Phosphodiesterase 4A/PDE4A Overexpression Lysate Summary
Description |
Phosphodiesterase 4A/PDE4A Transient Overexpression Lysate Expression Host: HEK293T
Plasmid: RC226235
Accession#: NM_001111307
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 |
PDE4A |
Applications/Dilutions
Dilutions |
|
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 |
98 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
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 Phosphodiesterase 4A/PDE4A Overexpression Lysate
Background
Enzymes of the cAMP-dependent phosphodiesterase type 4 (PDE4) family are important in hydrolyzing cAMP produced by G-protein coupled receptor (GPCR) stimulated adenylyl cyclases. In brain, more than 90% of cAMP formed by the stimulation of GPCRs is hydrolyzed by PDE4 enzymes. PDE4 enzymes are also important molecular targets for a variety of therapeutic agents like antidepressants, anti-asthmatics, and anti-inflammatory drugs. PDE4 family comprises 4 genes (PDE4A, B, C and D); each exhibiting multiple isozymes due to alternate splicing that leads to a larger number of distinct PDE4 variants. Members of the PDE4 family are regulated/activated by phosphorylation/dephosphorylation by cAMP-dependent protein kinase A and phosphatases. Protein-protein interactions and cellular trafficking of PDE4A enzymes play an important role in cAMP compartmentalization and cAMP-dependent signaling. In brain members of the PDE4A, B and D family are associated with GPCRs (adrenergic and dopaminergic) signaling.
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.
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