Mouse DKK3 HEK293 Overexpression Lysate

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Mouse DKK3 HEK293 Overexpression Lysate: Product Information

Product Description
This Mouse DKK3 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of DKK3 protein (Cat: 50247-M08H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Species
Mouse
Sequence Information
A DNA sequence encoding the extracellular domain (Met 1-Ile 349) of mouse DKK3 (NP_056629.1) precursor was expressed with a C-terminal polyhistidine tag.
Molecule Mass
The secreted recombinant mouse DKK3 consists of 338 amino acids and has a calculated molecular mass of 38 kDa. As a result of glycosylation, the recombinant protein migrates as an approximately 65 kDa protein in SDS-PAGE under reducing conditions.

Mouse DKK3 HEK293 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.

Mouse DKK3 HEK293 Overexpression Lysate: Alternative Names

Mouse AW061014 Overexpression Lysate; Mouse C87148 Overexpression Lysate

DKK3 Background Information

DKK3 (dickkopf related protein 3) is a member of the dickkopf-related family consisting of DKK1, DKK2, DKK3 and DKK4. It is a secreted protein, and also known as REIC (Reduced Expansion in Immortalized Cells). The DKK3 protein is proposed to function as a secreted tumor suppressor since it is downregulated in a number of cancer cells and prostate cancer tissue and may be a promising candidate molecule for therapeutic interference. DKK3 protein is also a negative regulator of beta-catenin and its downregulation contribute to an activation of the beta-catenin signaling pathway.
Full Name
dickkopf WNT signaling pathway inhibitor 3
References
  • Yue W, et al., 2008. Downregulation of Dkk3 activates beta-catenin/TCF-4 signaling in lung cancer. Carcinogenesis, 29 (1): 84-92.
  • Ding Z, et al., 2009. Promoter methylation and mRNA expression of DKK3 and WIF-1 in hepatocellular carcinoma. World J Gastroenterol, 15 (21):2595-601.
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