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Fluorescence labelled multiplex amplification primer set containing human genome DNA 22 loci, kit and application thereof

A fluorescent labeling and compound amplification technology, applied in the field of nucleic acid amplification, can solve the problems of destroying the PCR amplification process, failure to obtain typing results, and multi-inhibitor components, so as to improve amplification performance, avoid freeze-thaw process, The effect of improving specificity

Inactive Publication Date: 2017-04-26
AGCU SCIENTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Some samples have special sources such as blood stains in the soil, etc. Even after the DNA extraction step, there are still a large number of PCR inhibitors such as heme, humic acid, etc., which destroy the PCR amplification process, resulting in the inability to obtain a complete analysis. type result
Crude DNA extraction methods such as Chelex will also leave more inhibitor components

Method used

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  • Fluorescence labelled multiplex amplification primer set containing human genome DNA 22 loci, kit and application thereof
  • Fluorescence labelled multiplex amplification primer set containing human genome DNA 22 loci, kit and application thereof
  • Fluorescence labelled multiplex amplification primer set containing human genome DNA 22 loci, kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Test the sensitivity of the kit, 50pg Control DNA 9948 (ubit3.0 quantitative standard after dilution to 50pg) / 10μL system, 29 cycles of amplification.

[0064] A kit for simultaneously analyzing fluorescently labeled multiplexed amplification of 22 loci of human genomic DNA, comprising the following components: wherein the primers corresponding to the 22 loci and their primer concentrations are:

[0065]

[0066]

[0067] The above primer pairs are labeled in groups, and the 5' end of one primer in each primer pair is labeled with a fluorescent dye. The combinations used in this example are: D3S1358, D13S317, D7S820, D16S539, D1S1656, Penta E loci are marked with blue dyes; DYS391, TPOX, TH01, D2S1338, CSF1PO, Penta D loci are marked with green dyes; D19S433, The vWA, D21S11, D18S51, D6S1043 loci are marked with black dyes; the Amelogenin, D8S1179, D5S818, D12S391, FGA loci are marked with red dyes.

[0068] The kit also includes a molecular weight internal stand...

Embodiment 2

[0083] A fluorescence-labeled compound amplification inspection system with 22 loci was used to identify trace samples at the scene of the case. The same batch uses ABI’s STR kit Identifiler TM plus for comparative testing. A public security bureau provided 566 samples from different individuals: 250 samples of blood, cigarette butts, semen, costal cartilage samples, and 316 swabs. Genomic DNA was extracted with reference to "GA / T 383-2014 Forensic Science DNA Laboratory Inspection Specification".

[0084] A kit for simultaneously analyzing fluorescently labeled multiplexed amplification of 22 loci of human genomic DNA, comprising the following components: wherein the primers corresponding to the 22 loci and their primer concentrations are:

[0085]

[0086]

[0087]

[0088] The above primer pairs are labeled in groups, and the 5' end of one primer in each primer pair is labeled with a fluorescent dye. The combinations used in this example are: D3S1358, D13S317, ...

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Abstract

The invention provides a fluorescence labelled multiplex amplification primer set containing human genome DNA 22 loci, a kit and application thereof. The kit cooperates with a new hot start Taq enzyme and a corresponding premixed system of a PCR buffer solution to enhance the resistant ability of a PCR system to an inhibitor. At the same time, the PCR sensitivity is enhanced, all the 22 genome loci can be detected under a DNA template amount of 50pg, and the kit can be stored at 4DEG C for one year. The kit is suitable for forensic STR typing detection, parentage identification, individual recognition and other multiplex PCR application fields.

Description

technical field [0001] The invention belongs to the technology in the field of nucleic acid amplification, and specifically relates to a primer set, kit and application thereof for fluorescently labeled compound amplification containing 22 loci of human genome DNA. The system can be applied to forensic STR typing detection, kinship Identification and individual identification. Background technique [0002] Polymerase Chain Reaction (PCR) is a technique for rapidly amplifying specific DNA fragments in vitro. It is the cornerstone of modern molecular biology and one of the most important means. It is widely used in medical testing, biology Basic research, animal and plant inspection and other fields. Multiplex PCR is a kind of PCR technology classification, which refers to the composite amplification technology composed of more than 2 pairs of primers. The multiplex PCR technology used in practical applications usually refers to a multiplex amplification system with more tha...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/156
Inventor 郑卫国陈林丽张雷石美林于安路王艳芳郭育林
Owner AGCU SCIENTECH
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