Molecular and Toxicity Analyses of White Granulated Sugar and Other Processing Products Derived From Transgenic Sugarcane

Abstract

This study aimed to prepare the sugar industry for the possible introduction of genetically modified (GM) sugarcane and derived retail sugar products and to address several potential public concerns regarding the characteristics and safety of these products. GM sugarcane lines with integrated Cry1Ab and EPSPS foreign genes were used for GM sugar production. Traditional PCR, real-time fluorescent quantitative PCR (RT-qPCR), and enzyme-linked immunosorbent assay (ELISA) were performed in analyzing leaves, stems, and other derived materials during sugar production, such as fibers, clarified juices, filter mud, syrups, molasses, and final GM sugar product. The toxicity of GM sugar was examined with a feeding bioassay using Helicoverpa armigera larvae. PCR and RT-qPCR results showed that the leaves, stems, fibers, juices, syrups, filter mud, molasses, and white granulated sugar from GM sugarcane can be distinguished from those derived from non-GM sugarcane. The RT-qPCR detection method using short amplified product primers was more accurate than the traditional PCR method. Molecular analysis results indicated that trace amounts of DNA residues remain in GM sugar, and thus it can be accurately characterized using molecular analysis methods. ELISA results showed that only the leaves, stems, fibers, and juices sampled from the GM sugarcane differed from those derived from the non-GM sugarcane, indicating that filter mud, syrup, molasses, and white sugar did not contain detectable Cry1Ab and EPSPS proteins. Toxicity analysis showed that the GM sugar was not toxic to the H. armigera larvae. The final results showed that the GM sugar had no active proteins despite containing trace amounts of DNA residues. This finding will help to pave the way for the commercialization of GM sugarcane and production of GM sugar.

Keywords: PCR analysis; enzyme-linked immunosorbent assay; genetic modification sugar, genetic modification sugarcane; real-time fluorescent quantitative PCR; toxicity feeding bioassay.

Figure 1

Gel imaging analysis of all DNA extractions (without the digestion of RNA). Lane 1: DNA extracted from leaves; Lane 2: DNA extracted from stems; Lane 3: DNA extracted from fibers; Lane 4: DNA extracted from juices; Lane 5: DNA extracted from filter mud; Lane 6: DNA extracted from the syrup; Lane 7: DNA extracted from molasses; Lane 8: DNA extracted from the GM sugar; Lane 9: DNA extracted from the non-GM sugar.

Figure 2

Traditional PCR analysis of all samples. Lane M: DNA Marker; Lane 1: Leaf (GM); Lane 2: Stem (GM); Lane 3: Fiber (GM); Lane 4: Juice (GM); Lane 5: Filter mud (GM); Lane 6: Syrup (GM); Lane 7: Molasses (GM); Lane 8: Sugar (GM); Lane 9: Sugar (Non-GM).

Figure 3

RT-qPCR analysis of GM and non-GM sugar. (A): amplification plots of endogenous gene actin (CT value: GM leaf, 22.12; GM sugar, 31.42; non-GM sugar, 31.17); (B): dissociation curve of endogenous gene actin, the three samples share the same dissociation curve; (C): amplification plots of the endogenous gene Cry1Ab (CT value: GM leaf, 21.81; GM sugar, 31.44; non-GM sugar, no CT value); (D): dissociation curve of the endogenous gene Cry1Ab, GM leaf sample and GM sugar share the same dissociation curve, but the non-GM sugar has no dissociation curve.

Figure 4

ELISA of all samples.

Figure 5

Larvae feeding bioassay. (A): larvae feed with larva fodder mix with GM sugar; (B): larvae feed with larva fodder mix with non-GM sugar; (C): larvae feed with larva fodder mix with GM sugarcane stem material. (D): 1: Daily average weight of 10 larvae fed with larva fodder mix with GM sugar; 2: Daily average weight of 10 larvae fed with larva fodder mix with non-GM sugar; 3: Daily average weight of 10 larvae fed with larva fodder mix with GM sugarcane stem material.