lipofectamine rnaimax lipid reagent  (Thermo Fisher)


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    Name:
    Lipofectamine RNAiMAX Transfection Reagent
    Description:
    Lipofectamine RNAiMAX Transfection Reagent provides the highest transfection efficiencies on the widest variety of cell types for siRNA mediated gene knockdown experiments Lipofectamine RNAiMAX is a proprietary RNAi specific cationic lipid formulation designed specifically for the delivery of siRNA and miRNA into all cell types With Lipofectamine RNAiMAX Transfection Reagent you will get • Superior transfection efficiency requiring lower RNAi concentrations leading to more effective gene knockdown with minimal nonspecific effects• Easy optimization due to minimal cytotoxicity across a 10 fold concentration range of transfection reagent• Superior transfection efficiencies for miRNA antagonists and mimics• Compatibility with a broad range of cell types providing the most versatile approach to all of your gene silencing experiments• A simple and rapid protocol for consistent and reproducible resultsHigh knockdown in a wide range of cellsLipofectamine RNAiMAX Transfection Reagent transfects a wide range of cell types see figure For gene silencing Lipofectamine RNAiMAX Transfection Reagent s high efficiency transfections lead to the high levels of gene knockdown needed to achieve convincing results Simple high throughput ready transfectionsSimply mix Lipofectamine RNAiMAX Transfection Reagent with siRNA add to your cells incubate and measure gene knockdown The simplicity and speed combined with high transfection efficiency make Lipofectamine RNAiMAX Transfection Reagent ideal for high throughput siRNA transfections Transfection conditions can be easily established for automated or robotic systems used in such applications Find an optimized Lipofectamine RNAiMAX transfection protocol for your cell line
    Catalog Number:
    13778030
    Price:
    None
    Category:
    Cell Culture Transfection Reagents
    Applications:
    Cell Culture|RNAi|RNAi Transfection|RNAi, Epigenetics & Non-Coding RNA Research|Stem Cell & Primary Cell Transfections|Synthetic siRNA Transfection|siRNA|Transfection
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    Structured Review

    Thermo Fisher lipofectamine rnaimax lipid reagent
    Lipofectamine RNAiMAX Transfection Reagent provides the highest transfection efficiencies on the widest variety of cell types for siRNA mediated gene knockdown experiments Lipofectamine RNAiMAX is a proprietary RNAi specific cationic lipid formulation designed specifically for the delivery of siRNA and miRNA into all cell types With Lipofectamine RNAiMAX Transfection Reagent you will get • Superior transfection efficiency requiring lower RNAi concentrations leading to more effective gene knockdown with minimal nonspecific effects• Easy optimization due to minimal cytotoxicity across a 10 fold concentration range of transfection reagent• Superior transfection efficiencies for miRNA antagonists and mimics• Compatibility with a broad range of cell types providing the most versatile approach to all of your gene silencing experiments• A simple and rapid protocol for consistent and reproducible resultsHigh knockdown in a wide range of cellsLipofectamine RNAiMAX Transfection Reagent transfects a wide range of cell types see figure For gene silencing Lipofectamine RNAiMAX Transfection Reagent s high efficiency transfections lead to the high levels of gene knockdown needed to achieve convincing results Simple high throughput ready transfectionsSimply mix Lipofectamine RNAiMAX Transfection Reagent with siRNA add to your cells incubate and measure gene knockdown The simplicity and speed combined with high transfection efficiency make Lipofectamine RNAiMAX Transfection Reagent ideal for high throughput siRNA transfections Transfection conditions can be easily established for automated or robotic systems used in such applications Find an optimized Lipofectamine RNAiMAX transfection protocol for your cell line
    https://www.bioz.com/result/lipofectamine rnaimax lipid reagent/product/Thermo Fisher
    Average 98 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    lipofectamine rnaimax lipid reagent - by Bioz Stars, 2021-03
    98/100 stars

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    Related Articles

    Co-Culture Assay:

    Article Title: Bone Morphogenetic Protein 9 Regulates Early Lymphatic-Specified Endothelial Cell Expansion during Mouse Embryonic Stem Cell Differentiation
    Article Snippet: The VEGF-A levels in the culture medium were measured using a mouse VEGF-A164 ELISA kit (BioTechne; Mouse VEGF Quantikine ELISA Kit, MMV00) according to the manufacturer's instructions. .. FLK-1+ vascular precursors in co-culture on OP9 cell layer since 24 hr and adherent OP9 cells were transfected with 10 nM siRNA using Lipofectamine RNAiMAX transfection reagent (Life Technologies) according to the manufacturer's instructions. .. Stimulation with BMP9 was performed 24 hr after transfection in serum-free medium.

