Showing papers by "Dabing Zhang published in 2003"

A novel real‐time quantitative PCR method using attached universal template probe

Abstract: A novel real-time quantitative polymerase chain reaction (PCR) method using an attached universal template (UT) probe is described. The UT is an approximately 20 base attachment to the 5¢ end of a PCR primer, and it can hybridize with a complementary TaqMan probe. One of the advantages of this method is that different target DNA sequences can be detected employing the same UT probe, which substantially reduces the cost of real-time PCR setup. In addition, this method could be used for simultaneous detection using a 6-carboxy-fluoresceinlabeled UT probe for the target gene and a 5-hexachloro-fluorescein-labeled UT probe for the reference gene in a multiplex reaction. Moreover, the requirement of target DNA length for UT‐PCR analysis is relatively flexible, and it could be as short as 56 bp in this report, suggesting the possibility of detecting target DNA from partially degraded samples. The UT‐PCR system with degenerate primers could also be designed to screen homologous genes. Taken together, our results suggest that the UT‐PCR technique is efficient, reliable, inexpensive and less labor-intensive for quantitative PCR analysis.

Production of FaeG, the Major Subunit of K88 Fimbriae, in Transgenic Tobacco Plants and Its Immunogenicity in Mice

Abstract: Transgenic tobacco plants stably expressing recombinant FaeG, which is the major subunit and adhesin of K88ad fimbriae, were obtained. Analysis of sera from immunized mice indicates that in mice, the immunogenicity induced by plant-derived FaeG protein is comparable to that generated with traditional approaches.

Multiplex polymerase chain reaction method for detection of bovine materials in foodstuffs.

Abstract: Rapid identification of bovine materials in animal foodstuffs is essential for effective control of a potential source of bovine spongiform encephalophathy. A convenient polymerase chain reaction (PCR)-based assay was developed for detection and identification of a bovine-specific genomic DNA sequence in foodstuffs. Simultaneously the assay assessed the DNA quality of the experiment system by amplification of a highly conserved eucaryotic DNA region of the 18-S ribosomal gene, helping to check the reliability of the test result. The amplified bovine-specific PCR product was a genomic DNA fragment of lactoferrin, a low copy gene that was different from a commonly used bovine-specific mitochondria sequence for identification of bovine materials. The specificity of this method was confirmed by the absence of detectable homologous PCR product using reference foodstuff samples that lacked bovine-derived meat and bonemeals, or genomic DNA samples from vertebrates whose offals are commonly included in animal feeds. This method could detect the presence of bovine material in foodstuffs when the samples contained >0.02% bovine-derived meat and bone meal. Furthermore, it was not affected by prolonged heat treatment. The specificity, convenience, and sensitivity of this method suggest that it can be used for the routine detection of bovine-derived materials.

Molecular cloning and characterization of Stevia rebaudiana UDP-glucosyltransferase.

Abstract: We report here the cloning and characterization of a UDP-glucose flavonoid glucosyltransferase (srUFGT) in Stevia rebaudiana. The isolated cDNA was 1419 bp in length encoding 473 deduced amino acids with a predicted molecular mass of 53.2 kDa. The products of in vitro translation from an expression vector had anthocyanidins and steviol glucosyltransferase activity. Comparison of the activity of the recombinant UDP-glucosyltransferase toward a range of acceptor substrates suggests that it may participate in the synthesis of steviol glycosides. The results support the hypothesis that the flavonoid glucosyltransferases, which have a broad substrate specificity, may be not only involved in flavonoid glucosylation but also play a role in producing the water-soluble steviol-glycosides in S. rebaudiana.