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 co

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

Example Embodiment

[0036] Specific Example 1 Determination of Locus

[0037] On the basis of the company's last product, four autosomal loci commonly used in Europe, SE33, D2S441, D10S1248, and D22S1045, as well as the Y chromosome locus DYS391 of the auxiliary sex recognition locus Amel, and an indel 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, Penta D, WA, D2S441, 179, D2S441 , FGA, Amel, D16S539, D22S1045, SE33 and D10S1248 have 27 loci.

Example Embodiment

[0038] Specific embodiment 2 Fluorescent labeling conforms to the design of the loci combination scheme of the amplification system

[0039] In the present invention, fluorescent dyes are identified and selected, and six fluorescent markers of blue, green, yellow, red, purple, and orange are selected to construct a 6-color fluorescent combination scheme. On the basis of determining the 6-color fluorescence combination scheme, through a large number of repeated experiments, design the locus combination mode and the type of fluorescent label. Considering the production cost and the amplification efficiency of the primers of each locus, the 27 locus is divided into 5 groups, using FAM, HEX, TAM, ROX, and Alex 594 group labels, and the molecular weight internal standard uses the sixth color orange fluorescent dye Atto 633 is marked. After screening, a preferred fluorescent dye label was finally determined. The first group of FAM labels: D3S1358, TH01, D21S11, D18S51 and Penta E, the...

Example Embodiment

Specific embodiment three

[0040] The present invention has an autosomal STR locus fluorescence-labeled composite 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 allele typing standards

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

[0046] 6) Spectral calibration standards

[0047] The above PCR Master Mix includes: DMSO 10mM, Tris-buffer 125mM, potassium chloride 125mM, ammonium sulfate 65mM, deoxynucleotide triphosphates (dNTPs) 7.5mM, BSA2.5mg / ml, etc., which can be compatible with common amplification in the market Various inspection materials.

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

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

[0050] The aforementioned Alleli...

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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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