    Transfection:

    Article Title: Bone Morphogenetic Protein 9 Regulates Early Lymphatic-Specified Endothelial Cell Expansion during Mouse Embryonic Stem Cell Differentiation
    Article Snippet: The VEGF-A levels in the culture medium were measured using a mouse VEGF-A164 ELISA kit (BioTechne; Mouse VEGF Quantikine ELISA Kit, MMV00) according to the manufacturer's instructions. .. FLK-1+ vascular precursors in co-culture on OP9 cell layer since 24 hr and adherent OP9 cells were transfected with 10 nM siRNA using Lipofectamine RNAiMAX transfection reagent (Life Technologies) according to the manufacturer's instructions. .. Stimulation with BMP9 was performed 24 hr after transfection in serum-free medium.

    Article Title: Modification of N6-methyladenosine RNA methylation on heat shock protein expression
    Article Snippet: Control siRNA is from Qiagen (1027281). .. Each siRNA was transfected into HepG2 cells using Lipofectamine RNAiMAX (Invitrogen) for siRNA following the manufacture’s protocols. .. RNA isolation and purification Total RNA was isolated from HepG2 cells using TRIzol (Invitrogen), and contaminant DNA was removed using DNaseI.

    Article Title: Atmin modulates Pkhd1 expression and may mediate Autosomal Recessive Polycystic Kidney Disease (ARPKD) through altered non-canonical Wnt/Planar Cell Polarity (PCP) signalling
    Article Snippet: For the Atmin siRNA-mediated knockdown immunocytochemistry, E-cadherin (1:500, 3195, Cell signalling) was used. .. 2.3 Cell culture and siRNA knockdowns Mouse inner medullary collecting duct (mIMCD3) cells (gift from D. Norris, MRC Harwell) were grown in DMEM/F12 (Gibco) media supplemented with 2% fetal bovine serum (Life Technologies) and penicillin-streptomycin (Life Technologies) in 1% collagen-coated 6-well plates (VWR). mIMCD3 cells were transfected with Lipofectamine RNAimax transfection reagent (Thermo Fisher), using siRNA to knock down Atmin , Pkhd1 or Pkhd1 and Atmin , according to the manufacturer's instructions (Thermo Fisher); scrambled siRNA was used as a control (Thermo Fisher). .. 2.4 Quantitative RT-PCR analysis To measure gene expression levels, RNA was extracted using the RNeasy mini kit as per manufacturer's instructions (QIAGEN).

    Article Title: ID3 regulates the MDC1-mediated DNA damage response in order to maintain genome stability
    Article Snippet: The antibodies used for immunoblotting analysis, immunoprecipitation assay and immunostaining assay are provided in Supplementary Table . .. ID3 siRNA and generation of stable ID3 knockdown cell lines For the knockdown of ID3, MDC1, or ATM expression, cells were transiently transfected with siRNA using lipofectamine RNAiMAX (Invitrogen) according to the manufacturer’s instructions. .. The siRNA target sequences were as follows: ON-TARGETplus SMARTpool ID3 siRNA (ID3 CDS, NM_002167, GE Dharmacon), 5ʹ-GCACUCAGCUUAGCCAGGU-3ʹ, 5ʹ-GAACGCAGUCUGGCCAUCG-3ʹ, 5ʹ-GGGAACUGGUACCCGGAGU-3ʹ, 5ʹ-GGAAGGUGACUUUCUGUAA-3ʹ; MDC1 siRNA (MDC1 cDNA 58-76), 5ʹ-UCCAGUGAAUCCUUGAGGUdTdT-3ʹ; ATM siRNA (ATM cDNA 1266-1284), 5ʹ-GAUACCAGAUCCUUGGAGAdTdT-3ʹ; Negative control siRNA (Bioneer), 5ʹ-CCUACGCCACCAAUUUCGUdTdT-3ʹ.

