Sphingosine Kinase 1/SPHK1 Overexpression Lysate

Western Blot: Sphingosine Kinase 1 Overexpression Lysate (Adult Normal) [NBP2-06490] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Sphingosine Kinase 1.

• Reactivity: Hu
• Applications: WB

Sphingosine Kinase 1/SPHK1 Overexpression Lysate Summary

• Description:

  Sphingosine Kinase 1/SPHK1 Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC203398
  
  Accession#: NM_021972
  
  Protein Tag: C-MYC/DDK
  
  You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
• Gene: SPHK1

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.
  
  Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.
• Theoretical MW: 43.8 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for Sphingosine Kinase 1/SPHK1 Overexpression Lysate

• EC 2.7.1.91
• SK1
• sphingosine kinase 1
• SPHK1
• SPHKSK 1
• SPK 1
• SPK
• SPK1

Background

Sphingosine-1-phosphate (SPP) is a novel lipid messenger with both intracellular and extracellular functions. Intracellularly, it regulates proliferation and survival, and extracellularly, it is a ligand for EDG1 (MIM 601974). Various stimuli increase cellular levels of SPP by activation of sphingosine kinase (SPHK), the enzyme that catalyzes the phosphorylation of sphingosine. Competitive inhibitors of SPHK block formation of SPP and selectively inhibit cellular proliferation induced by a variety of factors, including platelet-derived growth factor (e.g., MIM 173430) and serum.[supplied by OMIM]

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Bioinformatics

• Gene Symbol: SPHK1