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ISSN
1672-9145
Acta Biochim Biophys Sin
2004, 36(9): 644–648 CN 31-1940/Q
Preparation of Polyclonal Antibodies of Rubisco Large and Small
Subunits and Their Application in the Functional Analysis of the Genes
Peng-Da MA1, Tian-Cheng LU1, Xiao-Fu ZHOU1,2,
Xiao-Juan ZHU1, and Xing-Zhi WANG1*
1The Institute of Genetics and Cytology, Northeast
Normal University, Changchun 130024, China;
2The Institute of Gene Technology, Jilin Normal
University, Siping 136000, China
Abstract Spinach Rubisco (ribulose-1,5-bisphosphate
carboxylase/oxygenase) large (rbcL) and small (rbcS) subunits were separated by
SDS-PAGE, and protein amount and purity were determined by Bradford assay.
Polyclonal antibodies against rbcL and rbcS subunit were generated in female
BALB/c mice and had no cross-reaction with each other. A total of 81 mg antigens were
used and 0.3 ml anti-sera with titer of 1:5000 were yielded. The antibodies
were also applicable to study rbcL and rbcS in tobacco plant Nicotiana
benthamiana. Potato virus X vector pGR107 induced silencing of rbcS
gene by Agrobacterium in Nicotiana benthamiana was performed. The
expression level of rbcL and rbcS was lower in rbcS silenced plants than
that in control plants as detected by the corresponding antibodies. This implied that the
expression of rbcL was regulated by rbcS.
Key words
ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco); rbcL; rbcS;
polyclonal antibody; virus-induced gene silencing (VIGS)
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Received: May 27, 2004 Accepted: August 3, 2004
This work was supported by a grant from the National Natural Science
Foundation of China (No. 30370767)
Abbreviations: Rubisco, ribulose-1,5-bisphosphate
carboxylase/oxygenase; rbcL, large subunits of Rubisco; rbcS, small subunits of
Rubisco; SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis;
PVX, potato virus X; ELISA, enzyme-linked immunosorbent assay; VIGS,
virus-induced gene silencing
*Corresponding author: Tel,
86-431-5269769; E-mail, [email protected]