[Construction of eukaryotic fusion expression vectors of LSDP5 and cellular localization].
OBJECTIVE
To construct the eukaryotic fusion expression vectors and localization of (lipid storage droplet protein 5, LSDP5) in COS7 cells.


METHODS
Eukaryotic expression plasmid pCMV5-HA was digested with Nde I and BamH I and pMD18-LSDP5 plasmid was digested with Bgl II and Nde I. Full length LSDP5 was subcloned into pCMV5-HA. After confirmed by restriction enzyme digestion analysis and sequencing, the plasmid pCMV5-HA-LSDP5 was transfected into COS7 cells using Lipofectamine 2000 and the expression and localization of the fusion protein was detected by fluorescent microscope.


RESULTS
Restriction digestions and sequencing assays showed that the recombinant plasmid of pCMV5-HA-LSDP5 was successfully constructed and the fusion proteins were detected in cytoplasm and were targeted to the surfaces of lipid droplets visualized by Bodipy 493/503 in COS7 cells.


CONCLUSIONS
The recombined plasmid pCMV5-HA-LSDP5 expressing the fusion protein of LSDP5 and HA-tagged is successfully constructed and the fusion protein distributes in cytoplasm and is co-localized with lipid droplets.
Liu
Fang
F
Department of Pathology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Zhang
Li-jun
LJ
Ye
Jing
J
Zhang
Li-ying
LY
Li
Fan-fan
FF
Li
Hang
H
Gu
Yu
Y
Li
Qing
Q
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D050356:Lipid Metabolism; D009124:Muscle Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*
D047908:Intracellular Signaling Peptides and Proteins; D009124:Muscle Proteins; D011993:Recombinant Fusion Proteins; C551268:lipid storage droplet protein 5, mouse