Human Enterovirus 71 VP0 Insect Cell Lysate (WB positive control)

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Human Enterovirus 71 VP0 Insect Cell Lysate (WB positive control): 产品信息

产品描述
This EV71 Enterovirus 71 VP0 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of Enterovirus 71 VP0 protein (Cat: 40013-V20B) from the overexpression lysate was verified.
表达宿主
Baculovirus-Insect cells
种属
EV71
蛋白构建信息
A DNA sequence encoding the amino acids (Met 1-Gln 323) of human enterovirus 71 genome polyprotein (Q66478-1), corresponding to the protein VP0 precusor, a component of immature procapsids, was fused with the N-terminal polyhistidine-tagged GST tag at the N-terminus.
分子量
The recombinant human enterovirus 71 protein VP0/GST chimera consists of 560 amino acids and has a calculated molecular mass of 63 kDa. It migrates as an approximately 58 kDa band in SDS-PAGE under reducing conditions.

Human Enterovirus 71 VP0 Insect Cell Lysate (WB positive control): 使用指南

制备方法
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.
裂解缓冲液
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.
使用建议
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.
缓冲液
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.
应用
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human Enterovirus 71 VP0 Insect Cell Lysate (WB positive control): 别称

EV71 VP0 Overexpression Lysate; EV71 VP4-VP2 Overexpression Lysate

Enterovirus 71 VP0 背景信息

Human enterovirus 71 genome polyprotein is a member of the picornaviruses polyprotein family. It contains two peptidase C3 domains, one RdRp catalytic domain, one SF3 helicase domain. Genome polyprotein is cleaved into the following 12 chains: Protein VP (VP4-VP2), Protein VP4 (P1A), Protein VP2 (P1B), Protein VP3 (P1C), Protein VP1 (P1D), Picornain 2A (P2A), Protein 2B (P2B), Protein 2C (P2C), Protein 3A (P3A), Protein 3B (P3B), Picornain 3C (Protease 3C) and RNA-directed RNA polymerase 3D-POL (P3D-POL). VP precursor is a component of immature procapsids. Capsid proteins VP1, VP2, VP3 and VP4 form a closed capsid enclosing the viral positive strand RNA genome. VP4 lies on the inner surface of the protein shell formed by VP1, VP2 and VP3. All the three latter proteins contain a beta-sheet structure called beta-barrel jelly roll. Together they form an icosahedral capsid composed of 6 copies of each VP1, VP2, and VP3, with a diameter of approximately 3 Angstroms. VP1 is situated at the 12 fivefold axes, whereas VP2 and VP3 are located at the quasi-sixfold axes.
参考文献
  • Brown B.A., et al., 1995, Virus Res. 39:195-206.
  • Tang W.-F. et al., 2007, J. Biol. Chem. 282:5888-5898.
  • Huang,S.C. et al., 2008, Virus Res. 131 (2):250-9.
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