Selected article for: "cationic lipid and par2529 vector"
Title: Oligomerization of a membrane protein correlates with its retention in the Golgi complex
  • Document date: 1993_9_2
    • ID: 5z1xminb_10
    Snippet: COS-'/and HeLa cells were maintained in DME with 10 and 5% FCS, respectively. COS-7 cells plated in 35-mm dishes (40-70% confluent) were transfected with an SV40-based expression vector using DEAE-dextran (Machamer et al., 1985) . Expression was analyzed at 40--48 h posttransfection. For expression using the vaccinia-T7 system, HeLa cells (40-70% confluent) were infected with the recombinant vaccinia virus vTF7-3 encoding T7 RNA polymerase (Fuers.....
    Document: COS-'/and HeLa cells were maintained in DME with 10 and 5% FCS, respectively. COS-7 cells plated in 35-mm dishes (40-70% confluent) were transfected with an SV40-based expression vector using DEAE-dextran (Machamer et al., 1985) . Expression was analyzed at 40--48 h posttransfection. For expression using the vaccinia-T7 system, HeLa cells (40-70% confluent) were infected with the recombinant vaccinia virus vTF7-3 encoding T7 RNA polymerase (Fuerst et al., 1986 ) at a multiplicity of infection of 10-20. After adsorption for 30 min at 37°C, the inoculum was replaced with 0.75 ml of serum-free medium containing 2/~g of a vector (pAR2529) encoding the appropriate gene behind the T7 promoter and 5-10/~1 of the cationic lipid ~TransfectACE" (GIBCO BRL, Gaithersburg, MD; Rose et al., 1991) . Expression level was varied by changing the amount of DNA added per well (0.02-5 ~tg/dish). In coexpression experiments, cells were transfected with 1/~g of DNA encoding each construct. Expression was analyzed by metabolic labeling starting 3 h after infection.

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