    Article Title: Cardiac Stem Cell Secretome Protects Cardiomyocytes from Hypoxic Injury Partly via Monocyte Chemotactic Protein-1-Dependent Mechanism
    Article Snippet: Densitometric quantification of blotting spots was performed using Quantity One software (Bio-Rad). .. Apoptosis Assay of HL-1 Cardiomyocytes Sca-1+/CD31− CSCshTERT were cultured on six-well plates at a density of 5 × 104 cells/well and transfected with 50 nM of MCP-1 siRNA duplexes (5′-CACAACCACCTCAAGCACT-3′) or NC siRNA (all from Bioneer) using Lipofectamine RNAiMAX (Invitrogen) for 48 h as suggested by the manufacturer. .. For CoCl2 - (Sigma-Aldrich) or H2 O2 -induced hypoxia, HL-1 cells, a cardiomyocyte cell line that continuously divides and spontaneously contracts while maintaining a differentiated cardiac phenotype [ ] was used.

    Article Title: Activation of intestinal olfactory receptor stimulates glucagon-like peptide-1 secretion in enteroendocrine cells and attenuates hyperglycemia in type 2 diabetic mice
    Article Snippet: The negative control siRNA did not affect the OR1A1 , OR1G1 , GNAL , CNGA2 , and GNAT3 expression levels in differentiated NCI-H716 cells. .. Endocrine-differentiated NCI-H716 cells were transfected with the siRNA duplexes using Lipofectamine RNAiMAX reagent (Invitrogen, Carlsbad, CA, USA) as previously described . .. Real-time quantitative PCR The expression of ORs and their downstream molecules after the siRNA transfection was determined using a StepOne real-time PCR instrument (Applied Biosystems, Foster City, CA, USA).

    Article Title: Cdc6 disruption leads to centrosome abnormalities and chromosome instability in pancreatic cancer cells
    Article Snippet: RNA interference Cdc6-specific siRNA with the following sequence: Cdc6_1: 5ʹ-AAC UUC CCA CCU UAU ACC AGA-3ʹ , Cdc6_2: 5ʹ-AAG AAU CUG CAU GUG UGA GAC-3ʹ and Cdc6_3: 5ʹ-CCA AGA AGG AGC ACA AGA U-3ʹ were synthesized by GenePharma (Shanghai, China). .. Cells were transfected with the siRNA using Lipofectamine RNAiMAX transfection reagents according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA, USA). .. Cell proliferation and colony formation assays PANC-1 cells were seeded in 12-well plates at a density of 2 × 104 cells per well.

    Article Title: The Vacuolar ATPase a2-subunit regulates Notch signaling in triple-negative breast cancer cells
    Article Snippet: Cells were transfected with siRNA (final concentration, 10 nM) using lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen) according to the manufacturer's instructions. .. Cells were transfected with siRNA (final concentration, 10 nM) using lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen) according to the manufacturer's instructions. .. For all experiments, data were collected 48 hrs post-transfection.

    Cell Culture:

    Article Title: Atmin modulates Pkhd1 expression and may mediate Autosomal Recessive Polycystic Kidney Disease (ARPKD) through altered non-canonical Wnt/Planar Cell Polarity (PCP) signalling
    Article Snippet: For the Atmin siRNA-mediated knockdown immunocytochemistry, E-cadherin (1:500, 3195, Cell signalling) was used. .. 2.3 Cell culture and siRNA knockdowns Mouse inner medullary collecting duct (mIMCD3) cells (gift from D. Norris, MRC Harwell) were grown in DMEM/F12 (Gibco) media supplemented with 2% fetal bovine serum (Life Technologies) and penicillin-streptomycin (Life Technologies) in 1% collagen-coated 6-well plates (VWR). mIMCD3 cells were transfected with Lipofectamine RNAimax transfection reagent (Thermo Fisher), using siRNA to knock down Atmin , Pkhd1 or Pkhd1 and Atmin , according to the manufacturer's instructions (Thermo Fisher); scrambled siRNA was used as a control (Thermo Fisher). .. 2.4 Quantitative RT-PCR analysis To measure gene expression levels, RNA was extracted using the RNeasy mini kit as per manufacturer's instructions (QIAGEN).

