Test strip technology for specific DNA detection using sybr Green I fluorescent dye

A fluorescent dye and specific technology, which is applied in the field of biotechnology applications, can solve the problems of inability to perform multiple detections and complicated labeling steps, and achieve the effects of rapid results display, low detection costs, and strong specificity

Active Publication Date: 2016-03-23
艾吉泰康(嘉兴)生物科技有限公司
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, the labeling steps of this method are complicated and cannot be used for multiple detection

Method used

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  • Test strip technology for specific DNA detection using sybr Green I fluorescent dye
  • Test strip technology for specific DNA detection using sybr Green I fluorescent dye
  • Test strip technology for specific DNA detection using sybr Green I fluorescent dye

Examples

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Embodiment 1

[0031] This example aims to detect influenza viruses A, B, H1, H3 and their mixtures. The nucleotide sequences of specific probes and target DNA are shown in Table 1.

[0032] Table 1. Influenza virus A, B, H1, H3 probes and target DNA sequences

[0033]

[0034] The detection method of this embodiment comprises the following steps:

[0035] (1) Selection of test strip material

[0036] Select nylon membrane (ZetaProbe TM , 15cm×15cmsheet, Bio-Rad) as the material of the test strip, the nylon membrane was cut into strips of 3cm×0.5cm.

[0037] (2) Preparation of probe sensing layer

[0038] Spot the probes A, B, H1, H3 (500nM, 0.5μL) on the cut nylon membrane at fixed intervals, and then irradiate the nylon membrane with 254nm ultraviolet light (UnitecLF-206LS) for 2 minutes at 60°C Bake for 5 minutes and finally store in a dark dry place.

[0039] (3) The target sequence of the sample to be tested is obtained

[0040] Primers were designed using probe A as a templat...

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Abstract

The invention provides a technology for rapidly detecting specific DNA (Deoxyribonucleic Acid) by applying an SYBR Green I fluorescent dye to a test paper strip. The technology comprises the following steps of selecting a nylon membrane as a material of the test paper strip; pointing probes onto the nylon membrane, then, irradiating with ultraviolet rays, baking, drying in the dark and storing, wherein the probes are specific nucleotide sequences to be detected; carrying out PCR (Polymerase Chain Reaction) amplification in a manner of taking samples to be detected as templates and taking primers, which are designed by taking the probes as templates, as primers, then, carrying out denaturation for 10 minutes at the temperature of 94 DEG C, and immediately lowering by 4 DEG C, so as to obtain sequences to be detected; flushing the nylon membrane with a buffer solution, dispensing a hybridization solution to the nylon membrane, hybridizing in the dark at room temperature, cleaning the nylon membrane sequentially with a buffer solution and deionized water, imaging with a 488nm-laser imaging scanner, and judging a result in accordance with whether the nylon membrane emits light or not, wherein the hybridization solution contains SYBR Green I and the sequences to be detected. The technology is simple, inexpensive and rapid and is applicable to clinical rapid diagnosis.

Description

technical field [0001] The invention relates to the application field of biotechnology, in particular to a technique for quickly detecting specific DNA on a test strip by using SYBRGreenI fluorescent dye. Background technique [0002] Pathogen detection has always been of great value in many research fields, especially in the medical and food industries. Rapid and accurate detection and identification of pathogens is the key to effective infection control and early treatment. However, standard pathogen culture requires not only incubation on medium for at least 8 hours, but also additional biochemical or immune recognition, which is tedious and time-consuming. In contrast, nucleic acid-based assays such as quantitative polymerase chain reaction (qPCR) and microarrays are faster and more sensitive, however, many of these methods are still complex and expensive. [0003] In recent years, reported lateral flow assays have contributed greatly to the development of paper-based ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6837C12Q2563/107C12Q2565/501C12Q2565/625
Inventor 李孟皇边红武韩凝武金霞
Owner 艾吉泰康(嘉兴)生物科技有限公司
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