allele-specific binding Search Results


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  • 99
    ATCC atcc 10231 derived allele
    Atcc 10231 Derived Allele, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore aso binding proteins streptavidin magnetic beads
    Aso Binding Proteins Streptavidin Magnetic Beads, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Thermo Fisher allele specific minor groove binding probes
    Allele Specific Minor Groove Binding Probes, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Genomatix matinspector software predicted allele specific binding
    Schematic presentation of location of analyzed polymorphisms in the human FADS2 gene promoter and surrounding TFBSs modified from Genomatix <t>MatInspector</t> software output. A: A BCL6 binding site is predicted when the major T allele of rs3834458 is present but lacking when the deletion is present. B: The minor T allele of rs968567 leads to prediction of binding sites for ELK1, STAT1, and STAT3. In contrast, no binding sites are predicted when the major C allele is present. C and D: Sequence that was recognized by Genomatix MatInspector to contain a TFBS and also rs3834458 (C) or rs968567 (D), respectively. Nucleotides marked in bold red show a high degree of matrix conservation at this position, i.e., information content is very high. Nucleotides in capitals denote the core sequence used by MatInspector. Underlined nucleotides highlight the position of the respective polymorphism; rc means that a TFBS sequence was found for the opposite strand.
    Matinspector Software Predicted Allele Specific Binding, supplied by Genomatix, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher allele specific taqman minor groove binding
    Schematic presentation of location of analyzed polymorphisms in the human FADS2 gene promoter and surrounding TFBSs modified from Genomatix <t>MatInspector</t> software output. A: A BCL6 binding site is predicted when the major T allele of rs3834458 is present but lacking when the deletion is present. B: The minor T allele of rs968567 leads to prediction of binding sites for ELK1, STAT1, and STAT3. In contrast, no binding sites are predicted when the major C allele is present. C and D: Sequence that was recognized by Genomatix MatInspector to contain a TFBS and also rs3834458 (C) or rs968567 (D), respectively. Nucleotides marked in bold red show a high degree of matrix conservation at this position, i.e., information content is very high. Nucleotides in capitals denote the core sequence used by MatInspector. Underlined nucleotides highlight the position of the respective polymorphism; rc means that a TFBS sequence was found for the opposite strand.
    Allele Specific Taqman Minor Groove Binding, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bio-Rad allele specific polymerase chain reaction
    Schematic presentation of location of analyzed polymorphisms in the human FADS2 gene promoter and surrounding TFBSs modified from Genomatix <t>MatInspector</t> software output. A: A BCL6 binding site is predicted when the major T allele of rs3834458 is present but lacking when the deletion is present. B: The minor T allele of rs968567 leads to prediction of binding sites for ELK1, STAT1, and STAT3. In contrast, no binding sites are predicted when the major C allele is present. C and D: Sequence that was recognized by Genomatix MatInspector to contain a TFBS and also rs3834458 (C) or rs968567 (D), respectively. Nucleotides marked in bold red show a high degree of matrix conservation at this position, i.e., information content is very high. Nucleotides in capitals denote the core sequence used by MatInspector. Underlined nucleotides highlight the position of the respective polymorphism; rc means that a TFBS sequence was found for the opposite strand.
    Allele Specific Polymerase Chain Reaction, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 88/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Becton Dickinson hla alleles
    Hydrogen bond interactions. The percentage occupancy of H-bonds averaged over the last 25 ns of simulation time between the nine residues (P1–P9) of the <t>MICA-TM</t> peptide and the <t>HLA</t> residues for the four complexes.
    Hla Alleles, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 91/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Romer Labs pepper bs3 alleles
    Hydrogen bond interactions. The percentage occupancy of H-bonds averaged over the last 25 ns of simulation time between the nine residues (P1–P9) of the <t>MICA-TM</t> peptide and the <t>HLA</t> residues for the four complexes.
    Pepper Bs3 Alleles, supplied by Romer Labs, used in various techniques. Bioz Stars score: 88/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc antibodies against pd l1
    A negative correlation between lncMX1–215 and <t>PD-L1</t> and galectin-9 expression was observed in HNSCC tissues. a Representative images of lncMX1–215 expression in normal and HNSCC tissues shown using the RNAscope technique. b The expression scores of lncMX1–215 in normal and HNSCC tissues were compared. c-f, Correlations between lncMX1–215 expression and TNM stage, pathological grade, gender and age were analyzed. g Representative images of PD-L1 and lncMX1–215 expression were shown. h The correlation between PD-L1 and lncMX1–215 expression was analyzed in an HNSCC TMA. i Representative images of galectin-9 and lncMX1–215 expression were shown. j The correlation between galectin-9 and lncMX1–215 expression was analyzed in an HNSCC TMA. The red triangle indicates an lncMX1–215 molecule. *: P
    Antibodies Against Pd L1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 142 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Ribobio lncmx1 215 specific aso
    <t>LncMX1–215</t> inhibits tumor proliferation capacity in vitro and in vivo . a, b The viability of HN4 and Cal27 cells after transfection with lncMX1–215 or <t>ASO</t> was determined using CCK8 assays. c, d Colony formation assays were performed with HN4 and Cal27 cells after transfection with lncMX1–215 or ASO. e, f EdU assays were performed after transfection with lncMX1–215 or ASO. g PARP and cleaved caspase 3 were detected via western blotting after transfection with lncMX1–215 for 48 h. h Cell apoptosis was analyzed via flow cytometry using an Annexin V/7-AAD kit after transfection for 48 h. i Cell cycle was analyzed using flow cytometry after transfection for 48 h. j Tumor growth curves for mice injected with cells treated with lncMX1–215 lentivirus or vector were analyzed. k Tumors derived from the xenograft model were resected and are shown for each group. l The weight of tumors resected from mice in the ectopic expression and vector groups was measured and analyzed. m The percentage of TUNEL-positive cells in each group was compared. n The relative Ki-67 staining score in each group was analyzed. *: P
    Lncmx1 215 Specific Aso, supplied by Ribobio, used in various techniques. Bioz Stars score: 94/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc metap2 specific antibody
    ASO‐21 does not affect splicing or expression of genomic targets with one mismatch ASO‐21 has a single mismatched nucleotide with three other sites in the mouse genome. The genes associated with these potential off‐target binding sites are shown. The location of the site, intronic length, position relative to nearby landmarks, the specific mismatch, and contiguous base pairs in the putative duplex between ASO‐21 and the off‐target site are shown. RT–PCR analysis of RNA isolated from the cortex of AD mice treated with either ASO‐C or ASO‐21, as in Fig 4 . The specific region of the transcript that is predicted to be affected by ASO‐21 binding was amplified, and products were separated on a 2% agarose gel. No aberrant splicing or changes in alternative splicing patterns were observed. Immunoblot analysis of <t>MetAP2</t> protein from the cortex of AD mice treated with either ASO‐C or ASO‐21, as in Fig 4 . ASO‐21 could form a putative duplex upstream of the MetAP2 start codon, potentially affecting expression. Graph depicting the quantitation of MetAP2 protein expression depicted in (C). Bars represent mean ± s.e.m. Source data are available online for this figure.
    Metap2 Specific Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore magna riptm rna binding protein immunoprecipitation kit
    Uniform 2′-MOE ASOs cause intron retention and decreased levels of STAT3 mRNA in association with the chromatin. ( A ) Purity of the cell fractions was assessed by qRT-PCR for a chromatin <t>RNA</t> marker ( STAT3 intron 21) and a cytoplasmic RNA marker ( RN7SL1 ) as well as by western blotting for proteins enriched in the indicated compartments. ( B ) HeLa cells were transfected with uniform 2′-MOE (UNI6a and UNI6b) or 2′-MOE gapmer ASO (GAP1i) for 24 h, followed by cell fractionation. STAT3 expression in each fraction was analyzed by qRT-PCR of the indicated regions of STAT3 mRNA. Expression of improperly spliced mRNA was visualized by polyacrylamide separation of STAT3 RT-PCR products. ( C ) STAT3 pre-mRNA levels were analyzed in the chromatin fraction by qRT-PCR. ( D ) Following RNAPII <t>immunoprecipitation,</t> successful pull-down was shown by western blot (right panel) and RNAPII-bound STAT3 was analyzed by qRT-PCR. Mean ± absolute deviation ( n = 2); * P
    Magna Riptm Rna Binding Protein Immunoprecipitation Kit, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 344 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    FFA Sciences water soluble acrylodan labeled fa binding protein
    Uniform 2′-MOE ASOs cause intron retention and decreased levels of STAT3 mRNA in association with the chromatin. ( A ) Purity of the cell fractions was assessed by qRT-PCR for a chromatin <t>RNA</t> marker ( STAT3 intron 21) and a cytoplasmic RNA marker ( RN7SL1 ) as well as by western blotting for proteins enriched in the indicated compartments. ( B ) HeLa cells were transfected with uniform 2′-MOE (UNI6a and UNI6b) or 2′-MOE gapmer ASO (GAP1i) for 24 h, followed by cell fractionation. STAT3 expression in each fraction was analyzed by qRT-PCR of the indicated regions of STAT3 mRNA. Expression of improperly spliced mRNA was visualized by polyacrylamide separation of STAT3 RT-PCR products. ( C ) STAT3 pre-mRNA levels were analyzed in the chromatin fraction by qRT-PCR. ( D ) Following RNAPII <t>immunoprecipitation,</t> successful pull-down was shown by western blot (right panel) and RNAPII-bound STAT3 was analyzed by qRT-PCR. Mean ± absolute deviation ( n = 2); * P
    Water Soluble Acrylodan Labeled Fa Binding Protein, supplied by FFA Sciences, used in various techniques. Bioz Stars score: 86/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Nordic-Mubio tritc goat anti mouse igg
    Uniform 2′-MOE ASOs cause intron retention and decreased levels of STAT3 mRNA in association with the chromatin. ( A ) Purity of the cell fractions was assessed by qRT-PCR for a chromatin <t>RNA</t> marker ( STAT3 intron 21) and a cytoplasmic RNA marker ( RN7SL1 ) as well as by western blotting for proteins enriched in the indicated compartments. ( B ) HeLa cells were transfected with uniform 2′-MOE (UNI6a and UNI6b) or 2′-MOE gapmer ASO (GAP1i) for 24 h, followed by cell fractionation. STAT3 expression in each fraction was analyzed by qRT-PCR of the indicated regions of STAT3 mRNA. Expression of improperly spliced mRNA was visualized by polyacrylamide separation of STAT3 RT-PCR products. ( C ) STAT3 pre-mRNA levels were analyzed in the chromatin fraction by qRT-PCR. ( D ) Following RNAPII <t>immunoprecipitation,</t> successful pull-down was shown by western blot (right panel) and RNAPII-bound STAT3 was analyzed by qRT-PCR. Mean ± absolute deviation ( n = 2); * P
    Tritc Goat Anti Mouse Igg, supplied by Nordic-Mubio, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti fus
    P54nrb preferentially associates with 2′-F-modified oligonucleotides. ( A ) Oligonucleotides used in this experiment. ( B ) ASO-binding proteins were isolated from HeLa cell extracts using a biotinylated gapmer 2′-F-PS-ASO (ISIS623496). Proteins were eluted from beads by competition with indicated gapmers. The affinity-selected proteins were analyzed by western blot. Ku70 served as a loading control. ( C ) Western analysis following ASO-pull down as described in panel (A) indicated that P54nrb has higher affinity for oligonucleotides with the 2′-F modification on the 3′ side. Ku70 served as a loading control. ( D ) Transfection of 2′-F-oligonucleotides reduced P54nrb but not <t>FUS</t> protein levels. HeLa cells were transfected with 2′-F-PS-ASO (ISIS404130) at a final concentration of 30 nM, and protein levels were determined by western analysis. ( E ) Western analysis of proteins isolated from HeLa cells following treatment of cells with siRNAs targeting P54nrb or <t>PSPC1</t> mRNA. Cells were transfected with siRNA at 3-nM final concentration and harvested after 36 h.
    Anti Fus, supplied by Abcam, used in various techniques. Bioz Stars score: 95/100, based on 75 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti pspc1
    <t>PSPC1</t> levels are reduced by 2′-F-modified oligonucleotides. ( A ) Indicated oligonucleotides were transfected into HeLa cells at a final concentration of 30 nM. After 24 h, levels of PSPC1 were evaluated by western. GAPDH served as a loading control. ( B ) A431 cells were incubated with ISIS404130 by free uptake for 40 h at the indicated concentrations. Levels of P54nrb, PSF and PSF proteins were evaluated by western. GAPDH served as a loading control.
    Anti Pspc1, supplied by Abcam, used in various techniques. Bioz Stars score: 93/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam phosphorylated gsk 3β
    MiR-195 blocked Wnt/β-catenin pathway. SW480 cells were transfected with miR-195 mimic, ASO of miR-195 or their corresponding controls. Then, the protein expressions of (A) β-catenin and Ki-67s, as well as (B) <t>GSK-3β</t> and p-GSK-3β
    Phosphorylated Gsk 3β, supplied by Abcam, used in various techniques. Bioz Stars score: 95/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore transwell assay cell migration
    Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using <t>Transwell</t> inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P
    Transwell Assay Cell Migration, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti nrf2
    NRAL controls the target of miR-340-5p. (a) Schematic diagram of the miR-340-5p binding site in the <t>Nrf2</t> 3’UTR or its mutant sequences based on TargetScan software. (b) Relative luciferase intensities of the luciferase constructors harbouring the wild-type or mutant Nrf2 3’UTRs were measured in 293T cells 48 h after transfection with the pri-miR-340-5p plasmid or ASO-miR-340-5p, n=3, ** p
    Anti Nrf2, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 820 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore active jnk1
    Effect of knocking down <t>JNK1</t> and/or -2 on JNK translocation to mitochondria, mitochondria bioenergetics, and liver injury. A , effect of JNK1 and -2 ASO on JNK translocation to mitochondria induced by APAP treatment. B , effect of silencing JNK1 and -2
    Active Jnk1, supplied by Millipore, used in various techniques. Bioz Stars score: 85/100, based on 20 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher oligreen
    ( A ) <t>Oligreen</t> dye-exclusion assays to determine bolasome binding capacity using 2 µg gapmer ASO. ( B ) Bolasome complexation and retention of phosphorothioate antisense gapmers following metaphor electrophoresis and SYBR Green II staining. Lanes 1–4: Amine-to-phosphate (N/P) ratio =0, 0.5, 2.5, 5.0. Abbreviation: ASO, antisense oligonucleotides.
    Oligreen, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 89/100, based on 424 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Isis Pharmaceuticals antisense oligonucleotide
    ( A ) <t>Oligreen</t> dye-exclusion assays to determine bolasome binding capacity using 2 µg gapmer ASO. ( B ) Bolasome complexation and retention of phosphorothioate antisense gapmers following metaphor electrophoresis and SYBR Green II staining. Lanes 1–4: Amine-to-phosphate (N/P) ratio =0, 0.5, 2.5, 5.0. Abbreviation: ASO, antisense oligonucleotides.
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    Abcam anti akt1
    miR-153 negatively regulates KLF5 expression in LSCC. (A and B) Western blot analysis of <t>AKT1</t> and FOXO3a expression in HEp-2 cells transfected with miR-153 mimics, ASO or NC. (C) Informatics prediction of miR-153 binding sites in the 3′-UTRs of
    Anti Akt1, supplied by Abcam, used in various techniques. Bioz Stars score: 96/100, based on 120 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore p irs1
    Effects of miR-494 on the insulin signaling pathway. The mimic form of miR-494 was overexpressed in cells. Cells were starved for 4 h before treatment with 0, 10, 100 nM of insulin for 20 min, and the levels of p-Akt (Ser473 and Thr308), p-AS160, p-p70S6K, and p-GSK-3α/β were measured in C 2 C 12 myoblasts (A) and CHO IR/IRS-1 cells (D). Bars show densitometric quantitation of phosphorylations of Akt, GSK-3α/β, AS160 and p70S6K in C 2 C 12 myoblasts (B) and CHO IR/IRS-1 cells (E). (C) The level of <t>p-IRS1</t> (Tyr608) was measured in insulin-treated C 2 C 12 myoblasts. GAPDH and actin were used as loading controls. Levels of phospho-proteins between miR-494 transfected cells and control were compared at each concentration of insulin. The values are expressed as the means ± SEM. *P
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    Image Search Results


    Schematic presentation of location of analyzed polymorphisms in the human FADS2 gene promoter and surrounding TFBSs modified from Genomatix MatInspector software output. A: A BCL6 binding site is predicted when the major T allele of rs3834458 is present but lacking when the deletion is present. B: The minor T allele of rs968567 leads to prediction of binding sites for ELK1, STAT1, and STAT3. In contrast, no binding sites are predicted when the major C allele is present. C and D: Sequence that was recognized by Genomatix MatInspector to contain a TFBS and also rs3834458 (C) or rs968567 (D), respectively. Nucleotides marked in bold red show a high degree of matrix conservation at this position, i.e., information content is very high. Nucleotides in capitals denote the core sequence used by MatInspector. Underlined nucleotides highlight the position of the respective polymorphism; rc means that a TFBS sequence was found for the opposite strand.

    Journal: Journal of Lipid Research

    Article Title: A common FADS2 promoter polymorphism increases promoter activity and facilitates binding of transcription factor ELK1

    doi: 10.1194/jlr.M900289-JLR200

    Figure Lengend Snippet: Schematic presentation of location of analyzed polymorphisms in the human FADS2 gene promoter and surrounding TFBSs modified from Genomatix MatInspector software output. A: A BCL6 binding site is predicted when the major T allele of rs3834458 is present but lacking when the deletion is present. B: The minor T allele of rs968567 leads to prediction of binding sites for ELK1, STAT1, and STAT3. In contrast, no binding sites are predicted when the major C allele is present. C and D: Sequence that was recognized by Genomatix MatInspector to contain a TFBS and also rs3834458 (C) or rs968567 (D), respectively. Nucleotides marked in bold red show a high degree of matrix conservation at this position, i.e., information content is very high. Nucleotides in capitals denote the core sequence used by MatInspector. Underlined nucleotides highlight the position of the respective polymorphism; rc means that a TFBS sequence was found for the opposite strand.

    Article Snippet: Additionally, the Genomatix MatInspector software predicted allele-specific binding of ELK1 to the region surrounding SNP rs968567.

    Techniques: Modification, Software, Binding Assay, Sequencing

    Hydrogen bond interactions. The percentage occupancy of H-bonds averaged over the last 25 ns of simulation time between the nine residues (P1–P9) of the MICA-TM peptide and the HLA residues for the four complexes.