    Article Title: Cardiac Stem Cell Secretome Protects Cardiomyocytes from Hypoxic Injury Partly via Monocyte Chemotactic Protein-1-Dependent Mechanism
    Article Snippet: Densitometric quantification of blotting spots was performed using Quantity One software (Bio-Rad). .. Apoptosis Assay of HL-1 Cardiomyocytes Sca-1+/CD31− CSCshTERT were cultured on six-well plates at a density of 5 × 104 cells/well and transfected with 50 nM of MCP-1 siRNA duplexes (5′-CACAACCACCTCAAGCACT-3′) or NC siRNA (all from Bioneer) using Lipofectamine RNAiMAX (Invitrogen) for 48 h as suggested by the manufacturer. .. For CoCl2 - (Sigma-Aldrich) or H2 O2 -induced hypoxia, HL-1 cells, a cardiomyocyte cell line that continuously divides and spontaneously contracts while maintaining a differentiated cardiac phenotype [ ] was used.

    Expressing:

    Article Title: ID3 regulates the MDC1-mediated DNA damage response in order to maintain genome stability
    Article Snippet: The antibodies used for immunoblotting analysis, immunoprecipitation assay and immunostaining assay are provided in Supplementary Table . .. ID3 siRNA and generation of stable ID3 knockdown cell lines For the knockdown of ID3, MDC1, or ATM expression, cells were transiently transfected with siRNA using lipofectamine RNAiMAX (Invitrogen) according to the manufacturer’s instructions. .. The siRNA target sequences were as follows: ON-TARGETplus SMARTpool ID3 siRNA (ID3 CDS, NM_002167, GE Dharmacon), 5ʹ-GCACUCAGCUUAGCCAGGU-3ʹ, 5ʹ-GAACGCAGUCUGGCCAUCG-3ʹ, 5ʹ-GGGAACUGGUACCCGGAGU-3ʹ, 5ʹ-GGAAGGUGACUUUCUGUAA-3ʹ; MDC1 siRNA (MDC1 cDNA 58-76), 5ʹ-UCCAGUGAAUCCUUGAGGUdTdT-3ʹ; ATM siRNA (ATM cDNA 1266-1284), 5ʹ-GAUACCAGAUCCUUGGAGAdTdT-3ʹ; Negative control siRNA (Bioneer), 5ʹ-CCUACGCCACCAAUUUCGUdTdT-3ʹ.

    Apoptosis Assay:

    Article Title: Cardiac Stem Cell Secretome Protects Cardiomyocytes from Hypoxic Injury Partly via Monocyte Chemotactic Protein-1-Dependent Mechanism
    Article Snippet: Densitometric quantification of blotting spots was performed using Quantity One software (Bio-Rad). .. Apoptosis Assay of HL-1 Cardiomyocytes Sca-1+/CD31− CSCshTERT were cultured on six-well plates at a density of 5 × 104 cells/well and transfected with 50 nM of MCP-1 siRNA duplexes (5′-CACAACCACCTCAAGCACT-3′) or NC siRNA (all from Bioneer) using Lipofectamine RNAiMAX (Invitrogen) for 48 h as suggested by the manufacturer. .. For CoCl2 - (Sigma-Aldrich) or H2 O2 -induced hypoxia, HL-1 cells, a cardiomyocyte cell line that continuously divides and spontaneously contracts while maintaining a differentiated cardiac phenotype [ ] was used.

    Concentration Assay:

    Article Title: The Vacuolar ATPase a2-subunit regulates Notch signaling in triple-negative breast cancer cells
    Article Snippet: Cells were transfected with siRNA (final concentration, 10 nM) using lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen) according to the manufacturer's instructions. .. Cells were transfected with siRNA (final concentration, 10 nM) using lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen) according to the manufacturer's instructions. .. For all experiments, data were collected 48 hrs post-transfection.

