Synthesis of a functional anti-phytochrome single-chain Fv protein in transgenic tobacco.
We have expressed a synthetic gene that encodes an antigen-binding single-chain FV protein (scFV) in transgenic tobacco plants. The scFV gene was created by polymerase chain reaction (PCR) amplification of the variable domain coding regions from a mouse monoclonal hybridoma cell line. The monoclonal cell line secretes an IgG1 antibody that binds to the plant regulatory photoreceptor protein, phytochrome. The cloned scFV gene was expressed initially in Escherichia coli and shown to produce a 28 kD, phytochrome-binding binding scFV protein. Transgenic tobacco plants expressing the scFV gene were also found to produce a functional scFV protein, and seeds from transgenic R1 progeny displayed aberrant phytochrome-dependent germination. The scFV from transgenic tobacco could be isolated, to near homogeneity, by a single phytochrome-Sepharose affinity chromatography step.
Owen
M
M
Department of Botany, University of Leicester, U.K.
Gandecha
A
A
Cockburn
B
B
Whitelam
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005813:Genes, Synthetic; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D010834:Phytochrome / Q000276:immunology*; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D014026:Nicotiana / Q000235:genetics*
D007135:Immunoglobulin Variable Region; D010834:Phytochrome