C myc

Manufactured by Thermo Fisher Scientific

Sourced in United States, China

The C-Myc is a lab equipment product designed for the detection and analysis of the c-Myc protein. It is a key tool for researchers studying cell proliferation and oncogenic pathways.

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Lab products found in correlation

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Anti-c-Myc antibody (8 citations) Anti-c-Myc magnetic beads (8 citations) Pierce™ Anti-c-Myc Agarose (7 citations) Anti-c-Myc Agarose (3 citations) Mouse anti-c-Myc-HRP (2 citations) Peirce c-Myc Tag IP/Co-IP Kit (2 citations) Pierce™ c-Myc Peptide (2 citations) C-Myc antibody (2 citations) Anti-c-Myc-Magnetic Dynabeads (2 citations) C-Myc Tag IP/Co-IP kit (2 citations)

60 protocols using c myc

Quantitative Real-Time PCR for mRNA and pri-miRNA Expression

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For mRNA and pri-miR expression analysis, 0.5 μg RNA was reversely transcribed to cDNA by use of the Omniscript RT Kit (Qiagen, Cat. # 205111) with Primer “random” (Roche, Cat. # 11034731001), following the manufacturer’s protocol. The cDNA was amplified using 1 µL of the cDNA preparation, the Taqman Gene Expression Master Mix (Applied Biosystems, Cat. # 4440040), and Taqman probes for hnRNPU, c-myc, GAPDH, and 18S (all from Thermo Fisher Scientific: hnRNPU Cat. # 4331182, Assay ID Hs00244919_m1; GAPDH, Cat. # 4331182, Assay ID Hs02786624_g1; c-myc, Cat. # 4453320, Assay ID Hs00153408_m1; 18S, Cat. # 4331182, Assay ID Hs99999901_s1; pri-miR-30c-1, Cat. # 4427012, Assay ID Hs03303371_pri; pri-miR-30c-2 Cat. # 4427012, Assay ID Hs03302833_pri) in a 7500 HT real-time PCR instrument (Applied Biosystems). GAPDH was used as an internal control and relative expression was calculated as ΔΔCT values.

Zietzer A., Hosen M.R., Wang H., Goody P.R., Sylvester M., Latz E., Nickenig G., Werner N, & Jansen F. (2020). The RNA-binding protein hnRNPU regulates the sorting of microRNA-30c-5p into large extracellular vesicles. Journal of Extracellular Vesicles, 9(1), 1786967.

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Lipogenic Gene Expression Analysis

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Gene Expression Assays for human FASN (Hs01005622_m1), SCD (Hs01682761_m1), c-MYC (Hs00153408_m1), ACC/ACACA (Hs01046047_m1), ACLY (Hs00982738_m1), SREBF1 (Hs01088679_g1), Hmgcr (Hs00168352_m1), SQS/FDFT1 (Hs00926054_m1), SREBF2 (Hs01081784_m1), LSS (Hs01552331_m1), LPL (Hs00173425_m1), CD36 (Hs00354519_m1), and β-Actin (4333762T), as well as for mouse Fasn (Mm00662319_m1), Acc/Acaca (Mm01304257_m1), Acly (Mm01302282_m1), Scd1 (Mm00772290_m1), Scd2 (Mm01208542_m1), SREBF1 (Mm00550338_m1), Cdk1 (Mm00772472_m1), Cdk2 (Mm00443947_m1), Skp2 (Mm00449925_m1), Birc5/Survivin (Mm00599749_m1), c-MYC (Mm00662319_m1), Hmgcr (Mm01282499_m1), Mvk (Mm00445773_m1), SQS/FDFT1 (Mm01598574_g1), SREBF2 (Mm01306292_m1), Lpl (Mm00434764_m1), Lss (Mm00461312_m1), Cd36 (Mm00432403_m1), and β-Actin (Mm00607939_s1) genes were purchased from Applied Biosystems (Foster City, CA, USA). Quantitative values for each gene were calculated by using the PE Biosystems Analysis software and expressed as number target (NT). NT = 2^−ΔCt, wherein the ΔCt value of each sample was calculated by subtracting the average Ct value of the target gene from the average Ct value of the β-Actin gene. Experiments were repeated three times in triplicate.

Jia J., Che L., Cigliano A., Wang X., Peitta G., Tao J., Zhong S., Ribback S., Evert M., Chen X, & Calvisi D.F. (2020). Pivotal Role of Fatty Acid Synthase in c-MYC Driven Hepatocarcinogenesis. International Journal of Molecular Sciences, 21(22), 8467.

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Western Blot and ChIP-Seq Workflow

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α-³²P dCTP (6,000 Ci/mmol) was purchased from PerkinElmer (Boston, MA). Antibodies used for Western, ChIP, Seq-ChIP, and immunohistochemistry (IHC) to PHB1, MATα1, c-MYC, MAFG, c-MAF, β-ACTIN, and IgG were purchased from Abcam (Cambridge, MA). Lipofectamine 2000 and RNAi-Max were purchased from ThermoFisher (Carlsbad, CA). siRNAs to PHB1 (Cat# 4392422), c-MYC (5′-CGAUUCCUUCUAACAGAAtt-3′), MAFG (5′-CGGACUAGAGAGAGUUGCGtt-3′), c-MAF (5′-GCAUCGUGUACUUACCAGUtt-3′) and MAT1A (5′-GCACAACGAAGACAUCACGtt-3′) were purchased from ThermoFisher. Bile acids and interleukin-6 (IL-6) were from Sigma (St. Louis, MS).

Fan W., Yang H., Liu T., Wang J., Li T.W., Mavila N., Tang Y., Yang J., Peng H., Tu J., Annamalai A., Noureddin M., Krishnan A., Gores G.J., Martínez-Chantar M., Mato J.M, & Lu S.C. (2017). Prohibitin 1 Suppresses Liver Cancers Tumorigenesis in Mice and Human Hepatocellular and Cholangiocarcinoma cells. Hepatology (Baltimore, Md.), 65(4), 1249-1266.

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Western Blot Analysis of Protein Expression

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With respect to the detection of protein expression, Western blot was carried out as previously described [24] (link). After 48 h of transfection, cells were harvested for analysis of protein expression. In brief, a total of 20 μg protein extract from tissues and cells was loaded onto polyacrylamide gels and transferred onto nitrocellulose membranes prior to blocking aspecific signals using defatted dry milk. Then, the membranes were incubated with the antibodies of proliferating cell nuclear antigen (PCNA; 1:1000; Thermo Fisher), B-cell lymphoma 2 (Bcl-2; 1:500; Thermo Fisher), Bcl-2-like protein 4 (Bax; 1:6000; Thermo Fisher), TRIM66 (1:1000; Thermo Fisher), β-catenin (1:2000; Affinity, Nanjing, China), c-myc (1:1000; Thermo Fisher), cyclinD1 (1:2000; Affinity), MMP2 (1:3000; Abcam, Cambridge, UK) and MMP9 (1:5000; Abcam) and GAPDH (1:400; Thermo Fisher). The blots were probed with secondary antibodies (1:4000; Thermo Fisher). At last, protein visualization was achieved using eyoECL Plus (Beyotime, Shanghai, China).

Wang W., Wang J., Li Y, & Zhao Y. (2022). Circ_0051079 silencing inhibits the malignant phenotypes of osteosarcoma cells by the TRIM66/Wnt/β-catenin pathway in a miR-625-5p-dependent manner. Journal of Bone Oncology, 35, 100436.

