Phe substitution from 14 of the identified substrates. Using this platform, we identified 34 peptides that are phosphorylated by c-Src. We have demonstrated that SILAC-based quantitative proteomics approach is suitable for identification of substrates of nonreceptor tyrosine kinases and can be coupled with peptide microarrays for high-throughput identification of substrate phosphopeptides.", "Category"=>"Primary Antibody", "Clonality"=>"Polyclonal", "Concentration"=>"1ug/ul", "Conjugation"=>"Unconjugated", "Cross Reactive Species"=>"Human, Mouse, Rat", "Datasheet URL"=>"https://www.biossusa.com/store/datasheets/bs-1135R", "Gene ID"=>"6714", "Gene ID Link"=>"Entrez Gene:
6714", "Host"=>"Rabbit", "images"=>"[{\"id\":\"15926\",\"status\":\"active\",\"visibility\":\"public\",\"origin_id\":\"1\",\"origin_notes\":\"\",\"product_id\":\"2139\",\"productconjugation_id\":null,\"application_id\":\"4\",\"application_note\":\"\",\"dilution\":\"1:400\",\"cross_reactive_species_id\":\"3\",\"cross_reactive_species_note\":\"\",\"description\":\"Paraformaldehyde-fixed, paraffin embedded rat brain; Antigen retrieval by boiling in sodium citrate buffer (pH6) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 30 minutes; Blocking buffer (normal goat serum) at 37\\u00b0C for 20min; Antibody incubation with SRC Polyclonal Antibody, Unconjugated (bs-1135R) at 1:400 overnight at 4\\u00b0C, followed by a conjugated secondary and DAB staining.\",\"sample_type_id\":\"2\",\"sample_details\":null,\"rating\":\"3\",\"disease_id\":null,\"sample_tissue_id\":\"2\",\"cell_line_id\":null,\"protocols_used\":\"Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\\u00b0C for 30min; Antibody incubation with (SRC) Polyclonal Antibody, Unconjugated (bs-1135R) at 1:400 overnight at 4\\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.\",\"additional_notes\":\"\",\"filepath_raw\":\"2139\\/15926_raw.jpg\",\"filepath_med\":\"2139\\/15926_med.jpg\",\"filepath_thumb\":\"2139\\/15926_thumb.jpg\",\"display_order\":\"1\",\"lock_version\":\"1\",\"created_at\":\"2017-04-17 17:35:30\",\"updated_at\":\"2017-04-17 17:35:31\",\"image_url\":\"http:\\/\\/biossantibodies.com\\/image-raw\\/15926\"},{\"id\":\"15927\",\"status\":\"active\",\"visibility\":\"public\",\"origin_id\":\"1\",\"origin_notes\":\"\",\"product_id\":\"2139\",\"productconjugation_id\":null,\"application_id\":\"4\",\"application_note\":\"\",\"dilution\":\"1:400\",\"cross_reactive_species_id\":\"2\",\"cross_reactive_species_note\":\"\",\"description\":\"Paraformaldehyde-fixed, paraffin embedded mouse testis; Antigen retrieval by boiling in sodium citrate buffer (pH6) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 30 minutes; Blocking buffer (normal goat serum) at 37\\u00b0C for 20min; Antibody incubation with SRC Polyclonal Antibody, Unconjugated (bs-1135R) at 1:400 overnight at 4\\u00b0C, followed by a conjugated secondary and DAB staining.\",\"sample_type_id\":\"2\",\"sample_details\":null,\"rating\":\"2\",\"disease_id\":null,\"sample_tissue_id\":\"20\",\"cell_line_id\":null,\"protocols_used\":\"Paraformaldehyde-fixed, paraffin embedded rat brain; Antigen retrieval by boiling in sodium citrate buffer (pH6) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 30 minutes; Blocking buffer (normal goat serum) at 37\\u00b0C for 20min; Antibody incubation with SRC Polyclonal Antibody, Unconjugated (bs-1135R) at 1:400 overnight at 4\\u00b0C, followed by a conjugated secondary and DAB staining.\",\"additional_notes\":\"\",\"filepath_raw\":\"2139\\/15927_raw.jpg\",\"filepath_med\":\"2139\\/15927_med.jpg\",\"filepath_thumb\":\"2139\\/15927_thumb.jpg\",\"display_order\":\"2\",\"lock_version\":\"1\",\"created_at\":\"2017-04-17 17:36:21\",\"updated_at\":\"2017-04-17 17:36:21\",\"image_url\":\"http:\\/\\/biossantibodies.com\\/image-raw\\/15927\"}]", "Immunogen Sequence"=>"2-100/541", "Isotype"=>"IgG", "Lead Time"=>"3 to 5 business days.", "Modification"=>"Unmodified", "Predicted Reactivity"=>"Dog, Cow", "Product URL"=>"https://www.biossusa.com/products/bs-1135R", "Purification"=>"Purified by Protein A.", "Size"=>"100ul", "Source"=>"KLH conjugated synthetic peptide derived from human SRC", "Storage"=>"0.01M TBS(pH7.4) with 1% BSA, 0.02% Proclin300 and 50% Glycerol. Shipped at 4°C. Store at -20°C for one year. Avoid repeated freeze/thaw cycles.", "Storage Buffer"=>"0.01M TBS(pH7.4) with 1% BSA, 0.02% Proclin300 and 50% Glycerol.", "Storage Condition"=>"Shipped at 4°C. Store at -20°C for one year. Avoid repeated freeze/thaw cycles.", "Subcellular Locations"=>"Cytoplasm, Nucleus, Cell membrane", "Swiss Prot"=>"P12931", "Swiss Prot Link"=>"Swiss Prot:
