有哪些信誉好的足球投注网站a genetically encoded tool kit for manipulating and monitoring membrane phosphatidylinositol 4,5-bisphosphate in intact cells基因编码的工具用于操作和监控膜磷脂酰肌醇4,5-bisphosphate在完整细胞.pdfVIP
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a genetically encoded tool kit for manipulating and monitoring membrane phosphatidylinositol 4,5-bisphosphate in intact cells基因编码的工具用于操作和监控膜磷脂酰肌醇4,5-bisphosphate在完整细胞
A Genetically Encoded Tool Kit for Manipulating and
Monitoring Membrane Phosphatidylinositol 4,5-
Bisphosphate in Intact Cells
Fabian Hertel, Agathe Switalski, Elisa Mintert-Jancke, Katharina Karavassilidou, Kirsten Bender, Lutz
´
Pott*, Marie-Cecile Kienitz
Institute of Physiology, Ruhr-University Bochum, Bochum, Germany
Abstract
Background: Most ion channels are regulated by phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P ) in the cell
2
membrane by diverse mechanisms. Important molecular tools to study ion channel regulation by PtdIns(4,5)P2 in living
¨
cells have been developed in the past. These include fluorescent PH-domains as sensors for Forster resonance energy
transfer (FRET), to monitor changes in plasma membrane For controlled and reversible depletion of PtdIns(4,5)P , voltage-
. 2
sensing phosphoinositide phosphatases (VSD) have been demonstrated as a superior tool, since they are independent of
cellular signaling pathways. Combining these methods in intact cells requires multiple transfections. We used self-cleaving
viral 2A-peptide sequences for adenovirus driven expression of the PH-domain of phospholipase-Cd1 (PLCd1) fused to
ECFP and EYFP respectively and Ciona intestinalis VSP (Ci-VSP), from a single open reading frame (ORF) in adult rat cardiac
myocytes.
Methods and Results: Expression and correct targeting of ECFP-PH-PLC
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