Journal:

Article Title: Reverse Sanger sequencing of RNA by MALDI-TOF mass spectrometry after solid phase purification

doi: 10.1093/nar/gnh089

Figure Lengend Snippet: MALDI mass spectra of the RNA in vitro transcription performed with pGEM 3Zf linearized with (A) HindIII (sequence of 60mer: 5′-PPP-GGGCGAAUUCGAGCUCGGUACCCGGGGAUCCUCUAGAGUCGACCUGCAGGCAUGCAAGCU-3′) or with (B) SmaI (sequence of 23mer: 5′-PPP-GGGCGAAUUCGAGCUCGGUACCC-3′) and transcribed with T7 RNA polymerase utilizing unmodified NTP after EtOH-precipitation and ZipTip purification. The byproducts, abortive products (2–12 nt) and slippage products (G4–G7), are labeled according to their length.

Article Snippet: RNA was synthesized by runoff transcription from a linearized plasmid DNA vector pGEM 3Zf (Promega, Mannheim, Germany) carrying two promotors directed at each other from opposite ends of a multiple cloning site.

Techniques: In Vitro, Sequencing, Purification, Labeling

Journal:

Article Title: Reverse Sanger sequencing of RNA by MALDI-TOF mass spectrometry after solid phase purification

doi: 10.1093/nar/gnh089

Figure Lengend Snippet: MALDI mass spectra of RNA transcripts after solid-phase purification by hybridization using magnetic beads No. 2 (A) or magnetic beads No. 1 (B). In vitro transcription was performed with pGEM 3Zf linearized (A) with HindIII (sequence of 60mer: 5′-GGGCGAAUUCGAGCUCGGUACCCGGGGAUCCUCUAGAGUCGACCUGCAGGCAUGCAAGCU-3′). Signals marked with open circle: leakage bond of the universal complementary sequence coated to magnetic beads; (B) with SmaI (sequence of 23mer: 5′-PPP-GGGCGAAUUCGAGCUCGGUACCC-3′) and transcribed with T7 RNA polymerase utilizing unmodified NTP.

Article Snippet: RNA was synthesized by runoff transcription from a linearized plasmid DNA vector pGEM 3Zf (Promega, Mannheim, Germany) carrying two promotors directed at each other from opposite ends of a multiple cloning site.

Techniques: Purification, Hybridization, Magnetic Beads, In Vitro, Sequencing

Journal: Cell reports

Article Title: Dysregulation of Streptococcus pneumoniae zinc homeostasis breaks ampicillin resistance in a pneumonia infection model

doi: 10.1016/j.celrep.2021.110202

Figure Lengend Snippet: (A) Phenotypic impact of Zn, PBT2, and PBT2 + Zn on S. pneumoniae 23F growth in cation-adjusted Mueller-Hinton broth (CA-MHB) supplemented as indicated. The data correspond with the mean (± standard error of the mean) absorbance 600 nm measurements from three independent biological experiments. Error bars, where not visible, are overlapped by the representative symbols. (B) RNA sequencing of S. pneumoniae 23F to determine relative gene expression, expressed as log 2 -fold change. Each dot represents a gene, distributed on the x axis in accordance with locus tag numbering for 23F. Genes more highly expressed in the presence of 8 μM PBT2 + 32 μM ZnSO 4 are present above the x axis (pink), with those below the x axis expressed at a lower level (purple). Genes of interest are annotated with their putative or characterized functions. (C and D) Whole cell metal ion accumulation of Mn (C) and Zn (D) in S. pneumoniae 23F in the presence of PBT2, Zn, and PBT2 + Zn grown in CA-MHB. Error bars indicate standard deviation of the mean from three biological replicates, ns = p > 0.05, ** = p < 0.01, **** = p < 0.0001, one-way analysis of variance with the Tukey post-test. (E) Development of resistance assays for S. pneumoniae 23F during serial passage in the presence of sub-inhibitory concentrations of PBT2 + ampicillin or vancomycin, as a positive control, in CA-MHB. Experiment was terminated at 15 days owing to a loss of viability in the presence of PBT2 + ampicillin. Data represent three biological replicates. (F) Enumeration of colony-forming units (CFUs) of S. pneumoniae from the lungs of BALB/c mice, following intranasal challenge with 5 × 10 5 CFU of strain DAW30 (n = 10). Colonization was examined at 24 h after the challenge. BALB/c mice were treated with combinations of ampicillin, PBT2, ampicillin + PBT2 or vehicle at 0 h and 6 h after infection via oral gavage (PBT2, vehicle) and/or subcutaneous ampicillin. Data represent the mean (± standard deviation) of two independent experiments with statistical analyses performed by the Mann-Whitney U test. ns = p > 0.05, *** = p = 0.001.

