Functional expression of the Aequorea victoria green fluorescent protein in insect cells using the baculovirus expression system.
A DNA fragment encoding the green fluorescent protein (GFP) was isolated via PCR from a jellyfish Aequorea victoria cDNA, cloned and sequenced. Subsequently, a recombinant baculovirus bearing the coding region of the GFP under the transcriptional control of the Autographa californica nuclear polyhedrosis virus (AcMNPV) polyhedrin gene promoter was constructed and isolated. High-level expression of GFP could be easily monitored in Spodoptera frugiperda (Sf9) insect cells after infection with recombinant baculovirus, due to the intrinsic fluorescence (lambda(max) = 508 nm) of the recombinant protein after excitation with blue light (lambda(max) = 400 nm). The functional recombinant GFP displayed an apparent molecular mass of approximately 43 kDa and the fluorescence emission spectrum of the recombinant protein was virtually identical to that of the native green fluorescent protein.
Reiländer
H
H
Max-Planck-Institut für Biophysik, Abt. Mol. Membranbiologie, Frankfurt/M, Germany.
Haase
W
W
Maul
G
G
D000818:Animals; D016367:Baculoviridae; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D015183:Restriction Mapping; D037401:Scyphozoa / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D018411:Spodoptera; D014162:Transfection
D017931:DNA Primers; D008164:Luminescent Proteins; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins