical system (BES). Induction of expression, addition of aromatic polymer-precursor and depletion of nitrogen source as indicated. The insert on the bottom right shows a schematic of the BES, which evolves H 2 at the cathode and O 2 at the anode as electron carrier and acceptor for autotrophic growth and production of aromatic polyesters from CO 2 and electricity.

Article Snippet: The plasmids pCM66T_P BAD - pct540 - phaC1437 and pCM66T_P BAD - hadA - phaC1437 were constructed by GenScript, cloning the synthetic operons containing pct540 & phaC1437 and hadA & phaC1437 , respectively, into pCM66T_P BAD -RFP (cf. supplementary material 2).

Techniques: Transgenic Assay, Expressing, Polymer

Journal: Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association

Article Title: Pharmaceutical agent cetylpyridinium chloride inhibits immune mast cell function by interfering with calcium mobilization.

doi: 10.1016/j.fct.2023.113980

Figure Lengend Snippet: Fig. 3. CPC effects on mitochondrial Ca2þ in Ag- stimulated RBL mast cells. RBL mast cells were transiently transfected with CEPIA2mt construct, and the mitochondrial Ca2+ levels were measured using a plate reader. Cells were sensitized with IgE for 1 h, pretreated with 0 or 10 μM CPC in BT buffer for 30 min, and exposed to ±0.005 μg/mL Ag with 0 or 10 μM CPC in BT buffer for 1 h. Representative graphs show CEPIA2mt fluorescence immediately following Ag/CPC exposure (A); average background fluores cence, from untransfected cells, was subtracted from all time points prior to plotting. Area under the curve (AUC) was calculated using the background- subtracted fluorescence time-series graph; these values were then normalized to Ag + 0 μM CPC on that day (B). Values presented in (A) are means ± SD for a single experiment with three replicates per treatment. Values presented in (B) are means ± SEM of at least three independent experiments, each experiment was a triplicate. Statistically significant result, as compared to Ag, is represented by ***p < 0.001, determined by an unpaired one-tailed t-test.

Article Snippet: RBL-2H3 cells were transfected with the mitochondrial calcium indicator pCMV CEPIA2mt (a gift from Masamitsu Iino; Addgene plasmid # 58,218; htt p://n2t.net/addgene: 58,218; PRID:Addgene_58,218) (Suzuki et al., 2014) via nucleofection using Amaxa Nucleofector kit T (Lonza), as done in (Weatherly et al., 2018).

Techniques: Transfection, Construct, Fluorescence, One-tailed Test