    Journal: PLoS ONE

    Article Title: Molecular Dynamics Simulation Reveals the Selective Binding of Human Leukocyte Antigen Alleles Associated with Behçet's Disease

    doi: 10.1371/journal.pone.0135575

    Figure Lengend Snippet: Hydrogen bond interactions. The percentage occupancy of H-bonds averaged over the last 25 ns of simulation time between the nine residues (P1–P9) of the MICA-TM peptide and the HLA residues for the four complexes.

    Article Snippet: Therefore, we aimed to investigate the link between BD and two specific HLA alleles associated with BD (HLA-A*26:01 and HLA-B*51:01) in terms of their binding affinity to the MICA-TM peptide using MD simulations, in comparison with two HLA alleles not associated with BD (HLA-A*11:01 and HLA-B*35:01).

    Techniques:

    Averaged decomposition energy contributions in HLA binding to MICA-TM. Per-residue decomposition energies and the energy components in terms of the electrostatic (ele) and van der Waals (vdW) interactions for the P1–P9 residues of MICA-TM.

    Journal: PLoS ONE

    Article Title: Molecular Dynamics Simulation Reveals the Selective Binding of Human Leukocyte Antigen Alleles Associated with Behçet's Disease

    doi: 10.1371/journal.pone.0135575

    Figure Lengend Snippet: Averaged decomposition energy contributions in HLA binding to MICA-TM. Per-residue decomposition energies and the energy components in terms of the electrostatic (ele) and van der Waals (vdW) interactions for the P1–P9 residues of MICA-TM.

    Article Snippet: Therefore, we aimed to investigate the link between BD and two specific HLA alleles associated with BD (HLA-A*26:01 and HLA-B*51:01) in terms of their binding affinity to the MICA-TM peptide using MD simulations, in comparison with two HLA alleles not associated with BD (HLA-A*11:01 and HLA-B*35:01).

    Techniques: Binding Assay

    Structural flexibilities of the HLA alleles bound with the MICA-TM peptide. Structural flexibilities were evaluated by B-factor. The ribbon color changes from blue (rigid) to red (flexible) to represent a low to high protein flexibility. Note that for clarity only the binding groove structure and the MICA-TM peptide are shown.

    Journal: PLoS ONE

    Article Title: Molecular Dynamics Simulation Reveals the Selective Binding of Human Leukocyte Antigen Alleles Associated with Behçet's Disease

    doi: 10.1371/journal.pone.0135575

    Figure Lengend Snippet: Structural flexibilities of the HLA alleles bound with the MICA-TM peptide. Structural flexibilities were evaluated by B-factor. The ribbon color changes from blue (rigid) to red (flexible) to represent a low to high protein flexibility. Note that for clarity only the binding groove structure and the MICA-TM peptide are shown.

    Article Snippet: Therefore, we aimed to investigate the link between BD and two specific HLA alleles associated with BD (HLA-A*26:01 and HLA-B*51:01) in terms of their binding affinity to the MICA-TM peptide using MD simulations, in comparison with two HLA alleles not associated with BD (HLA-A*11:01 and HLA-B*35:01).

    Techniques: Binding Assay

    Structural basis of HLA class I. (A) Schematic model of HLA buried in the transmembrane. (B) HLA (pink) contains the α 1 and α 2 subdomains that contribute to the peptide binding groove, while α 3 is the C-terminal domain in complex with ß 2 -microgluobulin ( ß 2 m) as a noncovalently supported protein (cyan). (C) Ribbon and (D) van der Waals surface representations of the MICA-TM nonapeptide (green stick model) occupied in the peptide binding sub-sites (S1–S9, shaded by different colors) of HLA-B*51:01.

    Journal: PLoS ONE

    Article Title: Molecular Dynamics Simulation Reveals the Selective Binding of Human Leukocyte Antigen Alleles Associated with Behçet's Disease

    doi: 10.1371/journal.pone.0135575

    Figure Lengend Snippet: Structural basis of HLA class I. (A) Schematic model of HLA buried in the transmembrane. (B) HLA (pink) contains the α 1 and α 2 subdomains that contribute to the peptide binding groove, while α 3 is the C-terminal domain in complex with ß 2 -microgluobulin ( ß 2 m) as a noncovalently supported protein (cyan). (C) Ribbon and (D) van der Waals surface representations of the MICA-TM nonapeptide (green stick model) occupied in the peptide binding sub-sites (S1–S9, shaded by different colors) of HLA-B*51:01.

    Article Snippet: Therefore, we aimed to investigate the link between BD and two specific HLA alleles associated with BD (HLA-A*26:01 and HLA-B*51:01) in terms of their binding affinity to the MICA-TM peptide using MD simulations, in comparison with two HLA alleles not associated with BD (HLA-A*11:01 and HLA-B*35:01).

    Techniques: Binding Assay

    Decomposition energy per HLA residue fingerprint plots. The HLA contribution to the MICA-TM binding is shown in terms of the electrostatic (ele) and van der Waals (vdW) interactions.

    Journal: PLoS ONE

    Article Title: Molecular Dynamics Simulation Reveals the Selective Binding of Human Leukocyte Antigen Alleles Associated with Behçet's Disease

    doi: 10.1371/journal.pone.0135575

    Figure Lengend Snippet: Decomposition energy per HLA residue fingerprint plots. The HLA contribution to the MICA-TM binding is shown in terms of the electrostatic (ele) and van der Waals (vdW) interactions.

    Article Snippet: Therefore, we aimed to investigate the link between BD and two specific HLA alleles associated with BD (HLA-A*26:01 and HLA-B*51:01) in terms of their binding affinity to the MICA-TM peptide using MD simulations, in comparison with two HLA alleles not associated with BD (HLA-A*11:01 and HLA-B*35:01).

    Techniques: Binding Assay

    A negative correlation between lncMX1–215 and PD-L1 and galectin-9 expression was observed in HNSCC tissues. a Representative images of lncMX1–215 expression in normal and HNSCC tissues shown using the RNAscope technique. b The expression scores of lncMX1–215 in normal and HNSCC tissues were compared. c-f, Correlations between lncMX1–215 expression and TNM stage, pathological grade, gender and age were analyzed. g Representative images of PD-L1 and lncMX1–215 expression were shown. h The correlation between PD-L1 and lncMX1–215 expression was analyzed in an HNSCC TMA. i Representative images of galectin-9 and lncMX1–215 expression were shown. j The correlation between galectin-9 and lncMX1–215 expression was analyzed in an HNSCC TMA. The red triangle indicates an lncMX1–215 molecule. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: A negative correlation between lncMX1–215 and PD-L1 and galectin-9 expression was observed in HNSCC tissues. a Representative images of lncMX1–215 expression in normal and HNSCC tissues shown using the RNAscope technique. b The expression scores of lncMX1–215 in normal and HNSCC tissues were compared. c-f, Correlations between lncMX1–215 expression and TNM stage, pathological grade, gender and age were analyzed. g Representative images of PD-L1 and lncMX1–215 expression were shown. h The correlation between PD-L1 and lncMX1–215 expression was analyzed in an HNSCC TMA. i Representative images of galectin-9 and lncMX1–215 expression were shown. j The correlation between galectin-9 and lncMX1–215 expression was analyzed in an HNSCC TMA. The red triangle indicates an lncMX1–215 molecule. *: P

    Article Snippet: Immunohistochemistry (IHC) and immunofluorescence were performed as described in our previous study [ ], with primary antibodies against PD-L1, galectin-9, H3K27ac and GCN5 (Cell Signaling Technology, Danvers, MA, USA), and Ki-67 (Proteintech, Rocky Hill, NJ, USA).

    Techniques: Expressing

    Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using Transwell inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using Transwell inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P

    Article Snippet: Immunohistochemistry (IHC) and immunofluorescence were performed as described in our previous study [ ], with primary antibodies against PD-L1, galectin-9, H3K27ac and GCN5 (Cell Signaling Technology, Danvers, MA, USA), and Ki-67 (Proteintech, Rocky Hill, NJ, USA).

    Techniques: Over Expression, Migration, Transfection, Allele-specific Oligonucleotide, Mouse Assay, Staining, Expressing, Binding Assay

    LncMX1–215 negatively regulates IFNα-induced PD-L1 and galectin-9 expression. a lncMX1–215 expression was detected in HN4 and Cal27 cells via PCR after transfection with expression plasmids for 24 h. b, c The protein and mRNA of PD-L1 and galectin-9 expression levels were detected after transfection with lncMX1–215 for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. d The lncMX1–215 knockdown efficiency after transfection with ASO for 24 h was analyzed. e, f After ASO transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h, PD-L1 and galectin-9 expression levels were detected using western blotting and qRT-PCR. g Surface PD-L1 expression was examined using flow cytometry after transfection with lncMX1–215 followed by 200 ng/ml IFNα stimulation for 24 h. h The galectin-9 concentration in medium supernatant was measured via ELISA after transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. i Galectin-9 knockdown efficiency was detected via western blotting after transfection with siRNA for 48 h. j Transfected cells were seeded in a 96-well plate and incubated with NK cells for 4 h at various effector/target (E:T) cell ratios as indicated. The specific lysis rate was measured using an LDH kit. k Relative CD274 mRNA and lncMX1–215 expression were detected using qRT-PCR after 200 ng/ml IFNα stimulation for indicated time. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: LncMX1–215 negatively regulates IFNα-induced PD-L1 and galectin-9 expression. a lncMX1–215 expression was detected in HN4 and Cal27 cells via PCR after transfection with expression plasmids for 24 h. b, c The protein and mRNA of PD-L1 and galectin-9 expression levels were detected after transfection with lncMX1–215 for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. d The lncMX1–215 knockdown efficiency after transfection with ASO for 24 h was analyzed. e, f After ASO transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h, PD-L1 and galectin-9 expression levels were detected using western blotting and qRT-PCR. g Surface PD-L1 expression was examined using flow cytometry after transfection with lncMX1–215 followed by 200 ng/ml IFNα stimulation for 24 h. h The galectin-9 concentration in medium supernatant was measured via ELISA after transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. i Galectin-9 knockdown efficiency was detected via western blotting after transfection with siRNA for 48 h. j Transfected cells were seeded in a 96-well plate and incubated with NK cells for 4 h at various effector/target (E:T) cell ratios as indicated. The specific lysis rate was measured using an LDH kit. k Relative CD274 mRNA and lncMX1–215 expression were detected using qRT-PCR after 200 ng/ml IFNα stimulation for indicated time. *: P

    Article Snippet: Immunohistochemistry (IHC) and immunofluorescence were performed as described in our previous study [ ], with primary antibodies against PD-L1, galectin-9, H3K27ac and GCN5 (Cell Signaling Technology, Danvers, MA, USA), and Ki-67 (Proteintech, Rocky Hill, NJ, USA).

