Anti srsf1

Manufactured by Thermo Fisher Scientific

Sourced in United States

Anti-SRSF1 is a laboratory reagent for use in molecular biology research. It is a protein-specific antibody that binds to the Serine/arginine-rich splicing factor 1 (SRSF1), which is involved in the regulation of pre-mRNA splicing. The core function of Anti-SRSF1 is to enable the detection and study of SRSF1 in various experimental systems.

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Lab products found in correlation

Anti gapdh, by Merck Group (2 mentions) Anti hnrnp f h, by Santa Cruz Biotechnology (2 mentions) Anti u1a, by Thermo Fisher Scientific (2 mentions) Anti actin, by Merck Group (1 mentions) Anti v5, by Thermo Fisher Scientific (1 mentions) R960 25, by Thermo Fisher Scientific (1 mentions) Anti h3s10p, by Cell Signaling Technology (1 mentions) Anti parp 1, by Santa Cruz Biotechnology (1 mentions) Anti srsf3, by Thermo Fisher Scientific (1 mentions) Anti mad2, by Fortis Life Sciences (1 mentions) Anti γh2ax, by Cell Signaling Technology (1 mentions) Anti p21, by Santa Cruz Biotechnology (1 mentions) Protein a g bead, by Thermo Fisher Scientific (1 mentions) Anti mettl3, by Abcam (1 mentions) Anti m6a, by Abcam (1 mentions) Anti histone h3, by Abcam (1 mentions) Pvdf membrane, by Merck Group (1 mentions) Anti α tubulin, by Abcam (1 mentions) Anti lamin b1, by Abcam (1 mentions) Rneasy minelute cleanup kit, by Qiagen (1 mentions)

Spelling variants of this product

SRSF1 (5 citations)

13 protocols using anti srsf1

Comprehensive Protein Extraction and Analysis

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For protein extraction, all cell lines were lysed in RIPA buffer or in a less stringent buffer (20 mM Tris HCl pH 8, 137 mM NaCl, 1% NP-40, 2 mM ethylenediaminetetraacetic acid (EDTA)) for the SRSF3 ISO2 (SRSF3-TR) detection. The homogenates were subjected to western blot analyses as reported (26 (link)). Antibodies used were anti-SLU7 (Novus, 1:1000), anti-PARP1 (Santa Cruz SC-7150; 1:1000, CA, USA), anti-Actin (Sigma A2066; 1:6000, MO, USA), anti-Sororin (kindly provided by Dr JM Peters, Austria), anti-SRSF3 (Thermo Fisher 33-4200; 1:500 and MBL RN080PW; 1:500, MA, USA), anti-WAPL (Cell signaling 77428S; 1:1000, MA, USA), anti-H3S10P (Cell signaling #9701; 1:1000), anti-γ-H2AX (Cell signaling #2577S; 1:1000), anti-SRSF1 (Thermo Fisher 32-4600; 1:1000), anti-MAD2 (Bethyl laboratories a300-301A-M; 1:1000, TX, USA), anti-P21 (Santa Cruz SC-397; 1:1000), anti-RNaseH1 (Thermo Fisher PA5-30974; 1:1000) and anti-V5 (Invitrogene R960-25; 1:1000).

Jiménez M., Urtasun R., Elizalde M., Azkona M., Latasa M.U., Uriarte I., Arechederra M., Alignani D., Bárcena-Varela M., Álvarez-Sola G., Colyn L., Santamaría E., Sangro B., Rodriguez-Ortigosa C., Fernández-Barrena M.G., Ávila M.A, & Berasain C. (2019). Splicing events in the control of genome integrity: role of SLU7 and truncated SRSF3 proteins. Nucleic Acids Research, 47(7), 3450-3466.

