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Human RHEB Insect Cell Lysate (WB positive control)

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Human RHEB Transfected / Overexpression Cell Lysate Product Information
Expressed Host:Baculovirus-Insect cells
Product Description:Baculovirus-Insect Cell lysate that Human RHEB transfected / overexpressed for Western blot (WB) positive control. The whole cell lysate is provided in 1X Sample Buffer (1X modified RIPA buffer+1X SDS loading buffer).
Sequence information:A DNA sequence encoding the human RHEB (Q15382)(Met1-Cys181) was fused with a polyhistide tag at the N-terminus.
Predicted N Terminal:His
Molecule Mass:The recombinant human RHEB consists of 199 amino acids and has a calculated molecular mass of 22.4 kDa. The recombinant protein migrates as an approximately 21 kDa band in SDS-PAGE under reducing conditions.
Human RHEB Transfected / Overexpression Cell Lysate Usage Guide
Preparation Method:Cell lysate was prepared by homogenization 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.
Quality Control Testing:12.5% SDS-PAGE Stained with Coomassie Blue after protein purification.
Stability:Samples are stable for up to twelve months from date of receipt.
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. 3. Store the lyophilized cell lysate at 4℃. After re-dissolution, recommend to aliquot it into smaller quantities and store at -80℃.
Storage Buffer:1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Storage Instruction:Store at 4℃. After re-dissolution, aliquot and store at -80℃.
Application notes:Western blot (WB): Use at an assay dependent dilution.
Other Applications: Not tested.
Optimal dilutions/concentrations should be determined by the end user.
RHEB / RHEB2 Background

RHEB is a recently discovered member of the Ras superfamily that may be involved in neural plasticity. This function is novel and not typically associated with the Ras proteins. RHEB gene is a member of the small GTPase superfamily and encodes a lipid-anchored, cell membrane protein with five repeats of the RAS-related GTP-binding region. RHEB is vital in regulation of growth and cell cycle progression due to its role in the insulin / TOR / S6K signaling pathway. The protein has GTPase activity and shuttles between a GDP-bound form and a GTP-bound form, and farnesylation of RHEB is required for this activity. Three pseudogenes have been mapped, two on chromosome 10 and one on chromosome 22.

Human RHEB / RHEB2 References
  • Karbowniczek, et al. (2004) Regulation of B-Raf kinase activity by tuberin and Rheb is mammalian target of rapamycin (mTOR)-independent. J Biol Chem. 279(29):29930-7.
  • Long, et al. (2005) Rheb binds and regulates the mTOR kinase. Curr Biol. 15(8):702-13.
  • Mizuki N, et al. (1996) Isolation of cDNA and genomic clones of a human Ras-related GTP-binding protein gene and its chromosomal localization to the long arm of chromosome 7, 7q36. Genomics. 34(1):114-8.
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    Catalog: 11382-H07BL-300
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