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    Thermo Fisher lipid mediated transfection
    a2V inhibition increases Notch signaling in TNBC A–D. TNBC cell line MDA-MB-231 was transfected with siRNA oligonucleotides against a2V or Notch1 along with scrambled control siRNA. Cells were harvested 48 hrs after <t>transfection.</t> Fold change in mRNA expression levels of (A) Notch receptors, (B) Notch Ligands and (C) Notch target genes is shown by qRT-PCR performed on the Notch signaling PCR array. Prior to fold-change calculation, the values were normalized to signal generated from endogenous control 18srRNA. (D) Protein level of Notch1 intracellular domain (N1ICD) following a2V gene silencing is shown by western blot analysis. β actin was used as loading control. E–I. MDA-MB-231 cells were treated with Vehicle Control (DMSO), Bafilomycin A1 (Baf A1 – 0.1 or 0.5 μM) or Gamma Secretase Inhibitor (GSI – 2 μM) for 24 hrs. (E) Protein level of Notch1 intracellular domain (N1ICD) following treatment with Baf A1 or GSI is shown by western blot. β actin was used as loading control. (F) Gene expression expression levels of Hes1 relative to endogenous control 18srRNA is shown. (G) Hes1 protein expression is shown by immunofluorescence (H and I). Independently, MDA-MB-231 and MDA-MB-468 were transfected with a RBP-j Notch reporter construct and then treated with Vehicle control, 0.5 μM BafA1 or 2 μM GSI for 24 hrs. Notch reporter levels in (H) MDA-MB-231 and (I) MDA-MB-468 as measured by luciferase assay. Data represent mean ± standard error, n = 4. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 compared to control. RBPj: Recombinant Binding Protein Suppressor of Hairless.
    Lipid Mediated Transfection, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/lipid mediated transfection/product/Thermo Fisher
    Average 99 stars, based on 1 article reviews
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    lipid mediated transfection - by Bioz Stars, 2021-03
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    a2V inhibition increases Notch signaling in TNBC A–D. TNBC cell line MDA-MB-231 was transfected with siRNA oligonucleotides against a2V or Notch1 along with scrambled control siRNA. Cells were harvested 48 hrs after transfection. Fold change in mRNA expression levels of (A) Notch receptors, (B) Notch Ligands and (C) Notch target genes is shown by qRT-PCR performed on the Notch signaling PCR array. Prior to fold-change calculation, the values were normalized to signal generated from endogenous control 18srRNA. (D) Protein level of Notch1 intracellular domain (N1ICD) following a2V gene silencing is shown by western blot analysis. β actin was used as loading control. E–I. MDA-MB-231 cells were treated with Vehicle Control (DMSO), Bafilomycin A1 (Baf A1 – 0.1 or 0.5 μM) or Gamma Secretase Inhibitor (GSI – 2 μM) for 24 hrs. (E) Protein level of Notch1 intracellular domain (N1ICD) following treatment with Baf A1 or GSI is shown by western blot. β actin was used as loading control. (F) Gene expression expression levels of Hes1 relative to endogenous control 18srRNA is shown. (G) Hes1 protein expression is shown by immunofluorescence (H and I). Independently, MDA-MB-231 and MDA-MB-468 were transfected with a RBP-j Notch reporter construct and then treated with Vehicle control, 0.5 μM BafA1 or 2 μM GSI for 24 hrs. Notch reporter levels in (H) MDA-MB-231 and (I) MDA-MB-468 as measured by luciferase assay. Data represent mean ± standard error, n = 4. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 compared to control. RBPj: Recombinant Binding Protein Suppressor of Hairless.