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RNA Immunoprecipitation Analysis

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Protein-A/G beads (Thermo Fisher Scientific) were resuspended by NT-2 buffer and incubated with anti-METTL3 (cat# ab195352, Abcam), anti-SRSF1 (cat# 32-4500, Thermo Fisher Scientific), anti-m6A (cat# ab208577, Abcam) antibodies at room temperature for 2 hours. Then, 100 μL cell lysates and 900 μL NET-2 (NET-2 buffer was consists of NT-2 buffer, EDTA, and DTT regents) buffer were added to eppendorf (EP) tubes containing Protein-A/G beads. The mixture was rotated and incubated overnight at 4°C. The immunoprecipitation was collected and purified by proteinase K and the expression of RNA was analyzed by RT-qPCR.

Tian Y., Xiao Y.H., Sun C., Liu B, & Sun F. (2023). N6-Methyladenosine Methyltransferase METTL3 Alleviates Diabetes-Induced Testicular Damage through Modulating TUG1/Clusterin Axis. Diabetes & Metabolism Journal, 47(2), 287-300.

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Western Blot Analysis of UPF1 Proteins

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Cells were lysed in RIPA lysis buffer (25 mM Tris-HCl, pH 7.6, 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, and 0.1% SDS). Fifteen microgram of proteins were loaded and separated by 8% (for UPF1 proteins) or 12% SDS-PAGE gel, then transferred to PVDF membrane (Millipore, USA). The following antibodies were used: Anti-UPF1 antibody-A (Cell Signaling Technology, USA), anti-UPF1 antibody-B (a gift from Jens Lykke-Andersen), anti-SRSF1 (Thermo Fisher Scientific, USA), anti-Lamin B1 (Abcam, UK), anti-α-tubulin (Abcam) and anti-Histone H3 (Abcam) antibodies. Horseradish peroxidase-conjugated anti-rabbit or anti-mouse IgGs (Thermo Fisher Scientific) were used as secondary antibodies.

Hong D., Park T, & Jeong S. (2019). Nuclear UPF1 Is Associated with Chromatin for Transcription-Coupled RNA Surveillance. Molecules and Cells, 42(7), 523-529.

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Identifying LINC00689 RNA-Binding Proteins

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RIP was performed to confirm the potential RNA-binding proteins of LINC00689 by a Magna RIP RNA-Binding Protein Immunoprecipitation Kit (Millipore) according to the manufacturer’s protocol. Briefly, cell lysis was conducted using RIP lysis buffer containing RNase and proteinase inhibitor, and then incubated with anti-U2AF2 (1:50, #70471, Cell Signaling Technology, Beverly, MA, USA), anti-SRSF1 (1:150, #32-4500, Thermo Fisher Scientific, Waltham, MA, USA), anti-PTBP1 (1:50, #32-4800, Thermo Fisher Scientific), anti-UPF1 (1:50, #9435, Cell Signaling Technology), or anti-IGF2BP2 (1:100, ab128175, Abcam) controlled by normal rabbit IgG (1:100, ab172730, Abcam) at 4°C overnight. After treatment with proteinase K buffer, the immunoprecipitated RNAs were extracted using the RNeasy MinElute Cleanup Kit (Qiagen, Duesseldorf, Germany), and then the enrichment of LINC00689 was measured by RT-qPCR.

Cao Y., Li J., Zhang G., Fang H., Du Y, & Liang Y. (2024). KLF15 transcriptionally activates LINC00689 to inhibit colorectal cancer development. Communications Biology, 7, 130.

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Protein Extraction and Immunoblotting

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Tissue was homogenized in Triton Lysis Buffer (12.5 mM HEPES pH 7.4, 50 mM NaCl, 500 μM EDTA, 5% glycerol, 0.5% Triton X-100, 50 mM sodium vanadate, 50 mM PMSF, 5 mM aprotinin, 5 mM, Leupeptin) using a TissueLyser II Homogenizer (Qiagen) before undergoing centrifugation at 10 000 rpm for 10 min at 4°C. The supernatant containing the protein lysate was transferred to a new tube and protein quantified with Pierce BCA Protein Assay Kit (Thermo Fisher Scientific) and analysed by western blot by incubating with anti-SRSF10 (Cell Signalling), anti-SRSF1 (Thermofisher), anti-SRSF5 (Atlas), anti-PPARα (Abcam), anti-Lamin (Abcam), anti-actin (Santa Cruz) and anti-Tubulin (Santa Cruz sc-5286) primary antibodies and imaged with an Odyssey infra-red scanner (LICOR).