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Vorinostat and CKD-581 Anticancer Protocol

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Suberoylanilide hydroxamic acid (SAHA, Vorinostat) was obtained from Sigma-Aldrich (St. Louis, MO, USA). CKD-581 (purity 98.80%, M.W. 588.72) was supplied from CKD Pharmaceutical Corporation (Seoul, Korea). CellTiter-Glo luminescent cell viability assay kit was purchased from Promega (Madison, WI, USA). Antibodies recognizing DACT1, DACT3, β-catenin, Cyclin D1, CDK4, p53, BCL-xL and BCL-2 were supplied from Santa Cruz Biotechnology (Dallas, TX, USA). Antibodies against histone H3, acetylated histone H3, histone H4, acetylated histone H4, acetylated tubulin, p21, phospho-p53 cleaved-caspase 3, AKT, phospho-AKT, p70S6K, phospho-p70S6K (Thr389), mTOR and phospho-mTOR (Ser2448) were obtained from Cell Signaling Technology (Danvers, MA, USA). Antibodies targeting DACT2, c-Myc and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Kim S.J., Kim S., Choi Y.J., Kim U.J, & Kang K.W. (2022). CKD-581 Downregulates Wnt/β-Catenin Pathway by DACT3 Induction in Hematologic Malignancy. Biomolecules & Therapeutics, 30(5), 435-446.

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Generation and Characterization of hiPSCs

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Human fibroblasts were cultured and reprogrammed using the CytoTune Sendai reprogramming vectors Oct4, Klf4, Sox2, and c-Myc (Thermo Fisher Scientific) as previously reported^{37 (link)}. The emergent iPSC colonies were picked under a stereomicroscope and expanded onto mitomycined human foreskin feeder layers. After the generation of a frozen stocks, iPSCs were preferentially adapted and cultured in feeder-free conditions^{34 (link)}. After ten passages, the clearance of the exogenous reprogramming factors and Sendai virus genome was confirmed by qPCR following the manufacturer’s instructions (Thermo Fisher Scientific). The absence of mycoplasma contamination was verified by the MycoAlert™ Mycoplasma Detection Kit (selective biochemical test of mycoplasma enzymes) according to the manufacturer’s instructions (Lonza).

Rodrigues A., Slembrouck-Brec A., Nanteau C., Terray A., Tymoshenko Y., Zagar Y., Reichman S., Xi Z., Sahel J.A., Fouquet S., Orieux G., Nandrot E.F., Byrne L.C., Audo I., Roger J.E, & Goureau O. (2022). Modeling PRPF31 retinitis pigmentosa using retinal pigment epithelium and organoids combined with gene augmentation rescue. NPJ Regenerative Medicine, 7, 39.

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Xenogeneic-free Derivation and Maintenance of hiPSCs

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The hiPSC line utilized was originally generated from human neonatal fibroblast cells isolated from a healthy female donor using non-integrative Sendai viral particles encoding human OCT4, KLF4, SOX2, and c-MYC genes (ThermoFisher) as previously described [24 (link)]. In this study, hiPSCs were cultured under either xenogeneic or xenogeneic-free conditions.
Under the xenogeneic condition, hiPSCs were maintained in mTeSR1 medium (STEMCELL Technologies) on surfaces coated with Growth Factor Reduced (GFR)-Matrigel (Corning; xenogeneic reagent derived from mouse sarcoma) with daily medium changes. Once at 80% confluency, hiPSCs were dissociated with 0.5 mM ethylenediaminetetraacetic acid (EDTA; ThermoFisher) and subcultured.
Under the xenogeneic-free condition, hiPSCs were cultured in chemically-defined, xenogeneic-free Essential 8 Medium (E8 medium; ThermoFisher) on surfaces coated with 10 μg/mL recombinant human Vitronectin XF™ (STEMCELL Technologies) with daily medium changes [33 (link)]. When cells reached 80% confluency, hiPSCs were dissociated with 0.5 mM EDTA and subcultured.

Luo J., Lin Y., Shi X., Li G., Kural M.H., Anderson C.W., Ellis M.W., Riaz M., Tellides G., Niklason L.E, & Qyang Y. (2020). Xenogeneic-free Generation of Vascular Smooth Muscle Cells from Human Induced Pluripotent Stem Cells for Vascular Tissue Engineering. Acta biomaterialia, 119, 155-168.