    Journal: Oncotarget

    Article Title: The Vacuolar ATPase a2-subunit regulates Notch signaling in triple-negative breast cancer cells

    doi:

    Figure Lengend Snippet: a2V inhibition increases Notch signaling in TNBC A–D. TNBC cell line MDA-MB-231 was transfected with siRNA oligonucleotides against a2V or Notch1 along with scrambled control siRNA. Cells were harvested 48 hrs after transfection. Fold change in mRNA expression levels of (A) Notch receptors, (B) Notch Ligands and (C) Notch target genes is shown by qRT-PCR performed on the Notch signaling PCR array. Prior to fold-change calculation, the values were normalized to signal generated from endogenous control 18srRNA. (D) Protein level of Notch1 intracellular domain (N1ICD) following a2V gene silencing is shown by western blot analysis. β actin was used as loading control. E–I. MDA-MB-231 cells were treated with Vehicle Control (DMSO), Bafilomycin A1 (Baf A1 – 0.1 or 0.5 μM) or Gamma Secretase Inhibitor (GSI – 2 μM) for 24 hrs. (E) Protein level of Notch1 intracellular domain (N1ICD) following treatment with Baf A1 or GSI is shown by western blot. β actin was used as loading control. (F) Gene expression expression levels of Hes1 relative to endogenous control 18srRNA is shown. (G) Hes1 protein expression is shown by immunofluorescence (H and I). Independently, MDA-MB-231 and MDA-MB-468 were transfected with a RBP-j Notch reporter construct and then treated with Vehicle control, 0.5 μM BafA1 or 2 μM GSI for 24 hrs. Notch reporter levels in (H) MDA-MB-231 and (I) MDA-MB-468 as measured by luciferase assay. Data represent mean ± standard error, n = 4. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 compared to control. RBPj: Recombinant Binding Protein Suppressor of Hairless.

    Article Snippet: Cells were transfected with siRNA (final concentration, 10 nM) using lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen) according to the manufacturer's instructions.

    Techniques: Inhibition, Multiple Displacement Amplification, Transfection, Expressing, Quantitative RT-PCR, Polymerase Chain Reaction, Generated, Western Blot, Immunofluorescence, Construct, Luciferase, Recombinant, Binding Assay

    a2V-ATPase inhibition enhances Wnt signaling in TNBC A. MDA-MB-231 cells were transfected with scrambled control or a2V siRNA and harvested after 48 hrs of transfection. Fold change in mRNA expression levels of Wnt signaling genes WNT4, β-catenin (CTNNB1), C-MYC and Cyclin D1 (CYCD1) was assessed by qRT PCR. Prior to fold--change calculation, the values were normalized to signal generated from endogenous control 18srRNA. Data represent mean ± standard error, n = 4. * P ≤ 0.05, ** P ≤ 0.01 compared to control siRNA. (B and C) TNBC cells were grown on chamber slides and treated with B. vehicle control or C. 0.1 μM Baf A1 for 4 hours. Cells were fixed, permeabilized and processed for immunofluorescence microscopy. Localization of β-catenin (green) is shown. Nucleus was stained with DAPI (blue). Scale bars: 10 μm.

    Journal: Oncotarget

    Article Title: The Vacuolar ATPase a2-subunit regulates Notch signaling in triple-negative breast cancer cells

    doi:

    Figure Lengend Snippet: a2V-ATPase inhibition enhances Wnt signaling in TNBC A. MDA-MB-231 cells were transfected with scrambled control or a2V siRNA and harvested after 48 hrs of transfection. Fold change in mRNA expression levels of Wnt signaling genes WNT4, β-catenin (CTNNB1), C-MYC and Cyclin D1 (CYCD1) was assessed by qRT PCR. Prior to fold--change calculation, the values were normalized to signal generated from endogenous control 18srRNA. Data represent mean ± standard error, n = 4. * P ≤ 0.05, ** P ≤ 0.01 compared to control siRNA. (B and C) TNBC cells were grown on chamber slides and treated with B. vehicle control or C. 0.1 μM Baf A1 for 4 hours. Cells were fixed, permeabilized and processed for immunofluorescence microscopy. Localization of β-catenin (green) is shown. Nucleus was stained with DAPI (blue). Scale bars: 10 μm.

    Article Snippet: Cells were transfected with siRNA (final concentration, 10 nM) using lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen) according to the manufacturer's instructions.

