DNA (Deoxyribonucleic Acid) library for detecting cholestatic jaundice pathogenic genes and application thereof

A DNA library, cholestasis technology, applied in the field of DNA library, can solve the problems of indistinguishable, unable to meet the requirements of multi-sample detection timeliness, low throughput, etc.

Active Publication Date: 2015-01-28
TONGJI HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI TECH
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Problems solved by technology

However, at present, the combination of clinical symptoms, biochemical and pathological tests sometimes cannot distinguish the two well; at the same time, the gene detection method based on Sanger sequenci

Method used

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  • DNA (Deoxyribonucleic Acid) library for detecting cholestatic jaundice pathogenic genes and application thereof
  • DNA (Deoxyribonucleic Acid) library for detecting cholestatic jaundice pathogenic genes and application thereof
  • DNA (Deoxyribonucleic Acid) library for detecting cholestatic jaundice pathogenic genes and application thereof

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specific Embodiment approach

[0054] 1. Reagents used in the method:

[0055] Ion AmpliSeq TM Library Kit 2.0, Ion PGM TM Template OT2200Kit v3, Ion Sequencing 200Kit v2, Ion Xpress Barcode Adapters 1-16Kit, Ion 318 TM Chip Kit v2

[0056] 2. Specimen collection and storage

[0057] (1) Specimen collection: The specimen is the peripheral blood of the patient. Blood is 5ml of venous blood taken routinely, treated with EDTA anticoagulant.

[0058] (2) Storage: It can be detected immediately, stored at 4°C for one week, and stored at -80°C for more than one week.

[0059] 3. Detection steps and result analysis:

[0060] (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.

[0061] (2) Super multiplex PCR amplification and library building of the target detection region: the whole exons of 60 genes involved in the present invention are used...

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Abstract

The invention discloses a DNA (Deoxyribonucleic Acid) library for detecting cholestatic jaundice pathogenic genes by virtue of a high-throughput sequencing technology and application thereof and in particular relates to a super-multiplet PCR primer which can cover the exon and adjacent areas of the genes according to 60 cholestatic jaundice pathogenic genes. The genome DNA of the sample is subjected to super-multiplet PCR amplification, the amplification product is sequenced by utilizing the high-throughput sequencing technology, the pathogenic mutation is searched, the genetic etiology of the cholestatic jaundice is clear, and a theoretical basis of genetics and molecular biology is provided for clinical diagnosis. The DNA library has the characteristics of accuracy, flexibility, rapidness and low cost. Moreover, according to the detection area of 60 genes, 15 common cholestatic genetic defects can be detected, and the DNA library has significance and clinical values on differential diagnosis of the cholestatic jaundice.

Description

technical field [0001] The invention relates to a DNA library for detecting cholestatic jaundice pathogenic genes by high-throughput sequencing technology and application thereof. Specifically, based on the causative gene of cholestatic jaundice, design super-multiplex PCR primers that can cover the exons and adjacent regions of the above genes, and perform super-multiplex PCR amplification on the sample genomic DNA, and use high-throughput sequencing technology to amplify the products Sequencing is performed to find pathogenic mutations, clarify the genetic etiology of cholestatic jaundice, 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] Cholestatic jaundice is a common clinical symptom in infancy, and its incidence is about 4 / 10000 live births. There are many causes of cholestatic jaundice in...

Claims

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Application Information

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IPC IPC(8): C40B40/06C12Q1/68
Inventor 汪道文王宏
Owner TONGJI HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI TECH
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