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STR (short tandem repeat) sequence high-throughput detection method with base selective controllable extension and detection reagent thereof

A detection method and selective technology, which is applied in the field of high-throughput DNA sequencing in biotechnology, can solve the problems of small throughput and inability to meet simultaneous detection, and achieve the goal of reducing complexity, reducing the number of base sites, reducing costs and consumption the effect of time

Inactive Publication Date: 2012-10-03
SOUTHEAST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It can be seen that the current detection method is based on capillary electrophoresis similar to the first-generation sequencing, and the detection throughput is small, which cannot meet the requirements of simultaneous detection of massive samples.

Method used

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  • STR (short tandem repeat) sequence high-throughput detection method with base selective controllable extension and detection reagent thereof
  • STR (short tandem repeat) sequence high-throughput detection method with base selective controllable extension and detection reagent thereof
  • STR (short tandem repeat) sequence high-throughput detection method with base selective controllable extension and detection reagent thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1: Using a high-throughput detection method for STR sequences with base-selective and controlled extension to perform multi-sample high-throughput gender detection on the STR sequence of the Amelogenin locus:

[0039] Using a high-throughput detection method for STR sequences with base-selective and controllable extension, the detection steps are:

[0040] (1) Selection and amplification of the locus: Add a pair of PCR amplification primers for the Amelogenin locus together with the DNA template of the sample to be tested into the amplification system, control appropriate reaction conditions, and perform PCR reaction to obtain the Amelogenin gene The STR sequence amplification product of the locus. Wherein, the 5' end of one of the pair of amplification primers is modified with a connectable group. Different samples need to be amplified in independent amplification systems.

[0041](2) Preparation of the STR sequence library on the detection chip: by spottin...

Embodiment 2

[0050] Example 2: D3S1358, TH01, D18S51, Penta E, D5S818, D13S317, D7S820, D16S539, CSF1PO, Penta D, TPOX, SE33, D8S1179, D21S11, vWA using a high-throughput detection method of base-selective and controlled extension of STR sequences , STR sequences of 16 loci of FGA are used for identification and detection of sample separation, and the library preparation adopts the chip direct preparation method:

[0051] Using a high-throughput detection method for STR sequences with base-selective and controllable extension, the detection steps are:

[0052] (1) Selection and amplification of loci: add a pair of PCR amplification primers corresponding to the above 16 loci to the amplification system together with the DNA template of the sample to be tested, control appropriate reaction conditions, and perform PCR reaction. The STR sequence amplification product of the locus was obtained. Different samples need to be amplified in independent amplification systems.

[0053] (2) Prepara...

Embodiment 3

[0064] Example 3: F13A1, F13B, FES / FPS, HPRTB, LPL, D22S1045, DYS19, DYS385 a / b, DYS388, DYS389 I / II, DYS390, DYS391 by a high-throughput detection method of base-selective and controlled extension of STR sequences 17 loci, DYS392, DYS393, DYS434, DYS437, and Y-GATA-H4, were tested for mixed samples at the same time, and the library was prepared using the magnetic bead preparation method:

[0065] Using a high-throughput detection method for STR sequences with base-selective and controllable extension, the detection steps are:

[0066] (1) Selection and amplification of loci: Due to the mixed sample system, nucleic acid identification fragments need to be introduced to distinguish different samples and different loci. Firstly, the corresponding nucleic acid identification fragments are introduced into a pair of PCR amplification primers corresponding to the above-mentioned 17 loci, and then the amplification primers introduced with the nucleic acid identification fragments ...

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Abstract

The invention relates to an STR (short tandem repeat) sequence high-throughput detection method with base selective controllable extension and a detection reagent thereof. The basic principle of an STR sequence detection method and a high-throughput DNA (deoxyribonucleic acid) sequencing technology are combined, and the method which runs on a high-throughput sequencing platform for detecting massive STR sequence samples and the corresponding detection reagent are provided. The method comprises the following steps of: directly or indirectly fixing STR sequences of the samples to be detected on a detection chip according to a library preparation method corresponding to a sequencing system, adding the appropriate detection reagent according to the detection flow process, controlling reaction conditions, adopting the base selective controllable extension technical scheme to detect fluorescence intensity signals emitted by all reaction sites where the samples are located, and finally getting the detection results of the massive samples by analyzing fluorescence signal photos of all the detection sites during the whole detection process. The greatest advantage of the method disclosed by the invention is that the number of the samples which can be detected every time is greatly increased, and detection cost and time consumption can be further greatly reduced.

Description

technical field [0001] The invention relates to a method for high-throughput detection of STR sequences with base-selective and controllable extension and its detection reagents, which belong to the field of high-throughput DNA sequencing in biotechnology, and in particular relate to the use of second-generation high-throughput sequencing technology to detect STR sequences method. Background technique [0002] The second generation of high-throughput DNA sequencing: the development and completion of the Human Genome Project and various model organism genome projects have had a huge impact on contemporary biological and medical research. People can understand the differences of life phenomena, the law of disease occurrence and development, and the interaction between drugs and living organisms at the genetic level. As far as gene sequence analysis is concerned, the focus of the post-genome era has shifted from the determination of the whole genome sequence of a single specie...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 李俊吉陆祖宏刘全俊
Owner SOUTHEAST UNIV
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