Human PDE9A Baculovirus-Insect cells Overexpression Lysate

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Human PDE9A Baculovirus-Insect cells Overexpression Lysate: Product Information

Product Description
This Human PDE9A overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of PDE9A protein (Cat: 11745-H20B) from the overexpression lysate was verified.
Expression Host
Baculovirus-Insect cells
Species
Human
Sequence Information
A DNA sequence encoding the full length of human PDE9A (O76083-2) (Met 1-Ala 533) was fused with the N-terminal polyhistidine-tagged GST tag at the N-terminus.
Molecule Mass
The recombinant human PDE9A consisting of 770 amino acids and has a calculated molecular mass of 89.5 kDa. It migrates as an approximately 75 kDa band in SDS-PAGE under reducing conditions.

Human PDE9A Baculovirus-Insect cells Overexpression Lysate: Usage Guide

Preparation Method
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
1.  Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2.  Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human PDE9A Baculovirus-Insect cells Overexpression Lysate: Alternative Names

Human HSPDE9A2 Overexpression Lysate

PDE9A Background Information

High affinity cGMP-specific 3',5'-cyclic phosphodiesterase 9A, also known as PDE9A, is a member of the cyclic nucleotide phosphodiesterase family and PDE9 subfamily. PDE9A is expressed in all tissues examined (testis, brain, small intestine, skeletal muscle, heart, lung, thymus, spleen, placenta, kidney, liver, pancreas, ovary and prostate) except blood. Highest levels of PDE9A is in brain, heart, kidney, spleen, prostate and colon. Isoform PDE9A2 is found in prostate. PDE9A mRNA is widely distributed throughout the rat and mouse brain, with the highest expression observed in cerebellar Purkinje cells. PDE9A is the only cGMP-specific PDE with significant expression in the forebrain, and as such is likely to play an important role in NO-cGMP signaling. PDE9A is highly conserved between species and is widely distributed throughout the rodent brain. PDE9A is probably involved in maintenance of low cGMP levels in cells and might play an important role in a variety of brain functions involving cGMP-mediated signal transduction. PDE9A hydrolyzes the second messenger cGMP, which is a key regulator of many important physiological processes. PDE9A represents a novel drug target worthy of further study.
Full Name
phosphodiesterase 9A
References
  • Andreeva, SG. et al., 2001,J Neurosci. 21 (22):9068-76.
  • van Staveren,WC. et al., 2002, J Neurocytol. 31 (8-9):729-41.
  • Almeida,CB. et al., 2008, Br J Haematol. 142 (5):836-44.
  • Kruse,L.S. et al., 2009, Brain Res. 1281 :25-34.
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