Article Snippet: S. pneumoniae Spain23F; ATCC 700669 multi-drug resistant Spain 23F ST81 lineage , ( ) , N/A.

Techniques: RNA Sequencing, Gene Expression, Standard Deviation, Positive Control, Infection, MANN-WHITNEY

Journal: Cell reports

Article Title: Dysregulation of Streptococcus pneumoniae zinc homeostasis breaks ampicillin resistance in a pneumonia infection model

doi: 10.1016/j.celrep.2021.110202

Figure Lengend Snippet: Combination of PBT2 and Zn resensitizes pathogenic S. pneumoniae strains to antibiotics of various classes

Article Snippet: S. pneumoniae Spain23F; ATCC 700669 multi-drug resistant Spain 23F ST81 lineage , ( ) , N/A.

Techniques:

Journal: Cell reports

Article Title: Dysregulation of Streptococcus pneumoniae zinc homeostasis breaks ampicillin resistance in a pneumonia infection model

doi: 10.1016/j.celrep.2021.110202

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: S. pneumoniae Spain23F; ATCC 700669 multi-drug resistant Spain 23F ST81 lineage , ( ) , N/A.

Techniques: Virus, Recombinant, SYBR Green Assay, RNA Sequencing, Expressing, Construct, Plasmid Preparation, Software, Microscopy, Targeted Proteomics

Journal: Frontiers in Endocrinology

Article Title: Transcriptional Regulation of Vih by Oct4 and Sox9 in Scylla paramamosain

doi: 10.3389/fendo.2020.00650

Figure Lengend Snippet: Primers used in this study.

Article Snippet: These dsRNAs were synthesized from the linearized template by using the T7 RiboMAXTM Express RNAi System (Promega).

Techniques: Sequencing, Transfection, Expressing

Journal: Frontiers in Endocrinology

Article Title: Transcriptional Regulation of Vih by Oct4 and Sox9 in Scylla paramamosain

doi: 10.3389/fendo.2020.00650

Figure Lengend Snippet: Oct4 or Sox9 RNA interference reduces the expression level of SpVih . (A) Relative expression of Oct4 in the eyestalk after injection of dsOct4, dsEGFP, and SPSS. (B) Relative expression of Sox9 in the eyestalk after injection of dsOct4, dsEGFP, and SPSS. (C) The relative expression of SpVih in the eyestalk after injection of dsSox9, dsOct4, dsEGFP, and SPSS. NC, negative control group; BC, blank control group; Bar with different letters indicates significant differences, P < 0.05.

Article Snippet: These dsRNAs were synthesized from the linearized template by using the T7 RiboMAXTM Express RNAi System (Promega).

Techniques: Expressing, Injection, Negative Control, Control

Journal: Frontiers in Endocrinology

Article Title: Transcriptional Regulation of Vih by Oct4 and Sox9 in Scylla paramamosain

doi: 10.3389/fendo.2020.00650

Figure Lengend Snippet: Oct4 or Sox9 RNA interference promotes the expression level of SpVtg . (A) Relative expression of SpVtg in the ovary after injection of dsSox9, dsOct4, dsEGFP, and SPSS. (B) The relative expression of SpVtg in hepatopancreas after injection of dsSox9, dsOct4, dsEGFP, and SPSS. NC, negative control group; BC, blank control group; Bar with different letters indicates significant differences, P < 0.05.

Article Snippet: These dsRNAs were synthesized from the linearized template by using the T7 RiboMAXTM Express RNAi System (Promega).

Techniques: Expressing, Injection, Negative Control, Control