    Techniques: Expressing, Polymerase Chain Reaction, Transfection, Allele-specific Oligonucleotide, Western Blot, Quantitative RT-PCR, Flow Cytometry, Cytometry, Concentration Assay, Enzyme-linked Immunosorbent Assay, Incubation, Lysis

    LncMX1–215 directly interacts with GCN5. a GCN5 and H3K27ac were detected after transfection with lncMX1–215 for 24 h and treatment with 200 ng/ml IFNα for 24 h. b GCN5 and H3K27ac were detected after transfection with ASO for 48 h. c, d GCN5, PD-L1, galectin-9 and H3K27ac were examined after transfection with lncMX1–215 or ASO for 24 h followed by treatment with 5 μM MS-275 for 24 h. e GCN5, PD-L1, galectin-9 and H3K27ac were detected after ectopic expression of GCN5 for 48 h in HN4 and Cal27 cells. f PD-L1 and galectin-9 were detected after cotransfection with GCN5 and lncMX1–215 for 48 h. g After transfection with lncMX1–215 for 48 h, cell lysates were precipitated with anti-GCN5 or IgG antibody. h GCN5 and H3K27ac expression was analyzed and quantified using immunofluorescence. i The correlation between H3K27ac and GCN5 and lncMX1–215 expression was analyzed in HNSCC tissue microarray. j Overexpression of GCN5-DYKDDDDK fusion plasmid in HN4 cells was verified by western blotting. P1 to P5 were truncations for full length of lncMX1–215 amplified by specific primers. k After transfection of HN4 cells with GCN5-DYKDDDDK fusion plasmid for 48 h, RNA immunoprecipitation was used to detect binding of GCN5 and lncMX1–215. The binding fragments were detected using PCR with primers P1 to P5. l Cell lysates were incubated with biotinylated lncMX1–215 or an unrelated probe, and the eluent and flow-through were analyzed via western blotting with anti-GCN5 antibody. L: lysate load; FT: flow-through; E: eluent. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: LncMX1–215 directly interacts with GCN5. a GCN5 and H3K27ac were detected after transfection with lncMX1–215 for 24 h and treatment with 200 ng/ml IFNα for 24 h. b GCN5 and H3K27ac were detected after transfection with ASO for 48 h. c, d GCN5, PD-L1, galectin-9 and H3K27ac were examined after transfection with lncMX1–215 or ASO for 24 h followed by treatment with 5 μM MS-275 for 24 h. e GCN5, PD-L1, galectin-9 and H3K27ac were detected after ectopic expression of GCN5 for 48 h in HN4 and Cal27 cells. f PD-L1 and galectin-9 were detected after cotransfection with GCN5 and lncMX1–215 for 48 h. g After transfection with lncMX1–215 for 48 h, cell lysates were precipitated with anti-GCN5 or IgG antibody. h GCN5 and H3K27ac expression was analyzed and quantified using immunofluorescence. i The correlation between H3K27ac and GCN5 and lncMX1–215 expression was analyzed in HNSCC tissue microarray. j Overexpression of GCN5-DYKDDDDK fusion plasmid in HN4 cells was verified by western blotting. P1 to P5 were truncations for full length of lncMX1–215 amplified by specific primers. k After transfection of HN4 cells with GCN5-DYKDDDDK fusion plasmid for 48 h, RNA immunoprecipitation was used to detect binding of GCN5 and lncMX1–215. The binding fragments were detected using PCR with primers P1 to P5. l Cell lysates were incubated with biotinylated lncMX1–215 or an unrelated probe, and the eluent and flow-through were analyzed via western blotting with anti-GCN5 antibody. L: lysate load; FT: flow-through; E: eluent. *: P

    Article Snippet: Immunohistochemistry (IHC) and immunofluorescence were performed as described in our previous study [ ], with primary antibodies against PD-L1, galectin-9, H3K27ac and GCN5 (Cell Signaling Technology, Danvers, MA, USA), and Ki-67 (Proteintech, Rocky Hill, NJ, USA).

    Techniques: Transfection, Allele-specific Oligonucleotide, Mass Spectrometry, Expressing, Cotransfection, Immunofluorescence, Microarray, Over Expression, Plasmid Preparation, Western Blot, Amplification, Immunoprecipitation, Binding Assay, Polymerase Chain Reaction, Incubation, Flow Cytometry

    Gain of H3K27 acetylation activates PD-L1 and galectin-9 expression. a Layered H3K27 acetylation enrichment in the upstream region of PD-L1 and LGALS9, as shown in the UCSC database. b PD-L1 and acetylation of histone 3 were detected and quantified after treatment with 200 ng/ml IFNα or 5 μM MS-275 for 24 h. # indicated the deference between combined group and each alone. c PD-L1, H3K27ac, H3K18ac and H3K9ac were detected and quantified after the indicated MS-275 treatment for 24 h. d PD-L1 and H3K27ac were detected and quantified after 5 μM MS-275 treatment for the indicated time. e After treatment of HN4 and Cal27 cells with 15 μM SAHA or 5 μM MS-275 for 24 h, ChIP assays were performed using anti-H3K27ac antibody. f The promoter activity of PD-L1 and LGALS9 was measured after transfection with lncMX1–215 for 24 h and then SAHA or MS-275 treatment for 24 h in 293 T cells. g, h The relative mRNA of CD274 and LGALS9 were detected after transfection with lncMX1–215 and then 15 μM SAHA or 5 μM MS-275 treatment for 24 h in HN4 and Cal27 cells. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: Gain of H3K27 acetylation activates PD-L1 and galectin-9 expression. a Layered H3K27 acetylation enrichment in the upstream region of PD-L1 and LGALS9, as shown in the UCSC database. b PD-L1 and acetylation of histone 3 were detected and quantified after treatment with 200 ng/ml IFNα or 5 μM MS-275 for 24 h. # indicated the deference between combined group and each alone. c PD-L1, H3K27ac, H3K18ac and H3K9ac were detected and quantified after the indicated MS-275 treatment for 24 h. d PD-L1 and H3K27ac were detected and quantified after 5 μM MS-275 treatment for the indicated time. e After treatment of HN4 and Cal27 cells with 15 μM SAHA or 5 μM MS-275 for 24 h, ChIP assays were performed using anti-H3K27ac antibody. f The promoter activity of PD-L1 and LGALS9 was measured after transfection with lncMX1–215 for 24 h and then SAHA or MS-275 treatment for 24 h in 293 T cells. g, h The relative mRNA of CD274 and LGALS9 were detected after transfection with lncMX1–215 and then 15 μM SAHA or 5 μM MS-275 treatment for 24 h in HN4 and Cal27 cells. *: P

    Article Snippet: Immunohistochemistry (IHC) and immunofluorescence were performed as described in our previous study [ ], with primary antibodies against PD-L1, galectin-9, H3K27ac and GCN5 (Cell Signaling Technology, Danvers, MA, USA), and Ki-67 (Proteintech, Rocky Hill, NJ, USA).

