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A DNA molecular detection method using 3D barcode

A DNA molecule and detection method technology, applied in the field of 3D barcode DNA molecule detection, can solve the problems of expensive, insensitive, time-consuming and expensive

Active Publication Date: 2021-03-23
THE FIRST AFFILIATED HOSPITAL OF ARMY MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods are insensitive, complex and time-consuming or expensive and require specialized laboratory training

Method used

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  • A DNA molecular detection method using 3D barcode
  • A DNA molecular detection method using 3D barcode
  • A DNA molecular detection method using 3D barcode

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] DNA molecular detection method using 3D barcode

[0033] (1) Preparation of DNA single-stranded bicyclic C1C2

[0034] Search the ctDNA gene sequence on NCBI, take 4 of the sequences as the DNA to be tested, and design 4 kinds of DNA single-strand circle C1 and DNA single-strand L2 according to each DNA sequence to be tested. The nucleotide sequences are as follows, italics are added The thick sequence is the complementary sequence of single-stranded loop C1 and single-stranded L2, and the underlined sequence is the complementary sequence of single-stranded L2 and the DNA to be tested.

[0035] Test DNA-1: 5'-GGAGCTGGTGACGTAGGCAAG-3' (SEQ ID NO.1)

[0036] DNA-2 to be tested: 5'-GTTGGAGCACGTGGTGTTGGG-3' (SEQ ID NO.2)

[0037] Test DNA-3: 5'-AATGATGCACGTCATGGTGGC-3'(SEQ ID NO.3)

[0038] Test DNA-4: 5'-AAAATAGGTAATTTGGGTCTA-3'(SEQ ID NO.4)

[0039] C1-1: 5'-ACTGTAACCATTCTTGTTTCCCAACCCGCCCTACCCAATATCATTTATCTGAATACCGTG-3' (SEQ ID NO. 5)

[0040] L2-1: 5'-PO4 3 -CACGG...

Embodiment 2

[0063] Test DNA Sensitivity Test

[0064] The test DNA1-4 and the control DNA were respectively in the molar mass ratio of 0:100, 0.1:99.9, 0.2:99.8, 0.3:99.7, 0.4:99.6, 0.5:99.5, 0.8:99.2, 1:99, 10:90 , mixed at a ratio of 100:0 to ensure that the total copy number is 1011copies / mL to form 4 kinds of target DNA solutions containing different concentrations of DNA to be tested, wherein the proportion of each DNA to be tested is 0%, 0.1%, 0.2 %, 0.3%, 0.4%, 0.5%, 0.8%, 1%, 10%, 100%. Detect target DNA solution according to the method of embodiment 1, the result is as follows Figure 4 As shown, the DNA-1 to be tested has a significant difference in the fluorescence intensity signal between 0.3% and 0.2%, and the fluorescence intensity of the ratio of 0.2% is almost the same as that of 0.1% and 0%, basically zero The fluorescence intensity of the DNA-1 ratio of 0.3% is significantly enhanced, and with the increase of the ratio, the fluorescence intensity continues to increase,...

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Abstract

The invention particularly relates to a DNA molecule detection method using a 3D bar code, belongs to the technical field of gene diagnosis, the detection method comprises steps of designing a DNA single-stranded ring C1 and a DNA single-stranded L2 according to a to-be-detected DNA sequence; completely hybridizing sequences at two ends of the DNA single-stranded L2 with a section of sequence of the DNA single chain ring C1; connecting sequence gaps at two ends of the DNA single-stranded L2 to form the DNA single-stranded double-ring C1C2; hybridizing and connecting with a probe on the 3D barcode; hybridizing DNA to be detected with DNA single-stranded double-loop C1C2 to obtain part of double-stranded DNA with restriction enzyme cutting sites, cutting the double-stranded DNA by restriction enzyme to release a topological structure, carrying out solid-phase RCA reaction on the surface of the 3D bar code, and finally carrying out fluorescence decoding. The DNA molecular detection method provided by the invention can realize detection of DNA single base mutation, has high detection sensitivity and strong specificity, and can detect 0.1% of mutation to the minimum.

Description

technical field [0001] The invention belongs to the field of gene diagnosis, and in particular relates to a 3D barcode DNA molecular detection method. Background technique [0002] Genetic testing is a technology that detects trace amounts of DNA through blood, other body fluids, or cells. It takes the peripheral venous blood or other tissue cells of the subject to amplify their genetic information, and then uses specific equipment to detect the DNA in the cells of the subject. Molecular information is used to detect and analyze the gene types and gene defects contained in it and whether its expression function is normal. It can diagnose diseases and predict the risk of diseases. Current DNA detection methods mainly include ARMS-PCR (Amplification Refractory Mutation System-PCR), ddPCR (Droplet Digital PCR) or sequencing. These methods are insensitive, complex and time-consuming or expensive and require specialized laboratory training. Therefore, it is necessary to establi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/682C12Q1/683
CPCC12Q1/682C12Q1/683C12Q2525/307C12Q2521/301C12Q2531/125C12Q2563/107
Inventor 阳莎陈鸣唱凯詹新宇汤晓琦肖瑶赵爽
Owner THE FIRST AFFILIATED HOSPITAL OF ARMY MEDICAL UNIV
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