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Fluorescence-labeled composite amplification kit for Y chromosome STR (short tandem repeat) gene loci capable of improving distinguishing capability and application thereof

A Y chromosome and fluorescent labeling technology, which is applied in the field of amplification kits and Y chromosome STR locus fluorescent labeling composite amplification kits, can solve the problems of unreported and used issues, and achieve the effect of improving the cumulative individual recognition ability

Active Publication Date: 2012-10-03
AGCU SCIENTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Therefore, in order to further improve the sensitivity and efficiency of the commercially available amplification kits, the applicant conducted in-depth research on the genetic polymorphism of the Y-STR locus in the male population, and based on this, developed a The Y-chromosomal STR loci fluorescence-labeled composite amplification kit with improved identification ability has not been reported and used at present, and there is no Y-STR fluorescent label detection kit in my country at present, so it is always expected by those skilled in the art solved problem

Method used

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  • Fluorescence-labeled composite amplification kit for Y chromosome STR (short tandem repeat) gene loci capable of improving distinguishing capability and application thereof
  • Fluorescence-labeled composite amplification kit for Y chromosome STR (short tandem repeat) gene loci capable of improving distinguishing capability and application thereof
  • Fluorescence-labeled composite amplification kit for Y chromosome STR (short tandem repeat) gene loci capable of improving distinguishing capability and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1 Determination of loci

[0044] 通过对DYS19、DYS381、DYS385a / b、DYS388、DYS389I / II、DYS390、DYS391、DYS392、DYS393、DYS425、DYS426、DYS437、DYS438、DYS439、DYS444、DYS447、DYS448、DYS449、DYS456、DYS458、DYS450、DYS468、 The polymorphism survey of 31 loci including DYS522, DYS527a / b, DYS630, DYS635, DYS710 and Y-GATA-H4 in the Chinese population, finally determined DYS19, DYS385a / b, DYS388, DYS389I / II, DYS390, DYS391 , DYS392, DYS393, DYS437, DYS438, DYS439, DYS444, DYS447, DYS448, DYS449, DYS456, DYS458, DYS522, DYS527a / b, DYS635 and Y-GATA-H4 and other 24 loci. It is compatible with existing kits, ensures the sharing and exchange of existing DNA data, and can improve the cumulative individual identification ability and cumulative non-parent exclusion rate of the DNA testing system, which is more in line with the technical requirements of DNA testing.

Embodiment 2

[0045] Example 2 Design of locus combination scheme for fluorescent labeling multiplex amplification system

[0046] The present invention identifies and selects fluorescent dyes, selects five fluorescent markers of blue, green, yellow, red and orange, and constructs a five-color fluorescent combination scheme.

[0047] On the basis of determining the 5-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 24 loci were divided into 4 groups, labeled with FAM, HEX, TAMRA, and ROX, and the molecular weight internal standard was marked with the fifth color orange fluorescent dye SIZ . After screening, a preferred fluorescent dye labeling method was finally determined: DYS391, DYS389I / II, DYS439, DYS438 and DYS449, using FAM labeling; DYS456, DYS458, DYS437, DYS635, DYS448 and DYS527a / b, u...

Embodiment 3

[0048] Example 3 Experimental process for the detection of amplified loci and their products

[0049] 1. Configuration of reaction system

[0050]

[0051] 2. Amplification thermal cycle experiment scheme

[0052] (1) Place the PCR amplification tube on the thermal cycler;

[0053] (2) Select the program recommended below to amplify;

[0054] (3) The amplified samples should be kept away from light;

[0055] Amplification program for thermal cycler

[0056]

[0057] 3. Fluorescent detection of the amplified product on a genetic analyzer

[0058] The sample loading mixture is composed of deionized formamide and molecular weight internal standard (AGCU Marker SIZ-500) in the system [(0.5μL AGCU Marker SIZ-500)×(injection number)+(12μL deionized formamide)×(injection number)〕. Mix 12.5 μL of the loading mixture with 1 μL of the amplification product or the allelic analysis standard (Y Allelic Ladder) in the system, avoiding the generation of air bubbles. Denature at ...

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Abstract

The invention provides a fluorescence-labeled composite amplification kit for Y chromosome STR (short tandem repeat) gene loci capable of improving distinguishing capability. When the kit is used for detecting DNA (deoxyribonucleic acid) gene, not only can 17 STR gene loci of DYS391, DYS389I / II, DYS439, DYS438, DYS456, DYS458, DYS437, DYS635, DYS448, Y-GATA-H4, DYS19, DYS392, DYS393, DYS390 and DYS385a / b, which can be analyzed by the commercial kit, be amplified and analyzed, but also at least one of STR gene loci of DYS449, DYS527a / b, DYS522, DYS388, DYS447 and DYS444 can be simultaneously amplified and analyzed, so that the accumulative individual distinguishing capability and cumulative probability of exclusion of the system are improved, and the individual distinguishing capability is improved overall.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and generally relates to an amplification kit used for detecting polymorphisms of short tandem repeat loci (Short Tandem Repeat, STR for short) in individuals. Specifically, the present invention relates to a Y chromosome STR locus fluorescent label multiplex amplification kit with improved identification capability. Background technique [0002] There are 3×10 in human genomic DNA 9 bp, 10% of which are tandem repeats, called satellite DNA. According to the length of the repeating unit, it can be divided into large satellites, medium satellites, small satellites and microsatellites. The repeating unit is only composed of 2-7 base pairs called microsatellites, also known as short tandem repeat loci ( Short Tandem Repeat, referred to as STR), the DNA sequence formed by this tandem repeat can produce hundreds of millions of genotype combinations, and the frequency of each combination...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 郑卫国葛斌文
Owner AGCU SCIENTECH
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