P12931", "Synonyms"=>"ASV; Avian sarcoma virus; c SRC; CDNA FLJ14219 fis clone NT2RP3003800 highly similar to Rattus norvegicus tyrosine protein kinase pp60 c src mRNA; cSrc; EC 2.7.10.2; Neuronal CSRC tyrosine specic protein kinase; Neuronal SRC; Oncogene SRC; OTTHUMP00000030931; OTTHUMP00000174476; OTTHUMP00000174477; p60 Src; p60-Src; p60Src; pp60c src; pp60c-src; pp60csrc; Proto oncogene tyrosine protein kinase Src; Proto-oncogene c-Src; Proto-oncogene tyrosine-protein kinase Src; Protooncogene SRC; protooncogene SRC Rous sarcoma; Src; SRC Oncogene; SRC_MOUSE; SRC1; Tyrosine kinase pp60c src; Tyrosine protein kinase SRC 1; Tyrosine protein kinase SRC1; v src avian sarcoma Schmidt Ruppin A2 viral oncogene homolog; V src sarcoma Schmidt Ruppin A 2 viral oncogene homolog avian; v src sarcoma Schmidt Ruppin A 2 viral oncogene homolog avian; vsrc avian sarcoma Schmidt Ruppin A2 viral oncogene homolog; SRC_HUMAN.", "Target Protein"=>"SRC"} -->
| Overview |
| bs-1135R |
| SRC Polyclonal Antibody |
| WB, ELISA, IHC-P, IHC-F, IF(IHC-P), IF(IHC-F), IF(ICC) |
| Human, Mouse, Rat |
| Dog, Cow |
| Specifications |
| Unconjugated |
| Rabbit |
| KLH conjugated synthetic peptide derived from human SRC |
| 2-100/541 |
| Polyclonal |
| IgG |
| 1ug/ul |
| Purified by Protein A. |
| 0.01M TBS(pH7.4) with 1% BSA, 0.02% Proclin300 and 50% Glycerol. |
| Shipped at 4°C. Store at -20°C for one year. Avoid repeated freeze/thaw cycles. |
| Target |
| 6714 |
| P12931 |
| Cytoplasm, Nucleus, Cell membrane |
| ASV; Avian sarcoma virus; c SRC; CDNA FLJ14219 fis clone NT2RP3003800 highly similar to Rattus norvegicus tyrosine protein kinase pp60 c src mRNA; cSrc; EC 2.7.10.2; Neuronal CSRC tyrosine specic protein kinase; Neuronal SRC; Oncogene SRC; OTTHUMP00000030931; OTTHUMP00000174476; OTTHUMP00000174477; p60 Src; p60-Src; p60Src; pp60c src; pp60c-src; pp60csrc; Proto oncogene tyrosine protein kinase Src; Proto-oncogene c-Src; Proto-oncogene tyrosine-protein kinase Src; Protooncogene SRC; protooncogene SRC Rous sarcoma; Src; SRC Oncogene; SRC_MOUSE; SRC1; Tyrosine kinase pp60c src; Tyrosine protein kinase SRC 1; Tyrosine protein kinase SRC1; v src avian sarcoma Schmidt Ruppin A2 viral oncogene homolog; V src sarcoma Schmidt Ruppin A 2 viral oncogene homolog avian; v src sarcoma Schmidt Ruppin A 2 viral oncogene homolog avian; vsrc avian sarcoma Schmidt Ruppin A2 viral oncogene homolog; SRC_HUMAN. |
| c-Src tyrosine kinase plays a critical role in signal transduction downstream of growth factor receptors, integrins and G protein-coupled receptors. We used stable isotope labeling with amino acids in cell culture (SILAC) approach to identify additional substrates of c-Src tyrosine kinase in human embryonic kidney 293T cells. We have identified 10 known substrates and interactors of c-Src and Src family kinases along with 26 novel substrates. We have experimentally validated 4 of the novel proteins (NICE-4, RNA binding motif 10, FUSE-binding protein 1 and TRK-fused gene) as direct substrates of c-Src using in vitro kinase assays and cotransfection experiments. Significantly, using a c-Src specific inhibitor, we were also able to implicate 3 novel substrates (RNA binding motif 10, EWS1 and Bcl-2 associated transcription factor) in PDGF signaling. Finally, to identify the exact tyrosine residues that are phosphorylated by c-Src on the novel c-Src substrates, we designed custom peptide microarrays containing all possible tyrosine-containing peptides (312 unique peptides) and their mutant counterparts containing a Tyr --> Phe substitution from 14 of the identified substrates. Using this platform, we identified 34 peptides that are phosphorylated by c-Src. We have demonstrated that SILAC-based quantitative proteomics approach is suitable for identification of substrates of nonreceptor tyrosine kinases and can be coupled with peptide microarrays for high-throughput identification of substrate phosphopeptides. |
| Application Dilution |
| WB |
1:300-5000 |
| ELISA |
1:500-1000 |
| IHC-P |
1:200-400 |
| IHC-F |
1:100-500 |
| IF(IHC-P) |
1:50-200 |
| IF(IHC-F) |
1:50-200 |
| IF(ICC) |
1:50-200 |
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