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iPSC Characterization by Immunofluorescence

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The iPSC clones were confirmed as IPSCs with a battery of rabbit anti-mouse IPSC markers including Oct3/4, Sox2, c-Myc, mKlf4, Nestin and SSEA-1 (Thermo Fisher Scientific, Waltham, MA, USA). The secondary antibody was an Alexa Fluor 594-conjugated goat anti-rabbit (Thermo Fisher Scientific), all used in accordance with the manufacturer’s conditions. To immobilize the iPSC clones, glass-bottom dishes were coated with Cell-TEK adhesive. The adherent iPSC was then fixed with 4% paraformaldehyde, after permeabilizing with TX-100 and blocking with normal goat serum. The iPSC clones were then incubated with the primary antibodies, washed, and incubated with the secondary antibodies. The dishes were finally mounted with Vectorshield mounting medium with DAPI (#H-1200) (Vector Laboratories, Burlingame, CA, USA) and viewed with an Olympus Fluoview-1000 confocal scanning system under different wavelengths.

Petrova S.C., Ahmad I., Nguyen C., Ferrell SD J.r., Wilhelm S.R., Ye Y, & Barsky S.H. (2020). Regulation of breast cancer oncogenesis by the cell of origin’s differentiation state. Oncotarget, 11(43), 3832-3848.

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Generation of iPSC from Blood Cells

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This study was approved by the Institutional Review Board of Severance Hospital, Yonsei University College of Medicine (3-2017-0167). Twenty milliliters of peripheral blood were obtained from one ARB patient (L40P and A195V mutations in BEST1 gene), one autosomal dominant BD patient (G96A mutation in BEST1 gene), and two normal controls. Human iPSC lines were generated using previously established methods.17 (link)18 (link) Mononuclear cells (MNC) were isolated from the blood samples and were expanded in MNC media for 7–10 days.
MNC were transfected with episomal vectors (OCT4, SOX2, c-MYC, KLF4, LIN28) (Thermo Fisher Scientific, Waltham, MA, USA) and cultured on extracellular matrices (BD, Franklin Lakes, NJ, USA). Culture on iPSC medium (STEMCELL Technologies, Vancouver, Canada) were performed for 1–2 weeks till colonization of iPSC. iPSC colonies were isolated and expanded on iPSC medium (STEMCELL Technologies). Differentiation was initiated after 10–20 passages of expansion to remove an epigenetic memory.

Lee J.H., Oh J.O, & Lee C.S. (2020). Induced Pluripotent Stem Cell Modeling of Best Disease and Autosomal Recessive Bestrophinopathy. Yonsei Medical Journal, 61(9), 816-825.

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Silencer siRNA Knockdown of Transcription Factors

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Silencer siRNA Transfection Kit (Thermo Fisher Scientific, Waltham, MA, USA) was used for the knockdown of selected genes (NF-κB, c-Myc and STAT3), according to the manufacturer’s instructions (Figure S1). The transcription factors were selected according to the literature, as the factors involved in the process of airway remodeling, but also as suggested elements of the relaxin pathway in this process [23 (link),101 (link),102 (link),103 (link),104 (link),105 (link)]. The cells were treated with 20 nM siRNA mixture against NF-κB (NCBI accession no. NM_001145138.1), c-Myc (NCBI accession no. NM_002467.4) and STAT3 mRNA (NCBI accession no. NM_003150.3), (Thermo Fisher Scientific, Waltham, MA USA) for 48 h. The same concentration of scrambled siRNAs was used as the negative control. The knockdown efficiency was evaluated after 48 h of transfection. The measurement of gene knockdown was analyzed according to the manufacturer’s protocol by qPCR. The data presented are normalized to the samples treated with control siRNA.

Wieczfinska J, & Pawliczak R. (2022). Relaxin Affects Airway Remodeling Genes Expression through Various Signal Pathways Connected with Transcription Factors. International Journal of Molecular Sciences, 23(15), 8413.

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