pFastBac-N

Protocol tips

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	recombinant bacmids were purified using HighPure MiniPrep Kit (Life Technologies) and used to transfect Spodoptera frugiperda (Sf9) cells grown in Sf-900 II Serum-free Medium (SFM) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin as manufacturer’s instruction (Life Technologies). Transfection was carried out using Cellfectin II reagent as previously described [8] and according to the manufacturer’s instructions (Invitrogen-Life Technologies).

Protocol tips

recombinant bacmids were purified using HighPure MiniPrep Kit (Life Technologies) and used to transfect Spodoptera frugiperda (Sf9) cells grown in Sf-900 II Serum-free Medium (SFM) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin as manufacturer’s instruction (Life Technologies). Transfection was carried out using Cellfectin II reagent as previously described [8] and according to the manufacturer’s instructions (Invitrogen-Life Technologies).

Publication protocol

To rescue recombinant baculoviruses encoding the IBV N gene, recombinant bacmids were purified using HighPure MiniPrep Kit (Life Technologies) and used to transfect Spodoptera frugiperda (Sf9) cells grown in Sf-900 II Serum-free Medium (SFM) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin as manufacturer’s instruction (Life Technologies). Transfection was carried out using Cellfectin II reagent as previously described [8] and according to the manufacturer’s instructions (Invitrogen-Life Technologies). Recombinant IBV N protein was expressed using passage 2 or higher passage recombinant baculovirus stocks (> 107 pfu/ml). The protein was expressed with a carboxy-terminal 6xHis tag, and purification using Ni-NTA Superflow resin (QIAGEN Inc., Valencia, CA) was performed as described previously [7]. Concentration of the purified protein was measured by the method of bicinchoninic acid (BCA) assay (Thermo Scientific, Rockford, IL) at an absorbance of 562 nm, using bovine serum albumin (Sigma-Aldrich, St. Louis, MO) as the protein standard. Aliquots of the protein were stored at − 80 C until used.

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