    Techniques: Expressing, Mass Spectrometry, Chromatin Immunoprecipitation, Activity Assay, Transfection

    LncMX1–215 inhibits tumor proliferation capacity in vitro and in vivo . a, b The viability of HN4 and Cal27 cells after transfection with lncMX1–215 or ASO was determined using CCK8 assays. c, d Colony formation assays were performed with HN4 and Cal27 cells after transfection with lncMX1–215 or ASO. e, f EdU assays were performed after transfection with lncMX1–215 or ASO. g PARP and cleaved caspase 3 were detected via western blotting after transfection with lncMX1–215 for 48 h. h Cell apoptosis was analyzed via flow cytometry using an Annexin V/7-AAD kit after transfection for 48 h. i Cell cycle was analyzed using flow cytometry after transfection for 48 h. j Tumor growth curves for mice injected with cells treated with lncMX1–215 lentivirus or vector were analyzed. k Tumors derived from the xenograft model were resected and are shown for each group. l The weight of tumors resected from mice in the ectopic expression and vector groups was measured and analyzed. m The percentage of TUNEL-positive cells in each group was compared. n The relative Ki-67 staining score in each group was analyzed. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: LncMX1–215 inhibits tumor proliferation capacity in vitro and in vivo . a, b The viability of HN4 and Cal27 cells after transfection with lncMX1–215 or ASO was determined using CCK8 assays. c, d Colony formation assays were performed with HN4 and Cal27 cells after transfection with lncMX1–215 or ASO. e, f EdU assays were performed after transfection with lncMX1–215 or ASO. g PARP and cleaved caspase 3 were detected via western blotting after transfection with lncMX1–215 for 48 h. h Cell apoptosis was analyzed via flow cytometry using an Annexin V/7-AAD kit after transfection for 48 h. i Cell cycle was analyzed using flow cytometry after transfection for 48 h. j Tumor growth curves for mice injected with cells treated with lncMX1–215 lentivirus or vector were analyzed. k Tumors derived from the xenograft model were resected and are shown for each group. l The weight of tumors resected from mice in the ectopic expression and vector groups was measured and analyzed. m The percentage of TUNEL-positive cells in each group was compared. n The relative Ki-67 staining score in each group was analyzed. *: P

    Article Snippet: A lncMX1–215-specific ASO was designed and synthesized by Ribobio (Guangzhou, China). siRNA targeting Galectin-9 was designed and synthesized by Biotend (Shanghai, China).

    Techniques: In Vitro, In Vivo, Transfection, Allele-specific Oligonucleotide, Western Blot, Flow Cytometry, Cytometry, Mouse Assay, Injection, Plasmid Preparation, Derivative Assay, Expressing, TUNEL Assay, Staining

    Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using Transwell inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using Transwell inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P

    Article Snippet: A lncMX1–215-specific ASO was designed and synthesized by Ribobio (Guangzhou, China). siRNA targeting Galectin-9 was designed and synthesized by Biotend (Shanghai, China).

    Techniques: Over Expression, Migration, Transfection, Allele-specific Oligonucleotide, Mouse Assay, Staining, Expressing, Binding Assay

    LncMX1–215 negatively regulates IFNα-induced PD-L1 and galectin-9 expression. a lncMX1–215 expression was detected in HN4 and Cal27 cells via PCR after transfection with expression plasmids for 24 h. b, c The protein and mRNA of PD-L1 and galectin-9 expression levels were detected after transfection with lncMX1–215 for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. d The lncMX1–215 knockdown efficiency after transfection with ASO for 24 h was analyzed. e, f After ASO transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h, PD-L1 and galectin-9 expression levels were detected using western blotting and qRT-PCR. g Surface PD-L1 expression was examined using flow cytometry after transfection with lncMX1–215 followed by 200 ng/ml IFNα stimulation for 24 h. h The galectin-9 concentration in medium supernatant was measured via ELISA after transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. i Galectin-9 knockdown efficiency was detected via western blotting after transfection with siRNA for 48 h. j Transfected cells were seeded in a 96-well plate and incubated with NK cells for 4 h at various effector/target (E:T) cell ratios as indicated. The specific lysis rate was measured using an LDH kit. k Relative CD274 mRNA and lncMX1–215 expression were detected using qRT-PCR after 200 ng/ml IFNα stimulation for indicated time. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: LncMX1–215 negatively regulates IFNα-induced PD-L1 and galectin-9 expression. a lncMX1–215 expression was detected in HN4 and Cal27 cells via PCR after transfection with expression plasmids for 24 h. b, c The protein and mRNA of PD-L1 and galectin-9 expression levels were detected after transfection with lncMX1–215 for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. d The lncMX1–215 knockdown efficiency after transfection with ASO for 24 h was analyzed. e, f After ASO transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h, PD-L1 and galectin-9 expression levels were detected using western blotting and qRT-PCR. g Surface PD-L1 expression was examined using flow cytometry after transfection with lncMX1–215 followed by 200 ng/ml IFNα stimulation for 24 h. h The galectin-9 concentration in medium supernatant was measured via ELISA after transfection for 24 h followed by 200 ng/ml IFNα stimulation for 24 h. i Galectin-9 knockdown efficiency was detected via western blotting after transfection with siRNA for 48 h. j Transfected cells were seeded in a 96-well plate and incubated with NK cells for 4 h at various effector/target (E:T) cell ratios as indicated. The specific lysis rate was measured using an LDH kit. k Relative CD274 mRNA and lncMX1–215 expression were detected using qRT-PCR after 200 ng/ml IFNα stimulation for indicated time. *: P

    Article Snippet: A lncMX1–215-specific ASO was designed and synthesized by Ribobio (Guangzhou, China). siRNA targeting Galectin-9 was designed and synthesized by Biotend (Shanghai, China).

    Techniques: Expressing, Polymerase Chain Reaction, Transfection, Allele-specific Oligonucleotide, Western Blot, Quantitative RT-PCR, Flow Cytometry, Cytometry, Concentration Assay, Enzyme-linked Immunosorbent Assay, Incubation, Lysis

    LncMX1–215 directly interacts with GCN5. a GCN5 and H3K27ac were detected after transfection with lncMX1–215 for 24 h and treatment with 200 ng/ml IFNα for 24 h. b GCN5 and H3K27ac were detected after transfection with ASO for 48 h. c, d GCN5, PD-L1, galectin-9 and H3K27ac were examined after transfection with lncMX1–215 or ASO for 24 h followed by treatment with 5 μM MS-275 for 24 h. e GCN5, PD-L1, galectin-9 and H3K27ac were detected after ectopic expression of GCN5 for 48 h in HN4 and Cal27 cells. f PD-L1 and galectin-9 were detected after cotransfection with GCN5 and lncMX1–215 for 48 h. g After transfection with lncMX1–215 for 48 h, cell lysates were precipitated with anti-GCN5 or IgG antibody. h GCN5 and H3K27ac expression was analyzed and quantified using immunofluorescence. i The correlation between H3K27ac and GCN5 and lncMX1–215 expression was analyzed in HNSCC tissue microarray. j Overexpression of GCN5-DYKDDDDK fusion plasmid in HN4 cells was verified by western blotting. P1 to P5 were truncations for full length of lncMX1–215 amplified by specific primers. k After transfection of HN4 cells with GCN5-DYKDDDDK fusion plasmid for 48 h, RNA immunoprecipitation was used to detect binding of GCN5 and lncMX1–215. The binding fragments were detected using PCR with primers P1 to P5. l Cell lysates were incubated with biotinylated lncMX1–215 or an unrelated probe, and the eluent and flow-through were analyzed via western blotting with anti-GCN5 antibody. L: lysate load; FT: flow-through; E: eluent. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: LncMX1–215 directly interacts with GCN5. a GCN5 and H3K27ac were detected after transfection with lncMX1–215 for 24 h and treatment with 200 ng/ml IFNα for 24 h. b GCN5 and H3K27ac were detected after transfection with ASO for 48 h. c, d GCN5, PD-L1, galectin-9 and H3K27ac were examined after transfection with lncMX1–215 or ASO for 24 h followed by treatment with 5 μM MS-275 for 24 h. e GCN5, PD-L1, galectin-9 and H3K27ac were detected after ectopic expression of GCN5 for 48 h in HN4 and Cal27 cells. f PD-L1 and galectin-9 were detected after cotransfection with GCN5 and lncMX1–215 for 48 h. g After transfection with lncMX1–215 for 48 h, cell lysates were precipitated with anti-GCN5 or IgG antibody. h GCN5 and H3K27ac expression was analyzed and quantified using immunofluorescence. i The correlation between H3K27ac and GCN5 and lncMX1–215 expression was analyzed in HNSCC tissue microarray. j Overexpression of GCN5-DYKDDDDK fusion plasmid in HN4 cells was verified by western blotting. P1 to P5 were truncations for full length of lncMX1–215 amplified by specific primers. k After transfection of HN4 cells with GCN5-DYKDDDDK fusion plasmid for 48 h, RNA immunoprecipitation was used to detect binding of GCN5 and lncMX1–215. The binding fragments were detected using PCR with primers P1 to P5. l Cell lysates were incubated with biotinylated lncMX1–215 or an unrelated probe, and the eluent and flow-through were analyzed via western blotting with anti-GCN5 antibody. L: lysate load; FT: flow-through; E: eluent. *: P

    Article Snippet: A lncMX1–215-specific ASO was designed and synthesized by Ribobio (Guangzhou, China). siRNA targeting Galectin-9 was designed and synthesized by Biotend (Shanghai, China).

    Techniques: Transfection, Allele-specific Oligonucleotide, Mass Spectrometry, Expressing, Cotransfection, Immunofluorescence, Microarray, Over Expression, Plasmid Preparation, Western Blot, Amplification, Immunoprecipitation, Binding Assay, Polymerase Chain Reaction, Incubation, Flow Cytometry

    ASO‐21 does not affect splicing or expression of genomic targets with one mismatch ASO‐21 has a single mismatched nucleotide with three other sites in the mouse genome. The genes associated with these potential off‐target binding sites are shown. The location of the site, intronic length, position relative to nearby landmarks, the specific mismatch, and contiguous base pairs in the putative duplex between ASO‐21 and the off‐target site are shown. RT–PCR analysis of RNA isolated from the cortex of AD mice treated with either ASO‐C or ASO‐21, as in Fig 4 . The specific region of the transcript that is predicted to be affected by ASO‐21 binding was amplified, and products were separated on a 2% agarose gel. No aberrant splicing or changes in alternative splicing patterns were observed. Immunoblot analysis of MetAP2 protein from the cortex of AD mice treated with either ASO‐C or ASO‐21, as in Fig 4 . ASO‐21 could form a putative duplex upstream of the MetAP2 start codon, potentially affecting expression. Graph depicting the quantitation of MetAP2 protein expression depicted in (C). Bars represent mean ± s.e.m. Source data are available online for this figure.

    Journal: EMBO Molecular Medicine

    Article Title: Therapeutic correction of ApoER2 splicing in Alzheimer's disease mice using antisense oligonucleotides

    doi: 10.15252/emmm.201505846

    Figure Lengend Snippet: ASO‐21 does not affect splicing or expression of genomic targets with one mismatch ASO‐21 has a single mismatched nucleotide with three other sites in the mouse genome. The genes associated with these potential off‐target binding sites are shown. The location of the site, intronic length, position relative to nearby landmarks, the specific mismatch, and contiguous base pairs in the putative duplex between ASO‐21 and the off‐target site are shown. RT–PCR analysis of RNA isolated from the cortex of AD mice treated with either ASO‐C or ASO‐21, as in Fig 4 . The specific region of the transcript that is predicted to be affected by ASO‐21 binding was amplified, and products were separated on a 2% agarose gel. No aberrant splicing or changes in alternative splicing patterns were observed. Immunoblot analysis of MetAP2 protein from the cortex of AD mice treated with either ASO‐C or ASO‐21, as in Fig 4 . ASO‐21 could form a putative duplex upstream of the MetAP2 start codon, potentially affecting expression. Graph depicting the quantitation of MetAP2 protein expression depicted in (C). Bars represent mean ± s.e.m. Source data are available online for this figure.

    Article Snippet: Membranes were blocked in 5% nonfat dry milk in 1× TBST and incubated with ApoER2 C‐terminal antibody ab108208 (1:2,000) (Abcam), ApoER2 exon 19‐specific antibody (1:1,000) (a gift from Joachim Herz) (Beffert et al , ), GFAP antibody (1:5,000) (ab7260, Abcam), SRSF1‐specific antibody (1:100)(a gift from Adrian Krainer), MetAP2‐specific antibody (1:200) (D3I1H, Cell Signalling), or β‐actin‐specific antibody (1:2,000) (Sigma‐Aldrich) followed by incubation with anti‐rabbit or anti‐mouse HRP‐conjugated antibody (1:5,000) (Thermo).

    Techniques: Allele-specific Oligonucleotide, Expressing, Binding Assay, Reverse Transcription Polymerase Chain Reaction, Isolation, Mouse Assay, Amplification, Agarose Gel Electrophoresis, Quantitation Assay

    Uniform 2′-MOE ASOs cause intron retention and decreased levels of STAT3 mRNA in association with the chromatin. ( A ) Purity of the cell fractions was assessed by qRT-PCR for a chromatin RNA marker ( STAT3 intron 21) and a cytoplasmic RNA marker ( RN7SL1 ) as well as by western blotting for proteins enriched in the indicated compartments. ( B ) HeLa cells were transfected with uniform 2′-MOE (UNI6a and UNI6b) or 2′-MOE gapmer ASO (GAP1i) for 24 h, followed by cell fractionation. STAT3 expression in each fraction was analyzed by qRT-PCR of the indicated regions of STAT3 mRNA. Expression of improperly spliced mRNA was visualized by polyacrylamide separation of STAT3 RT-PCR products. ( C ) STAT3 pre-mRNA levels were analyzed in the chromatin fraction by qRT-PCR. ( D ) Following RNAPII immunoprecipitation, successful pull-down was shown by western blot (right panel) and RNAPII-bound STAT3 was analyzed by qRT-PCR. Mean ± absolute deviation ( n = 2); * P

    Journal: Nucleic Acids Research

    Article Title: Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides

    doi: 10.1093/nar/gku184

    Figure Lengend Snippet: Uniform 2′-MOE ASOs cause intron retention and decreased levels of STAT3 mRNA in association with the chromatin. ( A ) Purity of the cell fractions was assessed by qRT-PCR for a chromatin RNA marker ( STAT3 intron 21) and a cytoplasmic RNA marker ( RN7SL1 ) as well as by western blotting for proteins enriched in the indicated compartments. ( B ) HeLa cells were transfected with uniform 2′-MOE (UNI6a and UNI6b) or 2′-MOE gapmer ASO (GAP1i) for 24 h, followed by cell fractionation. STAT3 expression in each fraction was analyzed by qRT-PCR of the indicated regions of STAT3 mRNA. Expression of improperly spliced mRNA was visualized by polyacrylamide separation of STAT3 RT-PCR products. ( C ) STAT3 pre-mRNA levels were analyzed in the chromatin fraction by qRT-PCR. ( D ) Following RNAPII immunoprecipitation, successful pull-down was shown by western blot (right panel) and RNAPII-bound STAT3 was analyzed by qRT-PCR. Mean ± absolute deviation ( n = 2); * P

    Article Snippet: RNA immunoprecipitation 5 × 106 HeLa cells were transfected with 50 nM ASO using 3 μg/ml Cytofectin transfection reagent for 24 h. RNA immunoprecipitation was performed using the Magna RIPTM RNA-Binding Protein Immunoprecipitation Kit (EMD Millipore) according to the manufacturer's instructions with 3 μg ChIPAb+ RNA Pol II monoclonal antibody (EMD Millipore) or 3 μg mouse IgG.

    Techniques: Quantitative RT-PCR, Marker, Western Blot, Transfection, Allele-specific Oligonucleotide, Cell Fractionation, Expressing, Reverse Transcription Polymerase Chain Reaction, Immunoprecipitation

    P54nrb preferentially associates with 2′-F-modified oligonucleotides. ( A ) Oligonucleotides used in this experiment. ( B ) ASO-binding proteins were isolated from HeLa cell extracts using a biotinylated gapmer 2′-F-PS-ASO (ISIS623496). Proteins were eluted from beads by competition with indicated gapmers. The affinity-selected proteins were analyzed by western blot. Ku70 served as a loading control. ( C ) Western analysis following ASO-pull down as described in panel (A) indicated that P54nrb has higher affinity for oligonucleotides with the 2′-F modification on the 3′ side. Ku70 served as a loading control. ( D ) Transfection of 2′-F-oligonucleotides reduced P54nrb but not FUS protein levels. HeLa cells were transfected with 2′-F-PS-ASO (ISIS404130) at a final concentration of 30 nM, and protein levels were determined by western analysis. ( E ) Western analysis of proteins isolated from HeLa cells following treatment of cells with siRNAs targeting P54nrb or PSPC1 mRNA. Cells were transfected with siRNA at 3-nM final concentration and harvested after 36 h.

    Journal: Nucleic Acids Research

    Article Title: 2′-Fluoro-modified phosphorothioate oligonucleotide can cause rapid degradation of P54nrb and PSF

    doi: 10.1093/nar/gkv298

    Figure Lengend Snippet: P54nrb preferentially associates with 2′-F-modified oligonucleotides. ( A ) Oligonucleotides used in this experiment. ( B ) ASO-binding proteins were isolated from HeLa cell extracts using a biotinylated gapmer 2′-F-PS-ASO (ISIS623496). Proteins were eluted from beads by competition with indicated gapmers. The affinity-selected proteins were analyzed by western blot. Ku70 served as a loading control. ( C ) Western analysis following ASO-pull down as described in panel (A) indicated that P54nrb has higher affinity for oligonucleotides with the 2′-F modification on the 3′ side. Ku70 served as a loading control. ( D ) Transfection of 2′-F-oligonucleotides reduced P54nrb but not FUS protein levels. HeLa cells were transfected with 2′-F-PS-ASO (ISIS404130) at a final concentration of 30 nM, and protein levels were determined by western analysis. ( E ) Western analysis of proteins isolated from HeLa cells following treatment of cells with siRNAs targeting P54nrb or PSPC1 mRNA. Cells were transfected with siRNA at 3-nM final concentration and harvested after 36 h.

    Article Snippet: Anti-PSPC1 (ab104238), anti-hnRNP K (ab32969), anti-TCP1-β (ab92746) and anti-FUS (ab23439) were purchased from Abcam.

    Techniques: Modification, Allele-specific Oligonucleotide, Binding Assay, Isolation, Western Blot, Transfection, Concentration Assay

    PSPC1 levels are reduced by 2′-F-modified oligonucleotides. ( A ) Indicated oligonucleotides were transfected into HeLa cells at a final concentration of 30 nM. After 24 h, levels of PSPC1 were evaluated by western. GAPDH served as a loading control. ( B ) A431 cells were incubated with ISIS404130 by free uptake for 40 h at the indicated concentrations. Levels of P54nrb, PSF and PSF proteins were evaluated by western. GAPDH served as a loading control.

    Journal: Nucleic Acids Research

    Article Title: 2′-Fluoro-modified phosphorothioate oligonucleotide can cause rapid degradation of P54nrb and PSF

    doi: 10.1093/nar/gkv298

    Figure Lengend Snippet: PSPC1 levels are reduced by 2′-F-modified oligonucleotides. ( A ) Indicated oligonucleotides were transfected into HeLa cells at a final concentration of 30 nM. After 24 h, levels of PSPC1 were evaluated by western. GAPDH served as a loading control. ( B ) A431 cells were incubated with ISIS404130 by free uptake for 40 h at the indicated concentrations. Levels of P54nrb, PSF and PSF proteins were evaluated by western. GAPDH served as a loading control.

