Cloning, prokaryotic expression of rice L-galactono-1,4-lactone dehydrogenase gene and preparation of anti-GLDH antibodies.

The full length of L-galactono-1,4-lactone dehydrogenase (GLDH) gene was amplified by RT-PCR from rice leaves and sequenced. The amplified gene was about 1 752 bp. The coding region for part of mature protein (789 bp) was subcloned into pET-30a(+). The recombinant plasmid pET30a-GLDHe was identified by enzyme digestion and DNA sequencing. Data of SDS-PAGE indicated that a 36 000 (relative molecular mass) protein was expressed. The antiserum against GLDH was generated by immunizing rabbits with the prokaryotic expressed protein. The polyclonal antibody was analyzed by Western blot for its specificity and titer. The antibody could recognize GLDH protein specifically and its titer was about 1∶1 000.