Jobbins A.M., Haberman N., Artigas N., Amourda C., Paterson H.A., Yu S., Blackford S.J., Montoya A., Dore M., Wang Y.F., Sardini A., Cebola I., Zuber J., Rashid S.T., Lenhard B, & Vernia S. (2022). Dysregulated RNA polyadenylation contributes to metabolic impairment in non-alcoholic fatty liver disease. Nucleic Acids Research, 50(6), 3379-3393.

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Western Blot Analysis of Signaling Proteins

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Total protein was extracted from cells using RIPA lysis buffer (Beyotime Institute of Biotechnnology) and quantified using a BCA protein assay kit (Beyotime Institute of Biotechnology). Proteins (40 µg per lane) were separated via 10% SDS-PAGE, then transferred onto PVDF membranes (Bio-Rad Laboratories, Inc.). After blocking with 5% skimmed milk for 1 h at room temperature, the membranes were incubated with the following primary antibodies at 4˚C overnight: Anti-AKT (cat. no. ab18785; 1:1,000), anti-phosphorylated (p)-AKT (cat. no. ab38449; 1:1,000), Anti-p-ERK (cat. no. ab201015; 1:1,000), anti-ERK (cat. no. ab32081; 1:1,000), anti-MST1R (cat. no. ab52927; 1:1,000), anti-cleaved caspase-3 (cat. no. ab32042; 1:1,000), anti-SRSF1 (Thermo Fisher Scientific, Inc.; cat. no. 32-4500, 1:1,000) and anti-β-actin (cat. no. ab8226; 1:1,000; all Abcam except for SRSF1). Following the primary antibody incubation, the membranes were incubated with an anti-rabbit secondary antibody (HRP-conjugated; Abcam; cat. no. ab7090, 1:5,000) at room temperature for 1 h. Enhanced chemiluminescence reagent (Thermo Fisher Scientific, Inc.) was used to visualize the protein bands. ImageJ software (version 2.0; National Institutes of Health) was used to quantify the intensity of the bands.

Zhou D., Zhu X., Wu X., Zheng J., Tou L, & Zhou Y. (2021). The effect of splicing MST1R in gastric cancer was enhanced by lncRNA FENDRR. Experimental and Therapeutic Medicine, 22(2), 798.

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Antibody Sources and Reagents

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Antibodies were obtained from the following sources; anti-MAP2, anti-beta-tubulin, and anti-synaptophysin from Cell Signaling Inc. (Beverly, MA), β-tubulin from LI-COR, Odyssey (Lincoln, NE), anti-SRSF1 from Invitrogen, anti-SRSF2 from Sigma Aldrich, anti-MCL1, anti-SRSF3, anti-SRSF4, and anti-HNRNP A1 from Santa Cruz Biotech, and anti-Grb2 from BD Biosciences. Mammalian protease inhibitors were obtained from Sigma-Aldrich (St Louis, MO). Bradford reagent was from BioWorld (Dublin, OH). MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) was obtained from Thermo Fisher Scientific (#M6494).

Sariyer R., De Simone F.I., Donadoni M., Hoek J.B., Chang S.L, & Sariyer I.K. (2017). Alcohol-mediated missplicing of Mcl-1 pre-mRNA is involved in neurotoxicity. Alcoholism, clinical and experimental research, 41(10), 1715-1724.

