title: [Site-directed mutagenesis and protein expression of SCN5A gene associated with congenital long QT syndrome].
abstract: OBJECTIVE
  To construct the sodium channel gene SCN5A-delQKP1507-1509 mutation associated with congenital long QT syndrome, and its eukaryotic expression vector, and to examine the expression of mutation protein in human embryonic kidney (HEK) 293 cells.
  
  
  METHODS
  Eukaryotic expression vector PEGFP-delQKP-hH1 for SCN5A-delQKP1507-1509 mutation was constructed by rapid site-directed mutagenesis. HEK293 cells were transfected with the wild or mutant vector using lipofectamine, and then subjected to confocal microscopy. The transfected cells were immunostained to visualize intracellular expression of the mutant molecules.
  
  
  RESULTS
  Direct sequence and electrophoresis analysis revealed 9 basic group absences at position 1507-1509. The delQKP1507-1509 mutation eukaryotic expression vector was expressed in HEK293 cells. Immunostaining of transfected cells showed the expression of both wild type and mutant molecules on the plasma membrane and there was no difference in the amount of protein, which suggested that the mutant delQKP1507-1509 did not impair normal protein expression in HEK293 cells.
  
  
  CONCLUSIONS
  Successful construction of mutant SCN5AdelQKP1507-1509 eukaryotic expression vector and expression of SCN5A protein in HEK293 cells provides a basis for further study on the functional effects of congenital long QT syndrome as a cause of SCN5A mutation.
authors.affiliation: Department of Pediatrics, First Affiliated Hospital of Medical College of Xi'an Jiaotong University, Xi'an, China. srm1975@sohu.com
authors.forename: Rui-Ming
authors.initials: RM
authors.lastname: Shi
authors.forename: Hua
authors.initials: H
authors.lastname: Qiang
authors.forename: Yan-Min
authors.initials: YM
authors.lastname: Zhang
authors.forename: Ai-Qun
authors.initials: AQ
authors.lastname: Ma
authors.forename: Jie
authors.initials: J
authors.lastname: Gao
mesh_terms: D015153:Blotting, Western; D057809:HEK293 Cells; D006801:Humans; D008133:Long QT Syndrome / Q000151:congenital / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D062554:NAV1.5 Voltage-Gated Sodium Channel / Q000032:analysis / Q000235:genetics* / Q000502:physiology
chemical_list: D062554:NAV1.5 Voltage-Gated Sodium Channel; C568320:SCN5A protein, human