Human IRE1 Baculovirus-Insect cells Overexpression Lysate


Human IRE1 Baculovirus-Insect cells Overexpression Lysate: 产品信息

This Human IRE1 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of IRE1 protein (Cat: 11905-HNCB) from the overexpression lysate was verified.
Baculovirus-Insect cells
A DNA sequence encoding the human ERN1 (O75460-1) (Pro 465-Leu 977) was expressed and purified with two additional amino acids (Gly & Pro ) at the N-terminus.
The secreted recombinant human ERN1 consists of 515 amino acids and predicts a molecular mass of 58.3 KDa. The apparent molecular mass of the protein is approximately 65 KDa in SDS-PAGE under reducing conditions due to glycosylation.

Human IRE1 Baculovirus-Insect cells Overexpression Lysate: 使用指南

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.
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.
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.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
稳定性 & 储存条件
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.
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human IRE1 Baculovirus-Insect cells Overexpression Lysate: 别称

Human hIRE1p Overexpression Lysate; Human IRE1 Overexpression Lysate; Human IRE1a Overexpression Lysate; Human IRE1P Overexpression Lysate

IRE1 背景信息

Endoplasmic reticulum stress and hypoxia are necessary components of malignant tumors growth and suppression of ERN1 (from endoplasmic reticulum to nuclei-1) signalling pathway, which is linked to the apoptosis and cell death processes, significantly decreases proliferative processes. An enhanced expression of TP53 gene in ERN1 knockdown glioma cells correlates with the decreased level of ubiquitin ligase MDM2 and increased expression level of USP7 which deubiquitinates TP53 and MDM2 and induces TP53-dependent cell growth repression and apoptosis. Thus, the expression of genes encoding TP53 and related to TP53 factors depends upon the endoplasmic reticulum stress signaling as well as on hypoxia, and correlates with suppression of glioma growth under ERN1 knockdown. The dependence of insulin-like growth binding proteins as well as IGF2BP3 and HTRA1 gene expressions in U87 glioma cells on ERN1 signaling enzyme function and hypoxia, indicating its participation in the regulation of metabolic and proliferative processes via IGF/INS receptors, because endoplasmic reticulum stress is an important component of tumor growth and metabolic diseases.
endoplasmic reticulum to nucleus signaling 1
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  • Oppermann F.S., et al., 2009, Mol. Cell. Proteomics 8:1751-64.
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