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A dna library for detecting multiple osteochondroma pathogenic genes and its application

A DNA library, osteochondroma technology, applied in the field of DNA library, can solve the problems of inability to meet multi-pathogenic gene diseases and multi-sample timeliness, complex operation, expensive and other problems

Active Publication Date: 2019-06-18
汪道文
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the traditional Sanger sequencing-based gene detection method has the disadvantages of low throughput, high price, and complicated operation. One reaction can only detect one amplified region, which cannot meet the timeliness requirements of multi-pathogenic gene diseases and simultaneous detection of multiple samples.

Method used

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  • A dna library for detecting multiple osteochondroma pathogenic genes and its application
  • A dna library for detecting multiple osteochondroma pathogenic genes and its application
  • A dna library for detecting multiple osteochondroma pathogenic genes and its application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] 1. Reagents used in the method:

[0072] Ion AmpliSeq TM Library Kit 2.0, Ion PGM TM Template OT2 200Kit v3, Ion Sequencing 200Kit v2, Ion Xpress Barcode Adapters 1-16Kit, Ion 318 TM ChipKit v2

[0073] 2. Specimen collection and storage

[0074] (1) Specimen collection: The specimen is the peripheral blood of the patient. Blood is 5ml of venous blood taken routinely, treated with EDTA anticoagulant.

[0075] (2) Storage: It can be detected immediately, stored at 4°C for one week, and stored at -80°C for more than one week.

[0076] 3. Detection steps and result analysis:

[0077] (1) Extraction of genomic DNA of specimens: DNA extraction of specimens was carried out according to the operation instructions of the blood DNA extraction kit of Tiangen Biochemical Technology (Beijing) Co., Ltd.

[0078] (2) Super multiplex PCR amplification and library building of the target detection area: the whole exons of 15 genes involved in the present invention are used ...

Embodiment 2

[0113] The primer pool of the present invention contains 8 multiple osteochondroma pathogenic genes newly discovered by the inventor. As shown in Table 1 above. The newly discovered 8 multiple osteochondroma pathogenic genes come from the research accumulation and family investigation of the inventors over the years. The inventors first searched the known multiple osteochondroma pathogenic genes through the KEGG signaling pathway and the homologous gene library to find the homologous genes of the known pathogenic genes and the key genes on the same pathogenic pathway. Then, targeted high-throughput sequencing of candidate disease-causing genes was carried out in the large-scale case sample library of Chinese yellow-race patients accumulated over the years, and bioinformatics analysis was carried out to screen and find disease-causing mutations. Cases with pathogenic mutations were screened for follow-up and further family analysis, and the entire family of the patient was seq...

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Abstract

The invention discloses a DNA library for detecting MO pathogenic gene mutations through a targeted high-throughput semiconductor sequencing technique and application of the DNA library. Specifically, a primer pool is designed according to 15 MO pathogenic genes, sample genome DNA is subjected to super-multiplex PCR amplification, and an amplified product is sequenced by using the high-throughput semiconductor sequencing technique to find pathogenic mutations so as to provide a theoretical basis of genetics and molecular biology for clinical diagnosis. The DNA library for detecting the MO pathogenic genes disclosed by the invention has the characteristics of accuracy, rapidness, flexibility and low cost, and 15 gene detection regions related in the invention comprehensively cover all known pathogenic genes of MO, thereby having important significance and clinical values in diagnosis and differential diagnosis of MO.

Description

technical field [0001] The invention relates to a DNA library for detecting and diagnosing multiple osteochondroma pathogenic genes through targeted high-throughput semiconductor sequencing technology and its application. Specifically, based on the pathogenic genes of multiple osteochondroma, supermultiplex PCR primers that can cover the exons and adjacent regions of the above genes are designed, and supermultiplex PCR amplification is performed on the sample genomic DNA, and the amplified products are sequenced using high-throughput Sequencing technology is used to search for pathogenic mutations, clarify the genetic etiology of multiple osteochondroma, and provide a theoretical basis for genetics and molecular biology for clinical diagnosis. It belongs to the gene detection technology in the clinical detection technology in the field of biomedicine. Background technique [0002] Multiple osteochondromas (Multiple osteochondromas, MO), also known as multiple exostoses, is a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C40B40/08C12Q1/6869C12Q1/6886C12Q1/6806C12M1/34
CPCC12Q1/6806C12Q1/6869C12Q1/6886C40B40/08C12Q2537/143C12Q2563/159C12Q2535/122C12Q2525/191C12Q2521/301
Inventor 汪道文周世媛李宗哲
Owner 汪道文