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人 Epcr/PROCR 基因ORF全长cDNA克隆(表达载体), C-Flag 标签

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Human PROCR cDNA Clone产品信息
NCBI注册码:BC014451
参考序列ORF长度:717bp
cDNA基因描述:Full length Clone DNA of Homo sapiens protein C receptor, endothelial (EPCR) with C terminal Flag tag.
分子别称:CCCA, EPCR, CCD41, CD201, bA42O4.2, PROCR
分子种属:Human
载体:pCMV3-C-FLAG
质粒:pCMV3-PROCR-Flag
限制性酶切位点:HindIII + NotI (6kb + 0.77kb)
Tag序列:FLAG Tag Sequence: GATTACAAGGATGACGACGATAAG
序列信息:Identical with the Gene Bank Ref. ID sequence.
测序引物:T7(TAATACGACTCACTATAGGG) BGH(TAGAAGGCACAGTCGAGG)
启动子:Enhanced CMV mammalian cell promoter
Application:Stable or Transient mammalian expression
抗生素(大肠杆菌):Kanamycin
抗生素(哺乳动物细胞):Hygromycin
运输方式:Each tube contains lyophilized plasmid.
储存方法:The lyophilized plasmid can be stored at room temperature for three months.
Human PROCR 基因质粒图
Human PROCR Gene cDNA Clone (full-length ORF Clone), expression ready, C-FLAG-tagged
Human PROCR 基因表达验证图
Human PROCR ORF mammalian expression plasmid, C-Flag tag
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Caption:
The plasmid was transfected into 293H adherent cells with Sinofection reagent (Cat# STF01). After 48 h, Immunofluorescence staining of cells. Cells were fixed with 4% PFA, permeabilzed with 0.3% Triton X-100 in PBS, blocked with 10% serum, and incubated with Mouse anti-Flag Tag monoclonal antibody (CST#8146S) at 37℃ 1 hour. Then cells were stained with Goat Anti-mouse IgG secondary antibody. The fluorescent signal is detected by fluorescence microscope. Each expression experiment has negative control.
FLAG Tag Info

FLAG-tag, or FLAG octapeptide, is a polypeptide protein tag that can be added to a protein using recombinant DNA technology. It can be used for affinity chromatography, then used to separate recombinant, overexpressed protein from wild-type protein expressed by the host organism. It can also be used in the isolation of protein complexes with multiple subunits.

A FLAG-tag can be used in many different assays that require recognition by an antibody. If there is no antibody against the studied protein, adding a FLAG-tag to this protein allows one to follow the protein with an antibody against the FLAG sequence. Examples are cellular localization studies by immunofluorescence or detection by SDS PAGE protein electrophoresis.

The peptide sequence of the FLAG-tag from the N-terminus to the C-terminus is: DYKDDDDK (1012 Da). It can be used in conjunction with other affinity tags, for example a polyhistidine tag (His-tag), HA-tag or Myc-tag. It can be fused to the C-terminus or the N-terminus of a protein. Some commercially available antibodies (e.g., M1/4E11) recognize the epitope only when it is present at the N-terminus. However, other available antibodies (e.g., M2) are position-insensitive.

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研究背景

Endothelial protein C receptor (EPCR), also known as activated protein C receptor (APC receptor) or PROCR, is a receptor for Protein C. Protein C plays an important role in many metabolism processes in humans and other animals after activated by binding to Endothelial protein C receptor (EPCR). Because of the EPCR is found primarily on endothelial cells (cells on the inside of blood vessels), activated protein C is found maily near endothelial cells. Protein C is pleiotropic, with two main functions: anticoagulation and cytoprotection. Which function will be performed depend on whether or not protein C remains bind to EPCR after activated. The anticoagulation occurs when it does not. In this case, protein C functions as an anticoagulant by irreversibly proteolytically inactivating Factor Va and Factor VIIIa, turning them into Factor Vi and Factor VIIIi respectively. When still bound to EPCR, activated protein C performs its cytoprotective effects, acting on the effector substrate PAR-1, protease-activated receptor-1. To a degree, APC's anticoagulant properties are independent of its cytoprotective ones, in that expression of one pathway is not affected by the existence of the other. 

参考资料
  • Nicolaes GA, et al. (2003). Congenital and acquired activated protein C resistance. Semin Vasc Med. 3 (1): 33-46.
  • Esmon CT. ( 2003). The protein C pathway. Chest 124 (3): 26-32.
  • Mosnier LO, et al. (2007)The cytoprotective protein C pathway. Blood. 109: 3161-72.
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    货号: HG13320-CF
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    • Human PROCR Gene cDNA Clone (full-length ORF Clone), expression ready, C-FLAG-tagged
    • Human PROCR ORF mammalian expression plasmid, C-Flag tag
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