Human PHKG1 Baculovirus-Insect cells Overexpression Lysate

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Human PHKG1 Baculovirus-Insect cells Overexpression Lysate: Product Information

Product Description
This Human PHKG1 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of PHKG1 protein (Cat: 11034-H09B) from the overexpression lysate was verified.
Expression Host
Baculovirus-Insect cells
Species
Human
Sequence Information
A DNA sequence encoding the human PHKG1 (NP_006204.1) (Met 1-Tyr 387) was fused with the GST tag at the N-terminus.
Molecule Mass
The recombinant human PHKG1/GST chimera consists of 612 amino acids and predicts a molecular mass of 71.3 kDa. It migrates as an approximately 65 kDa band in SDS-PAGE under reducing conditions.

Human PHKG1 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 PHKG1 Baculovirus-Insect cells Overexpression Lysate: Alternative Names

Human PHKG Overexpression Lysate

PHKG1 Background Information

Phosphorylase b kinase gamma catalytic chain, skeletal muscle isoform, also known as Phosphorylase kinase subunit gamma-1 and PHKG1, is a member of the protein kinase superfamily and CAMK Ser/Thr protein kinase family. PHKG1 is the catalytic member of a 16 subunit protein kinase complex which contains equimolar ratios of 4 subunit types. The complex is a crucial glycogenolytic regulatory enzyme. Muscle glycogenosis caused by phosphorylase kinase (Phk) deficiency may lead to exercise intolerance, weakness and musculatur atrophy. The gene encoding the muscle isoform of the Phk gamma subunit (gamma M) is one of the candidate genes in which mutations responsible for this condition should be sought. Muscle-specific deficiency of Phk causes glycogen storage disease, clinically manifesting in exercise intolerance with early fatiguability, pain, cramps and occasionally myoglobinuria.
Full Name
phosphorylase kinase, gamma 1 (muscle)
References
  • Wehner, M. et al., 1995, Hum Genet. 96 (5): 616-8.
  • Burwinkel, B. et al., 2003, Eur J Hum Genet. 11 (7): 516-26.
  • Winchester, JS. et al., 2007, Mol Genet Metab. 92 (3): 234-42.
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