Functional characterization of aroA from Rhizobium leguminosarum with significant glyphosate tolerance in transgenic Arabidopsis.
Glyphosate is the active component of the top-selling herbicide, the phytotoxicity of which is due to its inhibition of the shikimic acid pathway. 5-Enolpyruvylshikimate-3-phosphate synthase (EPSPS) is a key enzyme in the shikimic acid pathway. Glyphosate tolerance in plants can be achieved by the expression of a glyphosate-insensitive aroA gene (EPSPS). In this study, we used a PCR-based two-step DNA synthesis method to synthesize a new aroA gene (aroAR. leguminosarum) from Rhizobium leguminosarum. In vitro glyphosate sensitivity assays showed that aroAR. leguminosarum is glyphosate tolerant. The new gene was then expressed in E. coli and key kinetic values of the purified enzyme were determined. Furthermore, we transformed the aroA gene into Arabidopsis thaliana by the floral dip method. Transgenic Arabidopsis with the aroAR. leguminosarum gene was obtained to prove its potential use in developing glyphosate-resistant crops.
Department of Shanghai Key Laboratory of Agricultural Genetics and Breeding, Biotechnology Research Institute of Shanghai Academy of Agricultural Sciences, Shanghai 201-106, P. R. China.
Jing
J
Han
Yong-Sheng
YS
Tian
Jing
J
Xu
Li-Juan
LJ
Wang
Bo
B
Wang
Ri-He
RH
Peng
Quan-Hong
QH
Yao
D051229:3-Phosphoshikimate 1-Carboxyvinyltransferase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000502:physiology; D004351:Drug Resistance; D005998:Glycine / Q000031:analogs & derivatives* / Q000633:toxicity; D008969:Molecular Sequence Data; D010802:Phylogeny; D030821:Plants, Genetically Modified / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000502:physiology; D016961:Rhizobium leguminosarum / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment; D000097797:Glyphosate
D051229:3-Phosphoshikimate 1-Carboxyvinyltransferase; D005998:Glycine; D000097797:Glyphosate