Composite amplification kit for 44 human Y chromosome loci and application thereof

A Y-chromosome and compound amplification technology, applied in the field of compound amplification kits, can solve problems such as poor compatibility of the kits, limited number of STR loci, etc.

Active Publication Date: 2019-06-14
BEIJING PEOPLESPOT TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve technical problems such as the limitation of the number of STR loci detected simultaneously by multiplex amplification and the poor compatibilit

Method used

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  • Composite amplification kit for 44 human Y chromosome loci and application thereof
  • Composite amplification kit for 44 human Y chromosome loci and application thereof
  • Composite amplification kit for 44 human Y chromosome loci and application thereof

Examples

Experimental program
Comparison scheme
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Example Embodiment

[0074] Example 1 Development of a kit for compound amplification of 44 chromosomal loci

[0075] 1. Design of primers for compound amplification of 44 chromosomal loci

[0076] For the compound amplification system, especially the compound amplification system of 44 loci of the present invention, the requirements for the specificity of the primer sequence, the secondary structure, the stability of binding to the target sequence, and the amplification efficiency are extremely high. Continuous amplification-optimization-amplification cycle experiments have obtained 44 pairs of primers that specifically and efficiently amplify 44 loci. The specific sequences are shown in Table 1:

[0077] Table 1 Amplification primers for 44 loci and 2 IPCs

[0078]

[0079]

[0080] The above primers were fluorescently labeled with different colors according to the following grouping methods of the corresponding loci: the first group, IPC60, rs199815934, DYS456, DYS549, DYS439, DYS19, DYS392, DYS643, DY...

Example Embodiment

[0087] Example 2 Establishment of a compound amplification system and program for 44 chromosomal loci

[0088] The annealing temperature and cycle number were optimized for the amplification primers of 44 loci, and the optimal reaction program was obtained: 95℃ for 2 minutes; 94℃ for 5 seconds, 60℃ for 45 seconds, 72℃ for 45 seconds Clock, this step runs 30 cycles; 60℃ keep for 20 minutes; 4-10℃ keep.

Example Embodiment

[0089] Example 3 Application of compound amplification kit to genotyping 9948 cell line

[0090] The kit provided by the present invention is used to perform compound amplification of 44 Y chromosome loci on the 9948 cell line. The chelex 100 method was used to extract template DNA from the 9948 cell line. The amplification reaction was performed on the ABI 9700 thermal cycler, the electrophoresis and detection were performed on the ABI 3130 genetic analyzer, and the data analysis was performed using GeneMapper IDX v1.2 software. The reagent materials such as allelic ladders used can be obtained commercially and are conventional materials commonly used by those skilled in the art.

[0091] 1. Extract DNA using chelex 100 method (refer to "Forensic DNA Protocol", HumanPress, 1998 for specific methods):

[0092] (1) Take 3μl of the 9948 cell strain cultured in a 500μl centrifuge tube.

[0093] (2) Shake and mix the chelex solution to fully suspend chelex, add 195μl 5% chelex 100 (100-...

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Abstract

The invention relates to a composite amplification kit for 44 human Y chromosome loci and application thereof. The invention provides a composite amplification system of the 44 human Y chromosome loci. The system includes specific primers for amplifying the 44 loci including 41 Y chromosome STR loci and 3 Y chromosome Indel loci. The 44 pairs of specific primers are subjected to group fluorescencelabeling by the six-color fluorescent labeling technology. Through design and optimization of the primer sequence and the working concentration, simultaneous efficient specific sensitive amplification of the 44 human Y chromosome loci is achieved. A detection result of the composite amplification system has high individual recognition capability and good data compatibility, and can be used in practice for paternity testing and individual recognition, thereby effectively reducing the detection cost of human DNA typing and improving the detection working efficiency.

Description

technical field [0001] The invention relates to the technical fields of molecular genetics and biological detection, in particular to a composite amplification kit for 44 human Y chromosome loci and applications thereof. Background technique [0002] Short tandem repeat locus (Short Tandem Repeat, STR) is a genetic marker commonly used at present, which widely exists in prokaryotic and eukaryotic genomes, and is a kind of several repeating units composed of 2 to 6 nucleotides. Ten to more than one hundred nucleotide repeats. Different numbers of core sequences are arranged in tandem repeats, while the lengths are polymorphic. According to the conserved sequences at both ends, design primers for PCR, and then detect polymorphisms of STR genetic markers by polyacrylamide, agarose gel electrophoresis or capillary electrophoresis. Compared with other genetic markers, STR markers have small fragments and are easy to amplify, and are more suitable for trace and degraded samples....

Claims

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Application Information

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IPC IPC(8): C12Q1/6888C12Q1/6858C12N15/11
CPCY02A50/30
Inventor 不公告发明人
Owner BEIJING PEOPLESPOT TECH
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