Detection of hepatitis D virus by cDNA microarray method.

Background: Viral hepatitis is considered a major public health problem in most areas of the world. In acute and chronic infections, hepatitis D virus (HDV) infection often leads to a more severe disease. This study was designed to prepare microarrays for HDV detection.

[Relationship of serum levels of HGF and MMP-9 with disease activity of patients with systemic lupus erythematosus].

Objective: To determine the relationship of serum hepatocyte growth factor (HGF) and matrix metalloproteinase-9 (MMP-9) levels with the disease activity of systemic lupus erythematosus (SLE).

Methods: Serum levels of HGF and MMP-9 were measured by ELISA in 36 patients with SLE and 30 healthy subjects as controls.

Result: (1)Significantly increased serum level of HGF was found in SLE patients as compared with that in healthy controls (P<0.

[Co-expression of two spliceosomes of cyclin-dependent kinase-2 in SH-SY5Y cells].

Objective: To investigate the expression of cyclin-dependent kinase-2 (CDK-2) gene in SH-SY5Y cells.

Methods: The expression of CDK-2 gene was examined with reverse transcriptional (RT)-PCR, and the PCR products underwent electrophoresis on non-denaturing poly-acrylamide gel (PAG) followed by silver staining. The separated and purified DNAs were ligated into pMD18-T vector, and the positive clones identified by sequence analysis.

[DNA microarray for the detection of Yersinia pesits].

Objective: To develop a DNA microarray technique for fast diagnosis of plague.

Methods: Restriction display polymerase chain reaction (RD-PCR) and hybridization with fluorescently labeled cy5 were employed for detecting the sample DNA of Yersinia pestis, Yersinia pseudotuberculosis and Yersinia enterocolitica.

Results: The prepared microarray is capable of distinguishing Yersinia pestis from Yersinia pseudotuberculosis and Yersinia enterocolitica in the same genus.

[Preparation of the microarray for detecting hepatitis D virus].

Objective: To prepare the microarray for detection of hepatitis D virus (HDV).

Method: Several pairs of specific PCR primers were designed according to the conserved region of HDV genome. The DNA microarray were prepared by blotting the PCR products onto the surface of glass slides with the use of robotics, and restriction display PCR (RD-PCR) was employed to label the samples.

[Amplification and cloning of the N gene of SARS-associated coronavirus].

Objective: To amplify and clone the N gene of severe acute respiratory syndrome-associated coronavirus.

Method: Using primer Premier 5.0 software, two pairs of nested PCR primers were designed to amplify the N gene.

A tyrosine-modified hypocrellin B with affinity for and photodamaging ability towards calf thymus DNA.

An enhanced photodamaging ability towards CT-DNA was achieved in a tyrosine-modified hypocrellin B by improving the affinity of the sensitizer to DNA.

[Tumor infiltrating dendritic cells and Mucin1 gene expression in benign prostatic hyperplasia and prostate cancer].

Objective: To study the expression of Mucin1 gene and tumor infiltrating dendritic cells(TIDC) in the tissues of benign prostatic hyperplasia (BPH) and prostate cancer.

Methods: Mucin1 and TIDC were detected in 20 specimens of BPH and 30 specimens of prostate cancer by immunohistochemistry SP method.

Results: MUC1 expressed in both prostate cancer and BPH.

Endothelin-1 is a potent regulator in vivo in vascular calcification and in vitro in calcification of vascular smooth muscle cells.

We observed changes of endothelin content and endothelin mRNA in vivo in vascular calcification and in vitro in calcification of vascular smooth muscle cells to explore the role of endothelin in vascular calcification. Calcification model in vivo was induced by administration of Vitamin D(3) plus nicotine. Calcification of vascular smooth muscle cells (VSMCs) was induced by beta-glycerophosphate.

Possibility of using DNA chip technology for diagnosis of human papillomavirus.

To explore the application of DNA chip technology for the detection and typing of Human Papillomavirus (HPV), the HPV6, 11, 16 and 18 gene fragments were isolated and printed onto aminosilane-coated glass slides by a PixSys 5500 microarrayer as probes to prepare the HPV gene chips. HPV samples, after being labeled with fluorescent dye by restriction display PCR (RD-PCR) technology, were hybridized with the microarray, which was followed by scanning and analysis. The experimental condition for preparing the HPV gene chips was investigated, and the possibility of HPV genotyping using gene chips was discussed.

Inhibition of taurine transport by high concentration of glucose in cultured rat cardiomyocytes.

Cultured rat cardiomyocytes were treated with 10, 20, and 30 mmol/L glucose and 30 mmol/L glucose plus protein kinase C (PKC) inhibitor, Chelerythrine. In the 20 and 30 mmol/L glucose-treated cells, taurine contents reduced by 15% and 27% (P<.05), respectively, taurine transporter (TAUT) mRNA levels reduced by 47% and 64% (P<.

Rapid preparation of DNA microarray using PCR for hepatitis B and D virus detection.

Objective: To prepare DNA microarray for detecting both hepatitis B and D virus as (HBV and HDV).

Methods: With the assistance of Oligo6.4 software, specific PCR primers targeting the conserved region of HBV and HDV were designed.

[Gene sequence analysis of SARS-associated coronavirus by nested RT-PCR].

Objective: To explore an effective means for the detection of Severe Acute Respiratory Syndrome (SARS)- associated coronavirus.