    Techniques: Inhibition, Multiple Displacement Amplification, Transfection, Expressing, Quantitative RT-PCR, Generated, Immunofluorescence, Microscopy, Staining

    Lithium treatment stimulates NHEJ repair in a rat model of retinal I/R injury. ( a ) Location of γ -H2AX foci coincides with CREB1 phosphorylation, Nrf-1 and ligase IV in retina after following I/R surgery. The γ -H2AX-positive cells were primarily located in the INL and GCL. Scale bars: 100 μ m. ( b ) γ -H2AX expression was quantified after I/R surgery by counting foci. These data are graphically represented (12 h, 37.0±3.74; 1d, 2.66±0.6; 3d, 0.8±0.5; 7d, 0.3±0.5) compared with controls (12 h, 70.33±6.55; 1d, 11.3±0.2; 3d, 4.5±0.5; 7d, 1.8±0.9). ( c ) DNA NHEJ assay in vivo . (The strategy of the NHEJ repair assay is shown in Figure 3d .) At 24 h after I/R surgery, intravitreal injection was performed with the linearized plasmid pEGFP-N1 following transfecting reagent following the manufacturer's instructions ( in vivo -jetPEI). At 48 h after transfection, the retina is fixed and sliced. GFP is detectable in the RGC layer. Scale bars: 100 μ m. ( d ) A comparison of GFP-positive cells in the rat retina. Lithium pretreatment promotes DNA NHEJ activity (Sham, 0.67±0.62; Sham+Li, 1.25±0.72; I/R surgery, 5.5±1.19; I/R surgery+Li, 13.25±1.83). n =8 for each group, * P

    Journal: Cell Death & Disease

    Article Title: Lithium promotes DNA stability and survival of ischemic retinal neurocytes by upregulating DNA ligase IV

    doi: 10.1038/cddis.2016.341

    Figure Lengend Snippet: Lithium treatment stimulates NHEJ repair in a rat model of retinal I/R injury. ( a ) Location of γ -H2AX foci coincides with CREB1 phosphorylation, Nrf-1 and ligase IV in retina after following I/R surgery. The γ -H2AX-positive cells were primarily located in the INL and GCL. Scale bars: 100 μ m. ( b ) γ -H2AX expression was quantified after I/R surgery by counting foci. These data are graphically represented (12 h, 37.0±3.74; 1d, 2.66±0.6; 3d, 0.8±0.5; 7d, 0.3±0.5) compared with controls (12 h, 70.33±6.55; 1d, 11.3±0.2; 3d, 4.5±0.5; 7d, 1.8±0.9). ( c ) DNA NHEJ assay in vivo . (The strategy of the NHEJ repair assay is shown in Figure 3d .) At 24 h after I/R surgery, intravitreal injection was performed with the linearized plasmid pEGFP-N1 following transfecting reagent following the manufacturer's instructions ( in vivo -jetPEI). At 48 h after transfection, the retina is fixed and sliced. GFP is detectable in the RGC layer. Scale bars: 100 μ m. ( d ) A comparison of GFP-positive cells in the rat retina. Lithium pretreatment promotes DNA NHEJ activity (Sham, 0.67±0.62; Sham+Li, 1.25±0.72; I/R surgery, 5.5±1.19; I/R surgery+Li, 13.25±1.83). n =8 for each group, * P

    Article Snippet: The retinal neurocytes (4 × 106 cells per well in 6-well culture plates) in good condition underwent lipid-mediated transfection with Lipofectamine RNAiMAX (Invitrogen), as recommended by the manufacturer.

    Techniques: Non-Homologous End Joining, Expressing, In Vivo, Injection, Plasmid Preparation, Transfection, Activity Assay

    TRMT10A knockdown enhances total protein biosynthesis in rat β-cells. Total protein synthesis was measured in INS-1E cells transfected with control siRNA (siCT) or two siRNAs targeting rat TRMT10A (siTRMT10A #1 and #2). Protein biosynthesis was corrected by total protein content and expressed as % of siCT. INS-1E cells treated for 2 h with the inhibitor of translation cycloheximide (CY, 10 µM) were used as positive control (n = 4). *CY vs CT, § siTRMT10A vs siCT p

