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Method for detecting plant-derived components of kusnezoff monkshood roots

A technology of plant source and Aconitum aconitum, applied in biochemical equipment and methods, measurement/inspection of microorganisms, DNA/RNA fragments, etc., can solve problems such as lack of plant source components of Aconitum aconitum

Inactive Publication Date: 2021-07-23
青岛谱尼测试有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, there is no real-time fluorescent PCR detection method for the botanical components of Aconitum aconitum, and there is an urgent need to develop a rapid and simple technology for the detection of botanical components

Method used

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  • Method for detecting plant-derived components of kusnezoff monkshood roots
  • Method for detecting plant-derived components of kusnezoff monkshood roots

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] 1. Primers and probes used:

[0037] According to the gene sequence of Aconitum aconitum NADH dehydrogenase subunit 3 (NADH dehydrogenase subunit 3), design and synthesize specific amplification primers and probes for real-time fluorescent PCR:

[0038] Upstream primer: 5'-ATGTCAGAATTTGCACCTATTTGTAT-3',

[0039] Downstream primer: 5'-CCTATCCAGAAAAATTGTCGGC-3',

[0040] The fluorescent probe is: 5'-TATTTAGTGATCAGTCCGCTA-3'.

[0041] The above-mentioned fluorescent probes have a FAM fluorescent label at the 5' end and a MGB fluorescent label at the 3' end.

[0042] 2. The extraction and determination of sample DNA are carried out according to the following steps:

[0043] (1) Weigh 0.5g Aconitum aconitum and put it into a 50mL centrifuge tube;

[0044] (2) Add preheated 10mL CTAB extraction and lysis buffer (add 20µL proteinase K, 20µL RNA, 200µL β-mercaptoethanol before use), shake and suspend, and incubate at 65°C for 1h, shake continuously during the period, cool t...

Embodiment 2

[0065] 1. Primers and probes used:

[0066] According to the gene sequence of Aconitum aconitum NADH dehydrogenase subunit 3 (NADH dehydrogenase subunit 3), design and synthesize specific amplification primers and probes for real-time fluorescent PCR:

[0067] Upstream primer: 5'-ATGTCAGAATTTGCACCTATTTGTAT-3',

[0068] Downstream primer: 5'-CCTATCCAGAAAAATTGTCGGC-3',

[0069] The fluorescent probe is: 5'-TATTTAGTGATCAGTCCGCTA-3'.

[0070] The above-mentioned fluorescent probes have a FAM fluorescent label at the 5' end and a MGB fluorescent label at the 3' end.

[0071] 2. The extraction and determination of sample DNA are carried out according to the following steps:

[0072] (1) Weigh 0.5g of Aconitum samples, such as Aconitum aconitum, Aconitum japonica, Aconitum japonica and Aconitum melon-leaf, etc., and put the pulverized materials into different 50mL centrifuge tubes;

[0073] (2) Add preheated 10mL CTAB extraction and lysis buffer (add 20µL proteinase K, 20µL RNA, ...

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Abstract

The invention provides a primer and probe for detecting plant-derived components of kusnezoff monkshood roots by a real-time fluorescent PCR method, and a specific detection method. High sensitivity and specificity are achieved, and the primer and probe and the specific detection method can be used for rapidly detecting the plant-derived components of kusnezoff monkshood roots.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to a detection method of aconitum aconitum plant source components, in particular to a primer, a probe and a detection method for detecting the plant source components of aconitum aconitum. Background technique [0002] Aconitum aconitum is a plant of Ranunculaceae Aconitum aconitum ( Aconitum kusnezoffii Reichb. ) dry tuber root, pungent, bitter, hot in nature, has the effect of expelling wind and dampness, warming meridian and relieving pain, used for anemofrigid-damp arthralgia, joint pain, cold pain in trusted subordinates, cold colic pain and anesthesia pain relief. Although Aconitum aconitum has medicinal value, it is easy to be poisoned due to the difficulty in controlling the poisoning dose and therapeutic dose when used. The main ingredient that causes Aconitum poisoning is aconitine, which is extremely toxic to humans. Ordinary people will experience palpitation, heart ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/686C12N15/11
CPCC12Q1/6895C12Q1/686C12Q2600/166
Inventor 张斌王霞张艳红
Owner 青岛谱尼测试有限公司