Methods: The RNAs of the virus contained in the sputum samples from established SARS patients were extracted and reversely transcripted, followed by nested PCR using the reversely transcripted cDNA as the template. The PCR products were cloned then into the pMD18-T vectors, followed by sequence analysis.

Cloning and sequence analysis of Bacillus thuringiensis gene fragments isolated by restriction digest PCR.

Objective: To clone and analyze Bacillus thuringiensis gene fragments isolated by restriction digest PCR (RD-PCR).

Method: Specific primers were designed to amplify the genes of Bacillus thuringiensis israelensis (Bti), and the PCR products were classified and re-amplified by RD-PCR to obtain the fragments for subsequent purification and cloning into the pMD18-T vectors, followed by rapid identification. The recombinant plasmids were extracted from positive clones and the target gene fragments were sequenced.

Gene expression study of Saccharomyces cerevisiae with the Agilent 2100 bioanalyser.

This study explores the restriction display-polymerase chain reaction (RD-PCR) application of a new chip-based nucleic acid analysis system (Agilent 2100 bioanalyser) in a gene differential expression study. Total RNAs is extracted from Saccharomyces cerevisiae, double-stranded complementary DNA (cDNA) is synthesised by reverse transcription from the purified messenger RNA (mRNA), RD-PCR conducted to obtain the cDNA fragments and bioanalyser and agarose gel electrophoresis compared for the analysis of RD-PCR products. The bioanalyser proved to be faster and more sensitive in separating and detecting gene fragments, and was also able to compare different gene fragments quantitatively.

Changes in amount of ADM mRNA and RAMP2 mRNA in calcified vascular smooth muscle cells.

This work was aimed to explore the changes and significance of adrenomedullin (ADM) mRNA and receptor activity modifying protein 2 (RAMP2) mRNA in calcified vascular smooth muscle cells (VSMCs). Calcification of cultured rat VSMCs was produced by incubation with beta-glycerophosphate. Content of ADM released by VSMCs was measured by radioimmunoassay (RIA).

Design and application of 60mer oligonucleotide microarray in SARS coronavirus detection.

The 60mer oligonucleotide microarray was designed and applied to detecting of SARS (severe acute respiratory syndrome) coronavirus. Thirty 60mer specific oligos were designed to cover the whole genome of the first submitted coronavirus strain, according to the sequence of TOR2 (GENEBANK Accession: AY274119). These primers were synthesized and printed into a microarray with 12 ×12 spots.

Gene expression study of Saccharomyces cerevisiae with the Agilent 2100 bioanalyser.

This study explores the restriction display-polymerase chain reaction (RD-PCR) application of a new chip-based nucleic acid analysis system (Agilent 2100 bioanalyser) in a gene differential expression study. Total RNAs is extracted from Saccharomyces cerevisiae, double-stranded complementary DNA (cDNA) is synthesised by reverse transcription from the purified messenger RNA (mRNA), RD-PCR conducted to obtain the cDNA fragments and bioanalyser and agarose gel electrophoresis compared for the analysis of RD-PCR products. The bioanalyser proved to be faster and more sensitive in separating and detecting gene fragments, and was also able to compare different gene fragments quantitatively.

[Proliferation and denucleation of K562 cells induced by cytochalasin B].

Objective: To observe the effect of cytochalasin B on the denucleation of K562 cells.

Methods: K562 cells were grown in RPMI 1640 medium supplemented with 10% calf serum and 4 microL cytochalasin B (CB). Denucleation was induced in the cultured cells by CB, and the cells were examined by phase contrast microscopy and Giemsa staining respectively.

[Construction and identification of cDNA library of E.coli mRNA with poly(A) tracts].

Objective: To construct and identify the cDNA library of E.coli mRNA with poly(A) tracts.

Methods: The cDNA library of E.

[Effect of probe purification on the reutilization of gene chip].

Objective: To investigate the effect of probe purity on the reutilization of gene chips.

Methods: Purified and unpurified probes were respectively hybridized with the gene chip and after being scanned with Scanarry lite, a laser scanning device, the gene chip was treated with stripping solution to wash off the probes. After another round of scanning under the same condition to obtain the images, the gene chip was recycled for another hybridization.

[Cloning E. coli mRNAs with poly (A) tails by restriction digest polymerase chain reaction].

Objective: To investigate the polyadenylation at the 3' terminal of the mRNAs in E.coli.

Methods: mRNAs of E.

[Cloning of an expressed sequence tag with restriction display polymerase chain reaction].

Objective: To isolate gene fragments from SH-SY5Y cells by way of restriction display polymerase chain reaction (RD-PCR).

Methods: Total mRNA was extracted from SH-SY5Y cells followed by synthesis of the single-strand cDNA with Oligo (dT18) as the anchored primer, and the second strand was synthesized by nick translation. The double strands were cleft with restriction enzyme Sau3A I and the fragments ligated with a universal adapter to be amplified with the universal primers and selected primers.

[Purification of PCR products with cetyltrimethylammonium bromide].

Objective: To establish a method for purifying PCR products with cetyltrimethylammonium bromide (CTAB).

Method: Selective precipitation of the PCR product was performed using CTAB, which forms compound with DNA fragment in salt solution of appropriate concentration, but not with single strain oligonucleotide or dNTPs. The precipitation could be dissolved in 1.