title: [Cloning and expression of human pcsk9 cDNA and preparation of its polyclonal antibody].
abstract: OBJECTIVE
  To construct the prokaryotic expression vector of human proprotein convertase subtilisin/kexin type 9 (pcsk9), induce it to express and prepare the polyclonal antibody of human pcsk9.
  
  
  METHODS
  The sequence of pcsk9 cDNA was synthesized and amplied by PCR. By means of digestion and subcloning, the sequence was cloned into the expression vector PET-28b. The recombinant plasmid of pET-28b-pcsk9 was transformed into the competent cells of E.coli BL21 (DE3), induced to express, purified and identified. The New Zealand rabbits were immunized with the purified protein. The resulted polyclonal antibody was analyzed by ELISA and Western blotting.
  
  
  RESULTS
  The prokaryotic expression vector PET-28b-pcsk9 was constructed and expressed well in E.coli BL21 (DE3). We obtained the polyclonal antibody by immunizing the rabbits with the target protein. The purified antibody was proved to have a good specificity by Western blotting and its titer reached 1:13 200.
  
  
  CONCLUSIONS
  The expression vector of pcsk9 was successfully constructed and expressed. The polycolonal antibody of pcsk9 protein was obtained as well.
authors.affiliation: Department of Clinical Laboratory, Guangzhou Medical University, Guangzhou, China.
authors.forename: Fengmei
authors.initials: F
authors.lastname: Li
authors.forename: Hui
authors.initials: H
authors.lastname: Yu
authors.forename: Jinlong
authors.initials: J
authors.lastname: Song
authors.forename: Zhongmin
authors.initials: Z
authors.lastname: Liu
mesh_terms: D000818:Animals; D000906:Antibodies / Q000097:blood; D000918:Antibody Specificity; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D012697:Serine Endopeptidases / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification
chemical_list: D000906:Antibodies; D018076:DNA, Complementary; D011994:Recombinant Proteins; C472125:PCSK9 protein, human; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases; D012697:Serine Endopeptidases