PubMed Health⌕ Search

Biomedical subjects

Zhang Bao

Publications and source records attributed to Zhang Bao.

5 recordsLinked to original sources
PubMed2006

Oligonucleotide microarray with RD-PCR labeling technique for detection and typing of human papillomavirus.

Currently, screening for high-risk human papillomavirus (HPV) infection remains an important health concern throughout the world, because of the close association between certain types of HPV and cervical cancer. In this study, we explore the possibility of using approximately 70mer oligonucleotide microarray for detection and genotyping of HPV. The approximately 70mer type-specific oligonucleotide probes of four different types HPV were designed by using biological software Array designer 2.0, which analyzed the whole genome sequences of HPV and selected optimal probes. These probes were synthesized and printed onto the surface of glass slides in order to prepare a low-density microarray. HPV samples were labeled with fluorescence dyes Cy3 using a method of restriction display polymerase chain reaction (RD-PCR). HPV plasmid DNA was restricted with Sau3A I to produce multiple fragments that were ligated to adaptors subsequently and used as PCR template. PCR labeling was performed with the fluorescently labeled universal primer (Cy3-UP) whose sequence is designed according to the adaptor of the RD-PCR approaches. The labeled samples were hybridized with the oligonucleotide microarray. The scanning results showed that HPV DNA hybridized specifically with multiple spots correspondingly to show positive signals, whereas no signals were detected of all the negative and blank controls. These results demonstrated that approximately 70mer oligonucleotide microarray can be applied to HPV detection and genotyping. The application of RD-PCR in the sample labeling can increase significantly the sensitivity of the assay and will be especially useful for the discriminate diagnosis of multiple pathogens.

Carbocyanines↗
PubMed2004

Microarrays for the detection of HBV and HDV.

The increasing pace of development in molecular biology during the last decade has had a direct effect on mass testing and diagnostic applications, including blood screening. We report the model Microarray that has been developed for Hepatitis B virus (HBV) and Hepatitis D virus (HDV) detection. The specific primer pairs of PCR were designed using the Primer Premier 5.00 program according to the conserved regions of HBV and HDV. PCR fragments were purified and cloned into pMD18-T vectors. The recombinant plasmids were extracted from positive clones and the target gene fragments were sequenced. The DNA microarray was prepared by robotically spotting PCR products onto the surface of glass slides. Sequences were aligned, and the results obtained showed that the products of PCR amplification were the required specific gene fragments of HBV, and HDV. Samples were labeled by Restriction Display PCR (RD-PCR). Gene chip hybridizing signals showed that the specificity and sensitivity required for HBV and HDV detection were satisfied. Using PCR amplified products to construct gene chips for the simultaneous clinical diagnosis of HBV and HDV resulted in a quick, simple, and effective method. We conclude that the DNA microarray assay system might be useful as a diagnostic technique in the clinical laboratory. Further applications of RD-PCR for the sample labeling could speed up microarray multi-virus detection.

Base Sequence↗
PubMed2002

Construction of a cDNA fragment library from SH-SY5Y cells using restriction display PCR.

A complementary DNA (cDNA) fragment library from SH-SY5Y cells is constructed using a restriction display polymerase chain reaction (RD-PCR) technique. Messenger RNA (mRNA) is extracted from SH-SY5Y cells and single-strand cDNA synthesised using an anchored oligo primer (dT18). The second strand is produced by nick translation. The double strands are cleaved with the restriction enzyme Sau3AI and the fragments ligated with universal linker. The products are amplified with universal primers and selected primers, ligated into the pMDI8-T vector, and then sequenced. The library constructed contained 136 subgroups, each comprising seven to 12 cDNA fragments. RD-PCR proved a simple, effective way to construct a cDNA library, and this will contribute to the investigation of gene expression in the neuron in future microarray studies.

DNA, Complementary↗