With the development of genetically modified organism (GMO) detection techniques, the Polymerase Chain Reaction (PCR) technique has been the mainstay for GMO detection, and real-time PCR is the most effective and important method for GMO quantification. An event-specific detection strategy based on the unique and specific integration junction sequences between the host plant genome DNA and the integrated gene is being developed for its high specificity. This study establishes the event-specific detection methods for TC1507 and CBH351 maizes. In addition, the event-specific TaqMan real-time PCR detection methods for another seven GM maize events (Bt11, Bt176, GA21, MON810, MON863, NK603, and T25) were systematically optimized and developed. In these PCR assays, the fluorescent quencher, TAMRA, was dyed on the T-base of the probe at the internal position to improve the intensity of the fluorescent signal. To overcome the difficulties in obtaining the certified reference materials of these GM maizes, one novel standard reference molecule containing all nine specific integration junction sequences of these GM maizes and the maize endogenous reference gene, zSSIIb, was constructed and used for quantitative analysis. The limits of detection of these methods were 20 copies for these different GM maizes, the limits of quantitation were about 20 copies, and the dynamic ranges for quantification were from 0.05 to 100% in 100 ng of DNA template. Furthermore, nine groups of the mixed maize samples of these nine GM maize events were quantitatively analyzed to evaluate the accuracy and precision. The accuracy expressed as bias varied from 0.67 to 28.00% for the nine tested groups of GM maize samples, and the precision expressed as relative standard deviations was from 0.83 to 26.20%. All of these indicated that the established event-specific real-time PCR detection systems and the reference molecule in this study are suitable for the identification and quantification of these GM maizes. 相似文献
The effects of concentrated carrot protein (CCP) containing 15.4% (w/w) carrot (Daucus carota) antifreeze protein on texture properties of frozen dough and volatile compounds of crumb were studied. CCP supplementation lowered the freezable water content of the dough, resulting in some beneficial effects including holding loaf volume steadily and making the dough softer and steadier during frozen storage. Furthermore, SPME-GC-MS analysis showed CCP supplementation did not give any negative influences on volatile compounds of crumb and gave a pleasant aroma felt like Michelia alba DC from trans-caryophyllene simultaneously. Combining our previous results that CCP supplementation improves the fermentation capacity of the frozen dough, CCP could be used as a beneficial additive for frozen dough processing. 相似文献
ABSTRACT The mechanism of the effects of flumorph (a novel fungicide) was investigated by analyzing alterations of hyphal morphology, cell wall deposition patterns, F-actin organization, and other organelles in Phytophthora melonis. Calcofluor white staining suggested that flumorph did not inhibit the synthesis of cell wall materials, but disturbed the polar deposition of newly synthesized cell wall materials during cystospore germination and hyphal growth. After exposure to flumorph, zoospores were able to switch into cystospores accompanied with the formation of a cell wall, whereas cystospores failed to induce the isotropic-polar switch and did not produce germ tubes but continued the isotropic growth phase. In flumorph-treated hyphae, the most characteristic change was the development of periodic swelling ("beaded" morphology) and the disruption of tip growth. Newly synthesized cell wall materials were deposited uniformly throughout the diffuse expanded region of hyphae, in contrast to their normal polarized patterns of deposition. These alterations were the result of F-actin disruption, identified with the fluorescein isothiocynate (FITC)-phalloidin staining. The disruption of F-actin also was accompanied by disorganized organelles: each swelling of subapical hyphae was associated with a nucleus. Vesicles did not undergo polarized secretion to the apical hyphae, but diffused around nuclei for the subapical growth; thus, the cell wall was thickened with periodic expansion along the hyphae. Upon removing flumorph, normal tip growth and organized F-actin were observed again. These data, as well as data published earlier, suggest that flumorph may be involved in the impairment of cell polar growth through directly or indirectly disrupting the organization of F-actin. The primary site of action by flumorph in the disruption of the F-actin organization is under investigation. 相似文献
Low winter temperatures severely stress newly arriving insect species. Adaptive evolutionary changes in cold tolerance can facilitate their establishment in new environments. Ambrosia artemisiifolia, a noxious invasive plant, occurs throughout China. Ophraella communa, a biological control agent of A. artemisiifolia, mainly occurs in southern China. However, in 2012, it established populations in Beijing (39.98°N, 115.97°E) following introduction from Laibin (23.62°N, 109.37°E), implying cold adaptation. The mechanisms underlying its rapid evolution of cold tolerance remain unknown. We investigated the levels of cryoprotectants and energy reserves in adult O. communa from two latitudes. In high-latitude insects, we found high trehalose, proline, glycerol, total sugar, and lipid levels; five potential genes (Tret1a, Tret1b, Tret1-2, P5CS, and GST), responsible for regulating cold tolerance and involved in trehalose transport, proline biosynthesis, and glutathione S-transferase activation, were highly expressed. These hybridisation changes could facilitate cold temperature adaptation. We demonstrate the genetic basis underlying rapid adaptation of cold tolerance in O. communa, explaining its extension to higher latitudes. Thus, specialist herbivores can follow host plants by adapting to new temperature environments via rapid genetic evolution.