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人 PLBD2 基因ORF全长cDNA克隆(表达载体), C-Flag 标签

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Human PLBD2 cDNA Clone产品信息
NCBI注册码:NM_173542.3
参考序列ORF长度:1770bp
cDNA基因描述:Full length Clone DNA of Homo sapiens phospholipase B domain containing 2 with C terminal Flag tag.
分子别称:P76
分子种属:Human
载体:pCMV3-C-FLAG
质粒:
限制性酶切位点:
Tag序列:FLAG Tag Sequence: GATTACAAGGATGACGACGATAAG
序列信息:
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.
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

PLBD2 localizes to the lysosome, as its absence could plausibly lead to a serious yet unrecognized lysosomal storage disease. PLBD1 and PLBD2 are semi-orphans in the sense of being probable phospholipases of B class but with uncertain physiological substrates and thus functionalities. PLBD1 and PLBD2 constitute a small gene family (sequence homology class) within vertebrates though one that occurs expanded in some early diverging eukaryotes. PLBD2 presents a special difficulty in that a sequence of post-translational steps are apparently necessary for its activation. Without these, potential substrates can hardly be assayed. These steps include removal of the signal peptide, mannosylation appropriate to the lysosome targeting receptor, and self-catalytic proteolytic activation to expose the substrate binding site as this becomes appropriate.

参考资料
  • Morgan CP. et al., 2004), Biochem J. 382 (2): 441-9.
  • Kim W. et al., 2011, Mol Cell. 44 (2): 325-40.
  • Havugimana PC. et al., 2012. Cell. 150 (5): 1068-81.
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    货号: HG13947-CF
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