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PCR (polymerase chain reaction) primer for amplifying PKD (polystic kidney disease)1 exon ultra-long fragment, kit for detecting PKD1 gene mutation and application

An amplification primer and kit technology, applied in the field of genetic disease gene detection, can solve problems such as large workload and complicated procedures, and achieve the effects of large workload, simplified operation process, and reduced cost

Active Publication Date: 2017-08-01
THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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Problems solved by technology

The disadvantage of this method is: it needs to use 5-8 pairs of LR-PCR primers to amplify, and if any LR-PCR amplification fails, it needs to be re-prepared, and the LR-PCR product needs to be gel-cut, purified and accurately quantified; The pre-processing workload of this method is large and the procedure is complicated

Method used

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  • PCR (polymerase chain reaction) primer for amplifying PKD (polystic kidney disease)1 exon ultra-long fragment, kit for detecting PKD1 gene mutation and application
  • PCR (polymerase chain reaction) primer for amplifying PKD (polystic kidney disease)1 exon ultra-long fragment, kit for detecting PKD1 gene mutation and application
  • PCR (polymerase chain reaction) primer for amplifying PKD (polystic kidney disease)1 exon ultra-long fragment, kit for detecting PKD1 gene mutation and application

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Embodiment Construction

[0051] Preferred embodiments of the present invention will be described in more detail below with reference to the accompanying drawings.

[0052] Reagents used:

[0053] KOD FX Neo DNA Polymerase, Ion Ampliseq Library kit 2.0, Ion PGM TemplateOT2 200kit v3, Ion PGM Sequencing 200kit v2, Ion xpress Barcorde Adaptor33-48kit, Ion 316chip kit v2 BC.

[0054] 1. Extract the whole genome DNA of the sample to be tested;

[0055] Use OMEGA "Genomic DNA Extraction Kit" to extract peripheral blood DNA while removing RNA. The total volume of extracted DNA is about 100ul, the concentration is 20-200ng / μl; A260 / 280 is between 1.8-2.0.

[0056] 2. Ultra-long fragment PCR amplification of exon 2 to exon 46 of PKD1 gene;

[0057] The PCR amplification reagent uses KOD FXNeo (TOYOBO Life Science) DNA polymerase, which can amplify a fragment of up to 40kb using genomic DNA as a template. It has the characteristics of high amplification efficiency and high fidelity, and is suitable for fragm...

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Abstract

The invention relates to the field of disease transmission gene detection, in particular to a PCR (polymerase chain reaction) primer for amplifying a PKD (polystic kidney disease)1 exon ultra-long fragment, a kit for detecting PKD1 gene mutation and application. PKD1 exons are 2nd to 46th exons; the PCR primer has the primer with the base sequences shown by SEQ ID NO:1 and SEQ ID NO:2. The kit has the advantages that the diagnosis accuracy is high; the operation flow process is simple, convenient and efficient.

Description

technical field [0001] The present invention relates to the field of gene detection for genetic diseases, more specifically, to a PCR primer for amplifying PKD1 exon ultra-long fragments, a kit for detecting PKD1 gene mutations, and the PCR primer and kit as a method for adult polycystic polycystic Application of detection reagents for nephropathy-causing genes. Background technique [0002] Polycystic kidney disease (PKD) is a hereditary kidney disease in which multiple cystic masses occur in bilateral renal parenchyma. According to the age of onset and genetic characteristics, it can be divided into two types: autosomal recessive (infantile) polycystic kidney disease; autosomal dominant genetic (adult type) polycystic kidney disease. Adult-type polycystic kidney disease is relatively common, with an incidence rate of 1 / 400-1 / 1000 in the population; adult-type polycystic kidney disease is an autosomal dominant genetic disease, which is characterized by familial aggregation...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/156C12Q2600/16C12Q2535/122C12Q2537/143
Inventor 梅世月孔祥东白楠胡爽
Owner THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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