dh5α  (Toyobo)

Journal: PLoS ONE

Article Title: Pseudomonas aeruginosa serA Gene Is Required for Bacterial Translocation through Caco-2 Cell Monolayers

doi: 10.1371/journal.pone.0169367

Figure Lengend Snippet: Fly survival experiments. Virulence of the wild-type strain (WT), PAO1Tn:: serA mutant ( ΔserA ), PAO1Tn:: serA (pUCP19- serA ) complementary strain (+ serA ), E . coli DH5α (as a negative control), and WT in the presence of L -serine (10, 20, 30, 40, and 50 mM) was evaluated. Significant differences, based on the log-rank test, were observed between WT and DH5α (*: P

Article Snippet: E . coli DH5α strain was used as a control strain that does not penetrate epithelial cell monolayers [ ] and was purchased from TOYOBO, Japan.

Techniques: Mutagenesis, Negative Control

Journal: PLoS ONE

Article Title: Pseudomonas aeruginosa serA Gene Is Required for Bacterial Translocation through Caco-2 Cell Monolayers

doi: 10.1371/journal.pone.0169367

Figure Lengend Snippet: Bacterial adherence to Caco-2 cells for the wild-type strain (WT), PAO1Tn:: serA mutant ( ΔserA ), PAO1Tn:: serA (pUCP19- serA ) complementary strain (+ serA ), PAO1Tn:: flgE mutant ( ΔflgE ), E . coli DH5α (as a negative control), and WT in the presence of L -serine (20, 30, 40, and 50 mM). Bacterial adherence was determined based on the number of adhered bacteria per Caco-2 cell. The assay was performed in six replicates, and the results are expressed as the mean ± SD. Significant differences were observed between WT and ΔserA (*: P

Article Snippet: E . coli DH5α strain was used as a control strain that does not penetrate epithelial cell monolayers [ ] and was purchased from TOYOBO, Japan.

Techniques: Mutagenesis, Negative Control

Journal: PLoS ONE

Article Title: Pseudomonas aeruginosa serA Gene Is Required for Bacterial Translocation through Caco-2 Cell Monolayers

doi: 10.1371/journal.pone.0169367

Figure Lengend Snippet: Penetration activity of P . aeruginosa strains. (A) The wild-type strain (WT), PAO1Tn:: serA mutant ( ΔserA ), PAO1Tn:: serA (pUCP19- serA ) complementary strain (+ serA ), and PAO1Tn:: flgE mutant ( ΔflgE ) were inoculated onto the apical surfaces of Caco-2 cell monolayers at an MOI of 100, and the number of bacteria in the basolateral medium was counted at 6 h after infection. The assay was performed in triplicate, and the results are expressed as the mean ± SD. E . coli DH5α was used as a negative control. *#: P

Article Snippet: E . coli DH5α strain was used as a control strain that does not penetrate epithelial cell monolayers [ ] and was purchased from TOYOBO, Japan.

Techniques: Activity Assay, Mutagenesis, Infection, Negative Control

Journal: PLoS ONE

Article Title: Pseudomonas aeruginosa serA Gene Is Required for Bacterial Translocation through Caco-2 Cell Monolayers

doi: 10.1371/journal.pone.0169367

Figure Lengend Snippet: PGDH activity assay. (A) SDS-PAGE analysis to verify overexpression of the P . aeruginosa serA and synthesis of SerA proteins in crude extract isolated from DH5α (ptac-85- serA ) culture (SerA) and from DH5α (ptac-85) culture (mock). The red arrow indicates an overexpressed band at the expected size of 44 kDa for P . aeruginosa SerA. (B) Inhibitory effect of 50 mM L -serine on PGDH activity of SerA proteins (mU/mg) in crude extract isolated from DH5α (ptac-85- serA ) culture (SerA) and from DH5α (ptac-85) culture (mock). The assay was performed in five replicates, and the results are expressed as the mean ± SD. Significant differences were observed between SerA and mock (*: P

Article Snippet: E . coli DH5α strain was used as a control strain that does not penetrate epithelial cell monolayers [ ] and was purchased from TOYOBO, Japan.

Techniques: Activity Assay, SDS Page, Over Expression, Isolation

Journal: PLoS ONE

Article Title: Pseudomonas aeruginosa serA Gene Is Required for Bacterial Translocation through Caco-2 Cell Monolayers

doi: 10.1371/journal.pone.0169367

Figure Lengend Snippet: Motility assays. (A) Swimming motility of WT, ΔserA , + serA , and E . coli DH5α (as a negative control). A representative image from the swimming motility assay is shown. The major axis of swimming is the longest length of the swimming area. The assay was performed in triplicate, and the results are expressed as the mean ± SD. Significant differences were observed between WT and ΔserA (*: P

Article Snippet: E . coli DH5α strain was used as a control strain that does not penetrate epithelial cell monolayers [ ] and was purchased from TOYOBO, Japan.

Techniques: Negative Control, Motility Assay

Journal: The EMBO Journal

Article Title: An RNA-dependent protein kinase is involved in tunicamycin-induced apoptosis and Alzheimer's disease

doi: 10.1038/sj.emboj.7600049

Figure Lengend Snippet: Selection of Rzs that suppress Tm-induced cell death from a randomized library. ( A ) Schematic representation of the selection system. ( B ) Number of transformants after each round of selection. Vectors isolated from surviving cells were introduced into DH5α. ( C ) Individual Rzs, selected from the randomized library, were examined for their ability to protect cells against Tm-mediated apoptosis. SK-N-SH cells that expressed selected Rzs were exposed to 2 μg/ml Tm for 24 h. Apoptosis was assessed by TUNEL staining. ( D ) The activation of caspase-3 depended on the activation of PKR. Cells were treated with Tm for 24 h or untreated (control), and then caspase-3 activity was measured. Cells treated with z-VAD-fmk were examined as controls.

Article Snippet: For the first screening, transfected cells were exposed to Tm for 24 h. For the second screening, cells were transfected with candidate plasmids from the first screening and collected after incubation with Tm for 48 h. Expression vectors harboring Rzs were isolated from surviving cells and used to transform DH5α (TOYOBO, Tokyo, Japan).

Techniques: Selection, Isolation, TUNEL Assay, Staining, Activation Assay, Activity Assay