Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Primer combination for PCR (Polymerase Chain Reaction) identification of zaocys dhumnade medicinal material, standard decoction and traditional Chinese medicine formula granules as well as application and identification method of primer combination

A combination technology of traditional Chinese medicine formula granules and primers, which is applied in biochemical equipment and methods, recombinant DNA technology, microbial measurement/inspection, etc., can solve the problems of DNA loss of form and difficulty in identification, and achieve the effect of overcoming the interference of excipients

Pending Publication Date: 2022-06-24
GUANGDONG YIFANG PHARMA
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The technical problem to be solved by the present invention is to provide a primer combination for PCR identification of black snake medicinal materials, standard decoctions and traditional Chinese medicine formula granules, which has good specificity and can effectively solve the problem of difficult identification after DNA loses its form

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Primer combination for PCR (Polymerase Chain Reaction) identification of zaocys dhumnade medicinal material, standard decoction and traditional Chinese medicine formula granules as well as application and identification method of primer combination
  • Primer combination for PCR (Polymerase Chain Reaction) identification of zaocys dhumnade medicinal material, standard decoction and traditional Chinese medicine formula granules as well as application and identification method of primer combination
  • Primer combination for PCR (Polymerase Chain Reaction) identification of zaocys dhumnade medicinal material, standard decoction and traditional Chinese medicine formula granules as well as application and identification method of primer combination

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Example 1 Primer Design

[0068] Based on the NADH; Cytochrome Oxidase I; 12S rRNA; 16S rRNA; Cytochrome b sequence analysis of the Chinese Pharmacopoeia 2020 edition, the Cytochrome Oxidase I sequences of common snake species in the GeneBank database were homologous using BioEdit software Align and proofread the specific SNP site of the snakehead snake, and import the base sequence containing the SNP site into Primer Premier 5 software for primer design.

[0069] After the sequence comparison, it was found that the specific site of the black snake is T and the other is A or C. After determining the SNP site, make the SNP site close to the 3' end of the forward primer, and adjust the primer score by moving the upstream primer position. GC content, and with the help of PrimerPremier 5 software, the position of the reverse primer was further adjusted, and the optimal combination was determined by the final primer score and the product band score. In the design process, i...

Embodiment 2

[0072] Example 2 Establishment of identification method

[0073] (1) DNA extraction and concentration adjustment

[0074]Take 0.5 g of the dried sample, grind it into powder, put it in a 2 mL centrifuge tube, add 1.5 mL of CTAB precipitation solution preheated at 65 °C, 20 μL of proteinase K, mix well, heat it in a water bath at 65 °C for 60 min, cool it to room temperature, and use 10000 r Centrifuge at / min for 5 min, discard the supernatant, add 900 μL CTAB precipitation solution and 20 μL proteinase K, and operate in the same way. Add 900 μL of LCTAB extract and 10 μL of β-mercaptoethanol to the centrifuge tube in turn, mix well, heat in a water bath at 65°C for 120 min, centrifuge, and cool down to room temperature to take the supernatant; add an equal volume of chloroform-isoamyl alcohol (24:1 ), shake for 3 minutes, and mix well; centrifuge at 12000 r / min at 4°C for 10 minutes, take 750 μL of the supernatant into a new 2 mL centrifuge tube, add an equal volume of chlor...

Embodiment 3

[0086] Embodiment 3 Identification method verification

[0087] 14 snake samples of known species were selected (specifically shown in Table 1).

[0088] The samples were identified using the identification method established in Example 2. Among them, in the PCR system, the amount of DNA template is 50ng.

[0089] Table 1 Sample Sheet

[0090]

[0091]

[0092] Identification results such as figure 1 As shown in the figure, it can be seen from the figure that there is a 135bp band in the Wushao snake medicinal material, the Wushao snake standard decoction (lyophilized powder), and the Wushao snake traditional Chinese medicine formula granules; while other samples did not get this band. This result shows that the accuracy rate of the identification method in the present invention reaches 100%.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a primer combination for PCR (Polymerase Chain Reaction) identification of zaocys dhumnade medicinal materials, standard decoction and traditional Chinese medicine formula granules, the primer combination comprises a primer I and a primer II, the primer I is a DNA (Deoxyribose Nucleic Acid) molecule with a nucleotide sequence as shown in SEQ ID NO: 1, and the primer II is a DNA molecule with a nucleotide sequence as shown in SEQ ID NO: 2. The invention further discloses application of the primer combination and an identification method based on the primer combination. By implementing the method, the zaocys dhumnade medicinal material, the standard decoction and the traditional Chinese medicine formula granules can be effectively identified.

Description

technical field [0001] The invention relates to the technical field of identification of traditional Chinese medicines, in particular to a primer combination used for PCR identification of Chinese herbal medicines, standard decoctions and traditional Chinese medicine formula particles, and an application and identification method thereof. Background technique [0002] There are more than 200 species of snakes in my country, of which more than 20 are common snakes, including Agkistrodonacutus, Zaocys dhumnades, Bungarus multicinctus, Bungarusfasciatus, Ptyas mucosus, round snake. Daboia russelii, Naja naja, Lycodon rufozonatus, Elaphe carinata, Ptyas korros, Gloydiusbrevicaudus, Elaphe taeniura, Elaphe moellendorffi, etc. Has clinical efficacy. Due to the similar shapes of various snakes, it is difficult to identify medicinal materials, especially after the internal organs are cut off during processing, they must be dried and blackened. After the standard decoction is further...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12Q1/6888C12Q1/686C12Q1/6806C12N15/11
CPCC12Q1/6888C12Q1/686C12Q1/6806C12Q2565/113C12Q2527/125C12Q2565/125
Inventor 罗宇琴魏梅陈向东李国卫范耀耀黄上书宋叶钟春琳
Owner GUANGDONG YIFANG PHARMA
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com