Integrated detection DNA microarray chip for HBV, HCV and HIV-1 in blood

A microarray chip and blood technology, applied in the direction of DNA / RNA fragments, recombinant DNA technology, microbial measurement / inspection, etc., can solve the problems of high cost of reagents, difficulty in meeting the requirements of reagents, and low detection throughput

Active Publication Date: 2013-02-27
GUANGZHOU BDS BIOLOGICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the most advanced nucleic acid extraction and fluorescent PCR reagents at home and abroad are difficult to meet the requirements; if the Pooling method is not used to directly detect each sample, the low detection throughput and high reagent cost are unacceptable! how to solve this problem?

Method used

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  • Integrated detection DNA microarray chip for HBV, HCV and HIV-1 in blood
  • Integrated detection DNA microarray chip for HBV, HCV and HIV-1 in blood
  • Integrated detection DNA microarray chip for HBV, HCV and HIV-1 in blood

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1: the design of HBV, HCV, HIV-1 virus gene specific primer and probe

[0030] HBV designs a specific probe and a pair of specific primers in the 5'UTR region;

[0031] A total of 6 probes covering 4 genotypes and 6 subtypes were designed for the 5' NCR region of HCV. The six probes were mixed according to the equal concentration and spotted on the chip; HCV designed two pairs of PCR primers in the 5'NCR region, the outer primer sequence: sense: 5'-TGTGAGGAACTTCTGTCTTC-3', antisense: 5'-CATGATGCACGGTCTACGA-3'; Internal primer sequence: sense: 5'-GCCATGGCGTTAGTACGA-3', antisense: 5'-TCGCAAGCACCCTATCAGG-3'. The inner primer CY3 is fluorescently labeled.

[0032] HIV-1 is in the more conserved gag region, design a probe, the sequence is 5'-AGTAAGAATGTATAGCCCT-3'; design a pair of primers, the sequence is: upstream 5'-AACCACCTATCCCAGTAGGATAA-3', downstream 5'-GGTCCTTGTCTTATGTCCAGAAT-3' , downstream primer CY3 fluorescently labeled.

[0033] Put all the above ...

Embodiment 2

[0034] Embodiment 2: use the product of the present invention to detect 38 parts of blood samples

[0035] Get 38 clinical blood samples and negative, positive reference product, adopt product of the present invention to detect and screen.

[0036] Use a commercial extraction kit to simultaneously extract DNA and RNA viral nucleic acids for RT-PCR reactions.

[0037] Using a commercial RT-PCR reaction kit, RT-PCR reaction was carried out according to the instructions.

[0038] Take the reaction product for DNA microarray hybridization reaction:

[0039] Take a microarray chip, take 2ul of 38 samples and 2 reference PCR or RT-PCR products respectively, and add them to the corresponding hybridization area of ​​the chip, then add 5ul of hybridization solution (5×SSC) to each well, and put the chip into In the tank of the automatic hybridization washer, set the hybridization temperature to 50°C and the hybridization time to 40 minutes. Automatic washing and air drying after hyb...

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Abstract

The invention relates to a DNA microarray chip, in particular to an integrated high-flux detection DNA microarray chip for hepatitis B virus (HBV), hepatitis C virus (HCV) and human immunodeficiency virus 1 (HIV-1). A solid-phase carrier substrate is used, and an oligonucleotide probe is designed in view of genes of HBV, HCV and HIV-1 so as to prepare the DNA microarray chip; and when matched witha PCR primer and other components, the DNA microarray chip can simultaneously detect the three viruses in a plurality of samples with high sensitivity and high specificity. The DNA microarray chip ismainly used for screening the virus nucleic acid in blood and blood products.

Description

technical field [0001] The invention relates to a DNA microarray chip, in particular to an integrated high-throughput detection DNA microarray chip for hepatitis B virus (HBV), hepatitis C virus (HCV) and human immunodeficiency virus type 1 (HIV-1) in human blood. array chip. The present invention adopts solid-phase carrier substrates, and prepares oligonucleotide probes designed for HBV, HCV, and HIV-1 virus genes into DNA microarray chips, which can simultaneously detect multiple samples at one time with high sensitivity and high specificity The three types of viruses are mainly used in the screening of viral nucleic acids in blood and blood products. Background technique [0002] Ensuring the safety of blood and blood products and minimizing the risk of blood-transmitted viruses has always been a concern of countries all over the world. With the continuous improvement of detection technology, the risk of transfusion-related viruses has been greatly reduced. However, due...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 胡守旺张帆李明何蕴韶
Owner GUANGZHOU BDS BIOLOGICAL TECH CO LTD
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