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Targeted Gene Silencing with siRNA and 2'O-methyl RNA

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The following siRNA were from Dharmacon, GE Healthcare; siRNA 1; 5′-CGUGGAGUUUGUAC GGAAA-3′; siRNA 2- 5′-UGACCUAUGCAGUUC GAAA-3′; siRNA 3: 5′-UCUCGAAGCCGUAGUCG UA-3′; siRNA 4: 5′-CAGGAUUCAUGGAGCGGG A-3′. The following 2′O-methyl RNA were from Eurofins Genomics; UTR1: 5′-CUUCCUCCUGCCCGG-3′; UTR3: 5′-GAAACCCUGAGGGAG-3′. The following antibodies were used for immunoblotting and /or immunohistochemistry: anti-SRSF1 (Invitrogen); anti- α-tubulin (Fischer scientific); anti-ERK1/2 (Santa Cruz Biotechnology); anti-phospho ERK1/2 (Sigma). Cells were transfected with siRNA (0.2 μmol/L) or 2′O-methyl RNA (1 μmol/L) by using oligofectamine (Thermo Fischer Scientific).

Guyot M., Hilmi C., Ambrosetti D., Merlano M., Nigro C.L., Durivault J., Grépin R, & Pagès G. (2016). Targeting the pro-angiogenic forms of VEGF or inhibiting their expression as anti-cancer strategies. Oncotarget, 8(6), 9174-9188.

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Deoxynivalenol and Bortezomib Signaling Assay

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Deoxynivalenol (catalog # D0156) and cyanogen bromide-activated sepharose (catalog # S9142-1) are from Sigma-Aldrich. Bortezomib (catalog # ab142123) is from Abcam, and oprozomib (catalog # S7049) is from SelleckChem.
siRNAs that targeted host proteins were from the OnTARGETplus series from Horizon Discovery: siGRP78 (catalog # L-1008198-01), siRPLP1 (catalog # L-011135-00), siRPLP2 (catalog # L-004314-01), siSRSF1 (catalog # L-018672-01), siHNRNPA2B1 (L-011690-01), siDNAJB9 (catalog # L-012815-00), and siDNAJB12 (catalog # L-020585-00).
The majority of primary antibodies and horseradish peroxidase-conjugated secondary antibodies were from Abcam: anti-GRP78 (catalog # ab191023), anti-RPLP1 (catalog # ab121190), anti-RPLP2 (catalog # ab154958), anti-HNRNPA2B1(catalog # ab6102), anti-β-actin (catalog # ab8227), anti-GAPDH (catalog # ab8245), goat anti-rabbit-HRP (catalog # ab6721), and goat anti-mouse-HRP (catalog # ab6789). Anti-SRSF1 is from Invitrogen (catalog # 32-4600). Anti-HBsAg is from Fitzgerald Industries (catalog # 20-HR20). Anti-HBV polymerase is from Santa Cruz Biotech (catalog # Sc-81590). Anti-ubiquitin-HRP P4D1 is from Cytoskeleton (catalog # 140495).

Kao C.C., Nie Y., Ren S., De Costa N.T., Pandey R.K., Hong J., Smith D.B., Symons J.A., Beigelman L, & Blatt L.M. (2021). Mechanism of action of hepatitis B virus S antigen transport-inhibiting oligonucleotide polymer, STOPS, molecules. Molecular Therapy. Nucleic Acids, 27, 335-348.

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Antibody Characterization for Splicing Factors

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Antibodies used in this study were anti-SRSF1 (32–4500, Invitrogen), anti-U1-snRNP 70K (U1-70K) (H111, kindly provided by Akila Mayeda, Division of Gene Expression Mechanism, Fujita Health University), anti-U1 snRNP C (U1C) (4H12, Sigma-Aldrich), anti-U1 snRNP A (U1A) (PA5-27474, Thermo Fisher Scientific Pierce), anti-His-tag (D293-1, Medical & Biological Laboratories) and anti-GAPDH (Sigma-Aldrich).

Rahman M.A., Azuma Y., Nasrin F., Takeda J.I., Nazim M., Ahsan K.B., Masuda A., Engel A.G, & Ohno K. (2015). SRSF1 and hnRNP H antagonistically regulate splicing of COLQ exon 16 in a congenital myasthenic syndrome. Scientific Reports, 5, 13208.

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