    Article Snippet: Anti-PSPC1 (ab104238), anti-hnRNP K (ab32969), anti-TCP1-β (ab92746) and anti-FUS (ab23439) were purchased from Abcam.

    Techniques: Modification, Transfection, Concentration Assay, Western Blot, Incubation

    P54nrb preferentially associates with 2′-F-modified oligonucleotides. ( A ) Oligonucleotides used in this experiment. ( B ) ASO-binding proteins were isolated from HeLa cell extracts using a biotinylated gapmer 2′-F-PS-ASO (ISIS623496). Proteins were eluted from beads by competition with indicated gapmers. The affinity-selected proteins were analyzed by western blot. Ku70 served as a loading control. ( C ) Western analysis following ASO-pull down as described in panel (A) indicated that P54nrb has higher affinity for oligonucleotides with the 2′-F modification on the 3′ side. Ku70 served as a loading control. ( D ) Transfection of 2′-F-oligonucleotides reduced P54nrb but not FUS protein levels. HeLa cells were transfected with 2′-F-PS-ASO (ISIS404130) at a final concentration of 30 nM, and protein levels were determined by western analysis. ( E ) Western analysis of proteins isolated from HeLa cells following treatment of cells with siRNAs targeting P54nrb or PSPC1 mRNA. Cells were transfected with siRNA at 3-nM final concentration and harvested after 36 h.

    Journal: Nucleic Acids Research

    Article Title: 2′-Fluoro-modified phosphorothioate oligonucleotide can cause rapid degradation of P54nrb and PSF

    doi: 10.1093/nar/gkv298

    Figure Lengend Snippet: P54nrb preferentially associates with 2′-F-modified oligonucleotides. ( A ) Oligonucleotides used in this experiment. ( B ) ASO-binding proteins were isolated from HeLa cell extracts using a biotinylated gapmer 2′-F-PS-ASO (ISIS623496). Proteins were eluted from beads by competition with indicated gapmers. The affinity-selected proteins were analyzed by western blot. Ku70 served as a loading control. ( C ) Western analysis following ASO-pull down as described in panel (A) indicated that P54nrb has higher affinity for oligonucleotides with the 2′-F modification on the 3′ side. Ku70 served as a loading control. ( D ) Transfection of 2′-F-oligonucleotides reduced P54nrb but not FUS protein levels. HeLa cells were transfected with 2′-F-PS-ASO (ISIS404130) at a final concentration of 30 nM, and protein levels were determined by western analysis. ( E ) Western analysis of proteins isolated from HeLa cells following treatment of cells with siRNAs targeting P54nrb or PSPC1 mRNA. Cells were transfected with siRNA at 3-nM final concentration and harvested after 36 h.

    Article Snippet: Anti-PSPC1 (ab104238), anti-hnRNP K (ab32969), anti-TCP1-β (ab92746) and anti-FUS (ab23439) were purchased from Abcam.

    Techniques: Modification, Allele-specific Oligonucleotide, Binding Assay, Isolation, Western Blot, Transfection, Concentration Assay

    MiR-195 blocked Wnt/β-catenin pathway. SW480 cells were transfected with miR-195 mimic, ASO of miR-195 or their corresponding controls. Then, the protein expressions of (A) β-catenin and Ki-67s, as well as (B) GSK-3β and p-GSK-3β

    Journal: American Journal of Cancer Research

    Article Title: MicroRNA-195 suppresses colorectal cancer cells proliferation via targeting FGF2 and regulating Wnt/β-catenin pathway

    doi:

    Figure Lengend Snippet: MiR-195 blocked Wnt/β-catenin pathway. SW480 cells were transfected with miR-195 mimic, ASO of miR-195 or their corresponding controls. Then, the protein expressions of (A) β-catenin and Ki-67s, as well as (B) GSK-3β and p-GSK-3β

    Article Snippet: After blocked with 5% nonfat milk for 1 h at room temperature, the membranes were incubated with specific primary antibodies against CyclinB1 (ab32053), CyclinD1 (ab134175), Cyclin-Dependent Kinase 2 (CDK2; ab6433), FGF2 (ab16828), β-catenin (ab32572), Ki-67 (ab15580), Glycogen Synthase Kinase 3β (GSK-3β; ab93926), phosphorylated GSK-3β (p-GSK-3β; ab131097), Transcription Factor 4 (TCF-4; ab60727), Lymphoid Enhancer Binding Factor 1 (LEF-1; ab53293) and GAPDH (ab8245) (Abcam, Cambridge, UK), at 4°C overnight.

    Techniques: Transfection, Allele-specific Oligonucleotide

    Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using Transwell inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P

    Journal: Molecular Cancer

    Article Title: A novel IFNα-induced long noncoding RNA negatively regulates immunosuppression by interrupting H3K27 acetylation in head and neck squamous cell carcinoma

    doi: 10.1186/s12943-019-1123-y

    Figure Lengend Snippet: Overexpression of lncMX1–215 suppresses HNSCC metastasis. a, b Migration and invasion assays were performed with transfected cells using Transwell inserts (bar = 100 μm, 20 μm, respectively). c snail was detected in HN4 and Cal27 cells after transfection with lncMX1–215 or ASO for 48 h. d Representative images of lungs from mice in each experimental group. The arrows indicate individual metastatic nodules. Metastatic tumors in lung tissues were identified via hematoxylin and eosin staining. e The number of metastatic nodules in each group was analyzed. f Schematic diagram showing that IFNα-induced lncMX1–215 can negatively regulate PD-L1 and galectin-9 expression by interrupting GCN5/H3K27ac binding in HNSCC. *: P

    Article Snippet: Transwell assay Cell migration and invasion assays were performed using Transwell assay with uncoated polycarbonate inserts (Millipore, Darmstadt, Germany) for migration or BioCoat™ inserts (BD Biosciences, Franklin Lake, NJ, USA) for invasion.

    Techniques: Over Expression, Migration, Transfection, Allele-specific Oligonucleotide, Mouse Assay, Staining, Expressing, Binding Assay

    NRAL controls the target of miR-340-5p. (a) Schematic diagram of the miR-340-5p binding site in the Nrf2 3’UTR or its mutant sequences based on TargetScan software. (b) Relative luciferase intensities of the luciferase constructors harbouring the wild-type or mutant Nrf2 3’UTRs were measured in 293T cells 48 h after transfection with the pri-miR-340-5p plasmid or ASO-miR-340-5p, n=3, ** p

    Journal: Journal of Cell Communication and Signaling

    Article Title: NRAL mediates cisplatin resistance in hepatocellular carcinoma via miR-340-5p/Nrf2 axis

    doi: 10.1007/s12079-018-0479-x

    Figure Lengend Snippet: NRAL controls the target of miR-340-5p. (a) Schematic diagram of the miR-340-5p binding site in the Nrf2 3’UTR or its mutant sequences based on TargetScan software. (b) Relative luciferase intensities of the luciferase constructors harbouring the wild-type or mutant Nrf2 3’UTRs were measured in 293T cells 48 h after transfection with the pri-miR-340-5p plasmid or ASO-miR-340-5p, n=3, ** p

    Article Snippet: The membranes were incubated with anti-Keap1 (1:2000 dilution, Abcam, UK), anti-Nrf2 (1:4000 dilution, Abcam, UK), and anti-actin (1:6000, Santa Cruz, CA) primary antibodies and then with a secondary horseradish peroxidase-labeled goat anti-mouse IgG or goat anti-rabbit IgG antibody for 1 hr.

    Techniques: Binding Assay, Mutagenesis, Software, Luciferase, Transfection, Plasmid Preparation, Allele-specific Oligonucleotide

    Effect of knocking down JNK1 and/or -2 on JNK translocation to mitochondria, mitochondria bioenergetics, and liver injury. A , effect of JNK1 and -2 ASO on JNK translocation to mitochondria induced by APAP treatment. B , effect of silencing JNK1 and -2

    Journal:

    Article Title: Role of JNK Translocation to Mitochondria Leading to Inhibition of Mitochondria Bioenergetics in Acetaminophen-induced Liver Injury *Role of JNK Translocation to Mitochondria Leading to Inhibition of Mitochondria Bioenergetics in Acetaminophen-induced Liver Injury * S⃞

    doi: 10.1074/jbc.M708916200

    Figure Lengend Snippet: Effect of knocking down JNK1 and/or -2 on JNK translocation to mitochondria, mitochondria bioenergetics, and liver injury. A , effect of JNK1 and -2 ASO on JNK translocation to mitochondria induced by APAP treatment. B , effect of silencing JNK1 and -2

    Article Snippet: Addition of Active JNK1 or JNK2 to Isolated Mitochondria —In some experiments isolated mitochondria from control and APAP plus JNK inhibitor-treated mice (following 4-h APAP treatment) were incubated with purified active JNK1 or JNK 2 (Millipore, Billerica, MA) in incubation buffer (230 m m mannitol, 70 m m sucrose, 30 m m Tris-HCl, 5 m m KH2 PO4 , 1 m m EDTA, MgCl2 10 m m , ATP 500 μ m , pH 7.4) for 10 min at room temperature ( ).