    Journal: PLoS Genetics

    Article Title: tRNA Methyltransferase Homolog Gene TRMT10A Mutation in Young Onset Diabetes and Primary Microcephaly in Humans

    doi: 10.1371/journal.pgen.1003888

    Figure Lengend Snippet: TRMT10A knockdown enhances total protein biosynthesis in rat β-cells. Total protein synthesis was measured in INS-1E cells transfected with control siRNA (siCT) or two siRNAs targeting rat TRMT10A (siTRMT10A #1 and #2). Protein biosynthesis was corrected by total protein content and expressed as % of siCT. INS-1E cells treated for 2 h with the inhibitor of translation cycloheximide (CY, 10 µM) were used as positive control (n = 4). *CY vs CT, § siTRMT10A vs siCT p

    Article Snippet: RNA interference and plasmid transfections Cells were transfected overnight with 30 nM of a control siRNA (Qiagen), or two single siRNAs targeting rat or human TRMT10A using Lipofectamine RNAiMAX (Invitrogen). siRNA-lipid complexes were formed in Opti-mem (Invitrogen) for 20 min as previously described . hrGFP and TRMT10A-hrGFP plasmids were introduced by lipofection in INS-1E cells or dispersed rat and human islet cells using Lipofectamine 2000 (Invitrogen). siRNA sequences, plasmid and Lipofectamine concentrations are described in and .

    Techniques: Transfection, Positive Control

    TRMT10A knockdown sensitizes β-cells to FFA-, high glucose- and ER stress-induced apoptosis. INS-1E cells (A–E), primary rat β-cells (F) and dispersed human islets (G) were transfected with control siRNA (siCT) or siRNAs targeting rat (siTRMT10A #1 and #2) or human TRMT10A (siTRMT10A #3 and #4). 48 h after transfection cells were exposed or not (CT) to oleate (OL), palmitate (PAL) and 28 mM glucose (G28), or to the chemical ER stressors cyclopiazonic acid (CPA), tunicamycin (TU) and brefeldin (BR), for 24 (A–B) or 16 h (C–E). Apoptosis was examined by propidium iodide and Hoechst 33342 staining (A–D, F–G) or Western blot for cleaved caspase-3 (E). Results are means ± SE (n = 3–5). The blots are representative of 4 independent experiments. * Treated vs CT; § siTRMT10A vs siCT. One symbol p

    Journal: PLoS Genetics

    Article Title: tRNA Methyltransferase Homolog Gene TRMT10A Mutation in Young Onset Diabetes and Primary Microcephaly in Humans

    doi: 10.1371/journal.pgen.1003888

    Figure Lengend Snippet: TRMT10A knockdown sensitizes β-cells to FFA-, high glucose- and ER stress-induced apoptosis. INS-1E cells (A–E), primary rat β-cells (F) and dispersed human islets (G) were transfected with control siRNA (siCT) or siRNAs targeting rat (siTRMT10A #1 and #2) or human TRMT10A (siTRMT10A #3 and #4). 48 h after transfection cells were exposed or not (CT) to oleate (OL), palmitate (PAL) and 28 mM glucose (G28), or to the chemical ER stressors cyclopiazonic acid (CPA), tunicamycin (TU) and brefeldin (BR), for 24 (A–B) or 16 h (C–E). Apoptosis was examined by propidium iodide and Hoechst 33342 staining (A–D, F–G) or Western blot for cleaved caspase-3 (E). Results are means ± SE (n = 3–5). The blots are representative of 4 independent experiments. * Treated vs CT; § siTRMT10A vs siCT. One symbol p

    Article Snippet: RNA interference and plasmid transfections Cells were transfected overnight with 30 nM of a control siRNA (Qiagen), or two single siRNAs targeting rat or human TRMT10A using Lipofectamine RNAiMAX (Invitrogen). siRNA-lipid complexes were formed in Opti-mem (Invitrogen) for 20 min as previously described . hrGFP and TRMT10A-hrGFP plasmids were introduced by lipofection in INS-1E cells or dispersed rat and human islet cells using Lipofectamine 2000 (Invitrogen). siRNA sequences, plasmid and Lipofectamine concentrations are described in and .

    Techniques: Transfection, Staining, Western Blot