44 composite amplification kits and applications of human Y chromosome gene seats

A compound amplification and Y chromosome technology, which is applied in the field of compound amplification kits, can solve the problems of poor kit compatibility and limited number of STR loci, and achieve strong test material adaptability, good analysis data compatibility, and high compatibility sexual effect

Active Publication Date: 2022-08-09
BEIJING PEOPLESPOT TECH
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  • Abstract
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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 compatibility of the kits in the prior art, the present invention provides a multiplex amplification kit capable of simultaneously detecting 44 chromosomal loci and its application

Method used

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  • 44 composite amplification kits and applications of human Y chromosome gene seats
  • 44 composite amplification kits and applications of human Y chromosome gene seats
  • 44 composite amplification kits and applications of human Y chromosome gene seats

Examples

Experimental program
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Effect test

Embodiment 144

[0074] Example 1 Development of a 44-chromosomal locus composite amplification kit

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

[0076] For the composite amplification system, especially the composite amplification system of 44 loci in the present invention, the requirements for the specificity of the primer sequence, secondary structure, stability of binding with the target sequence, amplification efficiency, etc. are extremely high. Continuous amplification-optimization-amplification cycle experiments yielded 44 pairs of primers that specifically and efficiently amplify 44 loci, the specific sequences of which 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 corresponding loci: the first group, IPC60, rs199815934, DYS456, DYS549, DYS439, DYS19, DYS392, DYS643, DY...

Embodiment 24

[0087] Example 2 Establishment of a complex 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°C for 2 minutes; 94°C for 5 seconds, 60°C for 45 seconds, and 72°C for 45 seconds 1 hour, this step was run for 30 cycles; 60°C for 20 minutes; 4-10°C for incubation.

Embodiment 3

[0089] Example 3 Genotyping of 9948 cell line using compound amplification kit

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

[0091] 1. Use chelex 100 method to extract DNA (refer to "Forensic DNA Protocol", HumanPress, 1998 for the specific method):

[0092] (1) Take 3 μl of cultured 9948 cell line into a 500 μl centrifuge tube.

[0093] (2) Shake and mix the chelex solution to make the chelex fully suspended, add 195 μl of 5% chelex 1...

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Abstract

The invention involves 44 composite amplification kits and applications of human Y chromosome gene seats.The invention provides a composite amplification system for 44 gene seats in human Y chromosomes, including specific primers used to enlarge 44 gene seats.Chromosome indel.Use the six -color fluorescence markup technology to group the 44 -specific primers, and through the design and optimization of the primer sequence and working concentration, the 44 human Y chromosome gene seats are achieved at the same time as efficient, special, and sensitive.The detection results of the composite amplification system have high individual recognition ability and have good data compatibility, which can be used in practice for parental identification and individual identification, effectively reducing the detection cost of human DNA typing, increasing the improvementThe work efficiency of detection.

Description

technical field [0001] The invention relates to the technical field of molecular genetics and biological detection, in particular to a composite amplification kit of 44 human Y chromosome loci and its application. Background technique [0002] Short tandem repeat loci (Short Tandem Repeat, STR) is a commonly used genetic marker, which widely exists in prokaryotic and eukaryotic genomes. Ten to more than one hundred nucleotide repeats. Different numbers of core sequences are arranged in tandem repeats, while lengths are polymorphic. Based on the conserved sequences at both ends, primers are designed for PCR, and then polymorphisms of STR genetic markers can be detected by polyacrylamide, agarose gel electrophoresis or capillary electrophoresis. Compared with other genetic markers, STR loci have small fragments and are easy to amplify, and are more suitable for trace and degraded samples. Compared with previous technologies such as RFLP, STR composite amplification technolo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6888C12Q1/6858C12N15/11
CPCY02A50/30
Inventor 不公告发明人
Owner BEIJING PEOPLESPOT TECH
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