p62/SQSTM1 Overexpression Lysate
Novus Biologicals, part of Bio-Techne | Catalog # NBL1-16446
Key Product Details
Species
Human
Applications
Western Blot
Product Summary for p62/SQSTM1 Overexpression Lysate
p62/SQSTM1 Transient Overexpression Lysate
Expression Host: HEK293T
Plasmid: RC203214
Accession#: NM_003900
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.
Expression Host: HEK293T
Plasmid: RC203214
Accession#: NM_003900
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.
Product Specifications
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.
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.
TMW
47.5 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.
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.
Type
Overexpression
Scientific Data Images for p62/SQSTM1 Overexpression Lysate
![Western Blot: p62/SQSTM1 Overexpression Lysate [NBL1-16446]](https://resources.bio-techne.com/images/products/p62-SQSTM1-Overexpression-Lysate-Adult-Normal-Western-Blot-NBL1-16446-img0002.jpg "Western Blot: p62/SQSTM1 Overexpression Lysate [NBL1-16446]")
Western Blot: p62/SQSTM1 Overexpression Lysate [NBL1-16446]
Western Blot: p62/SQSTM1 Overexpression Lysate (Adult Normal) [NBL1-16446] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for p62/SQSTM1.Formulation, Preparation, and Storage
Formulation
RIPA buffer
Concentration
The exact concentration of the protein of interest cannot be determined for overexpression lysates. Please contact technical support for more information.
Shipping
The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store at -80C. Avoid freeze-thaw cycles.
Background: p62/SQSTM1
Abnormal function of p62/SQSTM1 is associated with a range of disease states such as neurodegeneration, cancer, and metabolic disorders (2). Mutations in the p62/SQSTM1 sequence have been linked to Paget's disease of the bone, amyotrophic lateral sclerosis, and frontotemporal lobar degeneration. In Parkinson's disease, p62/SQSTM1 has been linked to microglia activation and subsequent neuroinflammation (3). Functionally, p62/SQSTM1 is involved in a broad range of cellular processes such as amino acid sensing by interaction with mTORC1, oxidative stress response through interaction with Keap1, and targeting cargo for autophagy by interacting with ubiquitin labeled proteins (1).
To induce selective autophagy, p62/SQSTM1 forms long oligomers or helical filaments which interact with LC3 and ubiquitin labeled proteins and lead to the initiation of the autophagosome formation (2). p62/SQSTM1 is not only a selective autophagy receptor but also an autophagy substrate, as its engulfed by the autophagosome and degraded by the autophagolysosome. Monitoring LC3 levels is the standard for assessing autophagic flux, however monitoring p62/SQSTM1 levels by Western blot in the presence and absence of autophagy inhibitors (e.g., Chloroquine) is also a common practice (4). Besides its activity as a selective autophagy receptor, p62/SQSTM1 also plays a role as an adaptor in signaling cascades leading to NFkB activation downstream of TNF-R, IL-1 beta R, TrkA and p75NTR. Briefly, for NFkB signaling downstream of the TNF-R activation, p62/SQSTM1 engages RIP1 kinase and PKC iota/lambda through the ZZ and PB1 domains, respectively (5).
References
1.Katsuragi, Y., Ichimura, Y., & Komatsu, M. (2015). P62/SQSTM1 functions as a signaling hub and an autophagy adaptor. FEBS Journal. https://doi.org/10.1111/febs.13540
2. Sanchez-Martin, P., & Komatsu, M. (2018). p62/SQSTM1 - Steering the cell through health and disease. Journal of Cell Science. https://doi.org/10.1242/jcs.222836
3. Yao, L., Zhu, Z., Wu, J., Zhang, Y., Zhang, H., Sun, X., ... Lu, G. (2019). MicroRNA-124 regulates the expression of p62/p38 and promotes autophagy in the inflammatory pathogenesis of Parkinson's disease. The FASEB Journal. https://doi.org/10.1096/fj.201900363r
4 Klionsky, D. J., Abdelmohsen, K., Abe, A., Abedin, M. J., Abeliovich, H., Arozena, A. A., ... Zughaier, S. M. (2016). Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition). Autophagy. https://doi.org/10.1080/15548627.2015.1100356
5. Bitto, A., Lerner, C. A., Nacarelli, T., Crowe, E., Torres, C., & Sell, C. (2014). p62/SQSTM1 at the interface of aging, autophagy, and disease. Age. https://doi.org/10.1007/s11357-014-9626-3
Long Name
Sequestosome 1
Alternate Names
A170, EBIAP, ORCA, OSIL, PDB3, Sequestosome 1, SQSTM1, ZIP3
Gene Symbol
SQSTM1
Additional p62/SQSTM1 Products
Product Documents for p62/SQSTM1 Overexpression Lysate
Product Specific Notices for p62/SQSTM1 Overexpression Lysate
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.
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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