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Autosome STR gene locus fluorescent labeling composite amplification kit having enhanced identification ability, and applications thereof

An autosomal and fluorescent labeling technology, which is applied in the determination/testing of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problem of waste of DNA database data resources, shorten the time of adenylylation, and improve the balance The effect of shortening the amplification time

Inactive Publication Date: 2015-09-30
NINGBO HEALTH GENE TECHNOLOGIES CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, as the application of DNA as an identification method is becoming more and more extensive, users have higher and higher requirements for the number of loci, information volume, amplification time, application range of test materials, etc. of the kit. As the scale of DNA database construction continues to expand, the role of data comparison is becoming increasingly important. Since data comparison is based on the same STR locus, it is also necessary for the STR kit to be compatible at the locus, otherwise it will cause DNA Some data resources in the database are wasted

Method used

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  • Autosome STR gene locus fluorescent labeling composite amplification kit having enhanced identification ability, and applications thereof
  • Autosome STR gene locus fluorescent labeling composite amplification kit having enhanced identification ability, and applications thereof
  • Autosome STR gene locus fluorescent labeling composite amplification kit having enhanced identification ability, and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

specific Embodiment 1

[0036] Specific Example 1 Determination of Genetic Loci

[0037] On the basis of the company's previous product, four autosomal loci SE33, D2S441, D10S1248, and D22S1045 commonly used in Europe and auxiliary sex recognition loci Amel's Y chromosome locus DYS391 and an insertion-deletion fragment Y on the Y chromosome were added. -indel, there are D3S1358, TH01, D21S11, D18S51, Penta E, Y-indel, DYS391, D12S391, D6S1043, D2S1338, D1S1656, D5S818, D13S317, D7S820, D19S433, CSF1PO, XvSPO, D, D27D941 , FGA, Amel, D16S539, D22S1045, SE33 and D10S1248, a total of 27 loci.

specific Embodiment 2

[0038] Specific Example 2 Fluorescent labeling conforms to the design of the gene locus combination scheme of the amplification system

[0039] The present invention identifies and selects fluorescent dyes, selects six fluorescent markers of blue, green, yellow, red, purple and orange, and constructs a six-color fluorescent combination scheme. On the basis of determining the 6-color fluorescent combination scheme, through a large number of repeated experiments, the combination of loci and the type of fluorescent markers were designed. Considering the production cost and primer amplification efficiency of each locus, the 27 loci were divided into 5 groups, labeled with FAM, HEX, TAM, ROX and Alex 594, and the molecular weight internal standard was the sixth color orange fluorescent dye Atto 633 for marking. After screening, a preferred fluorescent dye marker was finally determined. The first group of FAM markers: D3S1358, TH01, D21S11, D18S51 and Penta E, the second group of H...

specific Embodiment 3

[0040] The present invention has the autosomal STR gene locus fluorescent label compound amplification kit with enhanced identification ability, and the kit includes:

[0041] 1)PCR Master Mix

[0042] 2) Primer Mix

[0043] 3) Control DNA 9948A

[0044] 4) Allelic Ladder allelic typing standard

[0045] 5) Size-500 orange fluorescent molecular weight internal standard

[0046] 6) Spectral calibration standard

[0047] The above PCR Master Mix includes: DMSO 10mM, Tris-buffer 125mM, potassium chloride 125mM, ammonium sulfate 65mM, deoxynucleotide triphosphates (dNTPs) 7.5mM, BSA 2.5mg / ml, etc., which can achieve compatible amplification common in the market Various inspection materials.

[0048] The above-mentioned Primer Mix includes all primers for amplifying 27 loci (see Table 1 for concentration), Taq enzyme 2-4U / 6.25μl, magnesium chloride 7.5Mm, etc.

[0049] The above-mentioned positive control substance is purchased human genomic DNA.

[0050] The Allelic Ladder ...

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Abstract

The present invention discloses an autosome STR gene locus fluorescent labeling composite amplification kit having enhanced identification ability, wherein 27 STR gene loci can be simultaneously amplified with the kit and comprise 24 autosome STR gene loci such as D3S1358, TH01, D21S11, D18S51, Penta E, D12S391, D6S1043, D2S1338, D1S1656, D2S441, D5S818, D13S317, D7S820, D19S433, CSF1PO, Penta D, vWA, D8S1179, TPOX, FGA, D16S539, D22S1045, SE33 and D10S1248, 2 Y chromosome gene loci such as Y-indel and DYS391, and a sex-determining locus Amel. According to the present invention, with the application of the kit to perform the DNA gene detection, all the gene loci of the current mainstream products at home and abroad are contained, the currently existing DNA database in most countries are covered, the compatibility problem is not required to be worried about, the accumulation individual recognition rate and the combined paternity non-exclusion probability of the system are improved, and the individual discriminability is improved on the whole.

Description

technical field [0001] The invention relates to a PCR amplification kit, in particular to a fluorescence composite amplification kit for simultaneous detection of 24 autosomal loci, 2 Y chromosome loci, and 1 gender locus in a single tube. The preparation method of the fluorescent compound amplification kit and the application of the kit in the field of forensic identification belong to the field of autosome typing and identification. Background technique [0002] Short tandem repeat (short tandem repeat, STR) is a type of DNA sequence with length polymorphism formed by tandem repetition of 2-6 bases in the human genome as the core unit. The number of core units varies and the number of repetitions is different. The genetic polymorphisms that make up the STR. STRs are widely distributed and numerous, accounting for about 10% of the human genome, and contain a huge amount of information. Different sequences can generate hundreds of millions of genotype combinations, and the ...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
CPCC12N15/11C12Q1/68
Inventor 金海英林锦锋刘亚举王万旭张兹均
Owner NINGBO HEALTH GENE TECHNOLOGIES CO LTD
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