    Techniques: Translocation Assay, Allele-specific Oligonucleotide

    Addition of purified active JNK1 or JNK2 inhibits mitochondria bioenergetics and induces MPT in redox altered mitochondria. Isolated mitochondria from control and JNK inhibitor plus APAP treated (redox altered due to GSH depletion and covalent binding)

    Journal:

    Article Title: Role of JNK Translocation to Mitochondria Leading to Inhibition of Mitochondria Bioenergetics in Acetaminophen-induced Liver Injury *Role of JNK Translocation to Mitochondria Leading to Inhibition of Mitochondria Bioenergetics in Acetaminophen-induced Liver Injury * S⃞

    doi: 10.1074/jbc.M708916200

    Figure Lengend Snippet: Addition of purified active JNK1 or JNK2 inhibits mitochondria bioenergetics and induces MPT in redox altered mitochondria. Isolated mitochondria from control and JNK inhibitor plus APAP treated (redox altered due to GSH depletion and covalent binding)

    Article Snippet: Addition of Active JNK1 or JNK2 to Isolated Mitochondria —In some experiments isolated mitochondria from control and APAP plus JNK inhibitor-treated mice (following 4-h APAP treatment) were incubated with purified active JNK1 or JNK 2 (Millipore, Billerica, MA) in incubation buffer (230 m m mannitol, 70 m m sucrose, 30 m m Tris-HCl, 5 m m KH2 PO4 , 1 m m EDTA, MgCl2 10 m m , ATP 500 μ m , pH 7.4) for 10 min at room temperature ( ).

    Techniques: Purification, Isolation, Binding Assay

    ( A ) Oligreen dye-exclusion assays to determine bolasome binding capacity using 2 µg gapmer ASO. ( B ) Bolasome complexation and retention of phosphorothioate antisense gapmers following metaphor electrophoresis and SYBR Green II staining. Lanes 1–4: Amine-to-phosphate (N/P) ratio =0, 0.5, 2.5, 5.0. Abbreviation: ASO, antisense oligonucleotides.

    Journal: International Journal of Nanomedicine

    Article Title: Bolaamphiphile-based nanocomplex delivery of phosphorothioate gapmer antisense oligonucleotides as a treatment for Clostridium difficile

    doi: 10.2147/IJN.S109600

    Figure Lengend Snippet: ( A ) Oligreen dye-exclusion assays to determine bolasome binding capacity using 2 µg gapmer ASO. ( B ) Bolasome complexation and retention of phosphorothioate antisense gapmers following metaphor electrophoresis and SYBR Green II staining. Lanes 1–4: Amine-to-phosphate (N/P) ratio =0, 0.5, 2.5, 5.0. Abbreviation: ASO, antisense oligonucleotides.

    Article Snippet: The fluorescent single-stranded nucleic acid dye, Oligreen (Thermo Fisher Scientific, Wilmington, DE, USA), does not effectively bind to single-stranded phosphorothioate bases following cationic complexation, leading to a large decrease in fluorescence emission intensity compared with free oligonucleotide.

    Techniques: Binding Assay, Allele-specific Oligonucleotide, Electrophoresis, SYBR Green Assay, Staining

    miR-153 negatively regulates KLF5 expression in LSCC. (A and B) Western blot analysis of AKT1 and FOXO3a expression in HEp-2 cells transfected with miR-153 mimics, ASO or NC. (C) Informatics prediction of miR-153 binding sites in the 3′-UTRs of

    Journal: Experimental and Therapeutic Medicine

    Article Title: MicroRNA-153 inhibits the proliferation and invasion of human laryngeal squamous cell carcinoma by targeting KLF5

    doi: 10.3892/etm.2016.3189

    Figure Lengend Snippet: miR-153 negatively regulates KLF5 expression in LSCC. (A and B) Western blot analysis of AKT1 and FOXO3a expression in HEp-2 cells transfected with miR-153 mimics, ASO or NC. (C) Informatics prediction of miR-153 binding sites in the 3′-UTRs of

    Article Snippet: The primary antibodies anti-cyclin B1 (mouse monoclonal antibody; ab72; 1:3,000), anti-cyclin D1 (rabbit polyclonal antibody; ab16663; 1:2,000), anti-kruppel-like factor 5 (KLF5; rabbit polyclonal antibody; ab24331; 1:2,000), anti-AKT1 (rabbit polyclonal antibody; ab32505; 1:1,000), anti-forkhead box O3 (FOXO3a; rabbit polyclonal antibody; ab12162; 1:1,000) and anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH; rabbit polyclonal antibody; ab181602, 1:1,000) were purchased from Abcam.

    Techniques: Expressing, Western Blot, Transfection, Allele-specific Oligonucleotide, Binding Assay

    Effects of miR-494 on the insulin signaling pathway. The mimic form of miR-494 was overexpressed in cells. Cells were starved for 4 h before treatment with 0, 10, 100 nM of insulin for 20 min, and the levels of p-Akt (Ser473 and Thr308), p-AS160, p-p70S6K, and p-GSK-3α/β were measured in C 2 C 12 myoblasts (A) and CHO IR/IRS-1 cells (D). Bars show densitometric quantitation of phosphorylations of Akt, GSK-3α/β, AS160 and p70S6K in C 2 C 12 myoblasts (B) and CHO IR/IRS-1 cells (E). (C) The level of p-IRS1 (Tyr608) was measured in insulin-treated C 2 C 12 myoblasts. GAPDH and actin were used as loading controls. Levels of phospho-proteins between miR-494 transfected cells and control were compared at each concentration of insulin. The values are expressed as the means ± SEM. *P

    Journal: PLoS ONE

    Article Title: MicroRNA-494, Upregulated by Tumor Necrosis Factor-?, Desensitizes Insulin Effect in C2C12 Muscle Cells

    doi: 10.1371/journal.pone.0083471

    Figure Lengend Snippet: Effects of miR-494 on the insulin signaling pathway. The mimic form of miR-494 was overexpressed in cells. Cells were starved for 4 h before treatment with 0, 10, 100 nM of insulin for 20 min, and the levels of p-Akt (Ser473 and Thr308), p-AS160, p-p70S6K, and p-GSK-3α/β were measured in C 2 C 12 myoblasts (A) and CHO IR/IRS-1 cells (D). Bars show densitometric quantitation of phosphorylations of Akt, GSK-3α/β, AS160 and p70S6K in C 2 C 12 myoblasts (B) and CHO IR/IRS-1 cells (E). (C) The level of p-IRS1 (Tyr608) was measured in insulin-treated C 2 C 12 myoblasts. GAPDH and actin were used as loading controls. Levels of phospho-proteins between miR-494 transfected cells and control were compared at each concentration of insulin. The values are expressed as the means ± SEM. *P

    Article Snippet: Antibody for p-IRS1 (Y612) was from Millipore (Billerica, MA, USA).

    Techniques: Quantitation Assay, Transfection, Concentration Assay

    Regulation of putative target genes by miR-494. (A) The secondary structure of mouse premature(pre)-miR-494 (miRbase, Accession MI0003532). (B) Alignment of the seed matches in conserved sites of Stxbp5, Igf1r, Rab35, Irs1 genes at 3’ untranslated region (3’ UTR) and seed types of each target gene. Entrez accession numbers and binding position at 3’ UTR are addressed for the representative transcript of each gene. Complementary sequences binding to miR-494 are shown in bold letters, with the m8 and A1 extensions highlighted in bold italic letters. Gene abbreviations: Stxbp5 , syntaxin binding protein 5; Igf1r , insulin-like growth factor 1 receptor; Irs1, insulin receptor substrate 1 (C) RT-PCR analysis for predicted target genes in miR-494 mimic- or miR-494 ASO-transfected C 2 C 12 cells. Gapdh gene was used as a loading control. (E) Immunoblot analysis of molecules related to the insulin signaling pathway. Cells were transfected with miR-494 mimic and subjected to immunoblot analysis. Antibodies for PTEN, p-PTP1B (S50) and IRS-1 were used. GAPDH was used as a loading control. The values were expressed as the means ± SEM and compared between miR-494 mimic transfected cells and control. *P

    Journal: PLoS ONE

    Article Title: MicroRNA-494, Upregulated by Tumor Necrosis Factor-?, Desensitizes Insulin Effect in C2C12 Muscle Cells

    doi: 10.1371/journal.pone.0083471

    Figure Lengend Snippet: Regulation of putative target genes by miR-494. (A) The secondary structure of mouse premature(pre)-miR-494 (miRbase, Accession MI0003532). (B) Alignment of the seed matches in conserved sites of Stxbp5, Igf1r, Rab35, Irs1 genes at 3’ untranslated region (3’ UTR) and seed types of each target gene. Entrez accession numbers and binding position at 3’ UTR are addressed for the representative transcript of each gene. Complementary sequences binding to miR-494 are shown in bold letters, with the m8 and A1 extensions highlighted in bold italic letters. Gene abbreviations: Stxbp5 , syntaxin binding protein 5; Igf1r , insulin-like growth factor 1 receptor; Irs1, insulin receptor substrate 1 (C) RT-PCR analysis for predicted target genes in miR-494 mimic- or miR-494 ASO-transfected C 2 C 12 cells. Gapdh gene was used as a loading control. (E) Immunoblot analysis of molecules related to the insulin signaling pathway. Cells were transfected with miR-494 mimic and subjected to immunoblot analysis. Antibodies for PTEN, p-PTP1B (S50) and IRS-1 were used. GAPDH was used as a loading control. The values were expressed as the means ± SEM and compared between miR-494 mimic transfected cells and control. *P

    Article Snippet: Antibody for p-IRS1 (Y612) was from Millipore (Billerica, MA, USA).

    Techniques: Binding Assay, Reverse Transcription Polymerase Chain Reaction, Allele-specific Oligonucleotide, Transfection