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Eugenol, a plant-derived small compound, shows great medicinal potential. However, whether and how eugenol regulates crop physiology remains elusive. Here we reported that eugenol induced Cd (cadmium) tolerance in the root of Brassica rapa. Roots were treated with eugenol and CdCl 2 simultaneously (eugenol + Cd) or pretreated with eugenol followed by CdCl 2 treatment (eugenol → Cd). Eugenol significantly attenuated Cd-induced growth inhibition, ROS accumulation, oxidative injury, and cell death, which were confirmed by in vivo histochemical analysis. Eugenol remarkably decreased free Cd 2+ accumulation in root. Eugenol intensified GSH (glutathione) accumulation in roots upon CdCl 2 exposure, which explained the decrease in free Cd 2+ and attenuation of oxidative injury. Eugenol stimulated endogenous H 2 S (hydrogen sulfide) generation by upregulating the expression of BrLCD ( l-cysteine desulfhydrase) and BrDCD ( d-cysteine desulfhydrase) as well as their enzymatic activities in CdCl 2 -treated root. Application of H 2 S biosynthesis inhibitor or H 2 S scavenger led to the decrease in endogenous H 2 S level in Cd-treated root, which further compromised all the above effects of eugenol. These findings suggested that eugenol triggered H 2 S → GSH signaling cassette in plants to combat Cd stress, which shed new light on eugenol-modulated plant physiology and the interaction between eugenol and H 2 S.

The purification and partial enzymology characteristics of polyphenol oxidase (PPO) from rape flower were studied. After preliminary treatments, the crude enzyme solution was in turn purified with ammonium sulfate, dialysis, and Sephadex G-75 gel chromatography. The optimal conditions and stability of PPO were examined at different pH values and temperatures. Subsequently, PPO was also characterized by substrate (catechol) concentrations, inhibitors, kinetic parameters, and molecular weight. Results showed that the optimal pH for PPO activity was 5.5 in the presence of catechol and that PPO was relatively stable at pH 3.5-5.5. PPO was moderately stable at temperatures from 60 to 70 °C, whereas it was easily denatured at 80-90 °C. Ethylenediaminetetraacetic acid, sodium chloride, and calcium chloride had little inhibitive effects on PPO, whereas citric acid, sodium sulfite, and ascorbic acid had strongly inhibitive effects. The Michaelis-Menten constant (K(m)) and maximal reaction velocity (V(max)) of PPO were 0.767 mol/L and 0.519 Ab/min/mL of the crude PPO solution, respectively. PPO was finally purified to homogeneity with a purification factor of 4.41-fold and a recovery of 12.41%. Its molecular weight was 60.4 kDa, indicating that the PPO is a dimer. The data obtained in this research may help to prevent the enzymatic browning of rape flower during its storage and processing.

We have developed a new immunoassay method to detect genetically modified (GM) maize and rape containing phosphinothricin-N-acetyltransferase (PAT). PAT encoded by Bialaphos resistance gene (bar) was highly expressed in soluble form in Escherichia coli BL21(DE3) and purified to homogeneity by Ni2+ affinity chromatography. A simple and efficient extraction and purification procedure of PAT from GM maize and rape was developed by means of the immunoaffinity column (IAC) as a cleanup tool. Purified polyclonal antibodies against PAT was produced and coupled covalently to CNBr-activated Sepharose 4B. Both the binding conditions and elution protocols were optimized. The IAC was successfully employed to isolate and purify the PAT from the various tissues of GM maize (Bt11 and Bt176) and rapes (MS1/RF1 and MS8/RF3). Enzyme linked immunosorbent assay (ELISA) procedures were established further on to measure the PAT protein. GM maize cannot be differentiated from non-GM maize by ELISA. But IAC-ELISA allowed 0.5% GMOs to be detected in MS1/RF1 and MS8/RF3 and 10% GMOs to be detected in Bt11 and Bt176, which makes this method an acceptable method to access PAT protein in GM rapes and maize.

Polymerase chain reaction (PCR) methods are very useful techniques for the detection and quantification of genetically modified organisms (GMOs) in food samples. These methods rely on the amplification of transgenic sequences and quantification of the transgenic DNA by comparison to an amplified reference gene. Reported here is the development of specific primers for the rapeseed (Brassica napus) BnACCg8 gene and PCR cycling conditions suitable for the use of this sequence as an endogenous reference gene in both qualitative and quantitative PCR assays. Both methods were assayed with 20 different rapeseed varieties, and identical amplification products were obtained with all of them. No amplification products were observed when DNA samples from other Brassica species, Arabidopsis thaliana, maize, and soybean were used as templates, which demonstrates that this system is specific for rapeseed. In real-time quantitative PCR analysis, the detection limit was as low as 1.25 pg of DNA, which indicates that this method is suitable for use in processed food samples which contain very low copies of target DNA.