Digoxin probe for human-derived MANF gene detection and application thereof

A technology of gene detection and digoxin, which is applied in the field of gene technology, can solve problems such as hybridization probes, and achieve the effects of improving detection sensitivity, convenient and simple operation, and strong specificity

Inactive Publication Date: 2017-09-22
SHANGHAI TONGJI HOSPITAL
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This method has high detection specificity, and there is curren

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  • Digoxin probe for human-derived MANF gene detection and application thereof
  • Digoxin probe for human-derived MANF gene detection and application thereof

Examples

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Embodiment 1

[0031] This embodiment relates to a method for determining virus titer by dot hybridization, comprising the following steps:

[0032] 1. DNA extraction of rAAV8-MANF virus

[0033] Viral DNA was extracted with a commercial kit, and the concentration was determined to control the extraction quality OD260 / 280 to 1.8. In this example, Tiangen virus DNA extraction kit was used, and the specific operation was carried out according to the instructions.

[0034] 2. Dot hybridization

[0035] In this example, Roche DIG High Prime DNA Labeling and Hybridization Detection Kit II was used. The specific operation is as follows:

[0036] 1. DNA immobilization

[0037] (1) Preparation of standard products: serially dilute the linearized human-derived MANF recombinant plasmid standard products by 2 times;

[0038] (2) Sample pretreatment: Denature the standard and sample DNA at 95°C for 5 minutes, then place on ice to cool;

[0039] (3) Spotting: Take 1 ul of each DNA sample and spot i...

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Abstract

The invention discloses a Digoxin probe for human-derived MANF gene detection and application thereof. The sequence of the Digoxin probe is shown as SEQ NO:1. The invention further relates to the application of the Digoxin probe for human-derived MANF gene detection in a recombinant virus titer detection method. The invention further relates to a dot blot hybridization detection method which involves the Digoxin probe in claim 1. The Digoxin probe for human-derived MANF gene detection is creatively designed for the first time in at home and has high specificity. The designed Digoxin probe can be applied to relevant recombinant virus titer detection, has high detection specificity and sensitivity and brings great convenience to relevant experimental study.

Description

technical field [0001] The invention belongs to the field of gene technology and relates to a probe, in particular to a digoxin probe for human-derived MANF gene detection and an application thereof. Background technique [0002] Neurotrophic factors play an important role in the survival, growth, and differentiation of nerve cells, so they are used in research on neuroprotection, regeneration, and functional restoration. Mesencephalic Astrocyte-derived Neurotrophic Factor (MANF) was first isolated in 2003 from the culture medium of rat midbrain type I astrocytes subcultured in vitro. The molecular weight is 20kDa, which belongs to the conservative MANF protein family. The protein is a secreted protein, mainly expressed in neurons, and to a lesser extent in glial cells. Studies have shown that cerebral ischemic injury and a variety of endoplasmic reticulum stress in nerve cells can up-regulate the expression of MANF. In vitro experiments have confirmed that MANF can promot...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
Inventor 靳令经蒋明王玏朱玮符星
Owner SHANGHAI TONGJI HOSPITAL
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