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

Fluorescent PCR detection primer set, probe composition, kit, detection method for simultaneous identification of bupleurum scorzonerifolium and bupleurum chinense and application

A detection kit and detection primer technology, applied in the field of molecular biology detection and detection, can solve the problems of large limitations, high cost of supporting instruments, complicated identification operations, etc., and achieve significant differences between species, high accuracy and specificity. strong effect

Inactive Publication Date: 2017-08-18
BIOTECH RES CENT SHANDONG ACADEMY OF AGRI SCI
View PDF2 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the molecular means of research on the identification of Bupleurum at home and abroad is mainly DNA barcode technology, and the identification operation is complicated by comparing the ITS sequence of Bupleurum by DNA sequencing means, the cost of equipment is high, and the limitations are large

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
  • Fluorescent PCR detection primer set, probe composition, kit, detection method for simultaneous identification of bupleurum scorzonerifolium and bupleurum chinense and application
  • Fluorescent PCR detection primer set, probe composition, kit, detection method for simultaneous identification of bupleurum scorzonerifolium and bupleurum chinense and application
  • Fluorescent PCR detection primer set, probe composition, kit, detection method for simultaneous identification of bupleurum scorzonerifolium and bupleurum chinense and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] 1. DNA extraction of Bupleurum and counterfeit samples:

[0059] The plant DNA extraction kit was used for extraction, and the specific operation steps were found in the kit instruction manual. The purity and concentration of the extracted genomic DNA were determined by UV spectrophotometer. The measured OD260 / OD280 values ​​are all around 1.8-1.9, and the concentration is above 10ng / μL, indicating that the DNA is of high purity and moderate concentration, which meets the requirements of PCR amplification.

[0060] 2. Selection of target genes and design of primers:

[0061] The ITS2 sequence is a non-coding sequence between the 5.8S and 28S rRNA genes. Because of its special structure and function in the overall ribosomal rDNA, it is both conservative and highly variable. There may be large differences among different species or individuals of the same species. Therefore, using the ITS2 sequence as the identification target gene of Bei Bupleurum has the characteristi...

Embodiment 2

[0071] Example 2 specificity verification

[0072] Utilizing the primers and probes designed by the present invention, Bupleurum bupleurum, Bupleurum chinensis, Bupleurum chinensis Yinzhou, Bupleurum xiaoyehei, Bupleurum coniferae, Bupleurum bamboo leaves, Bupleurum membranaceus, Bupleurum bupleurum, Narrow The total genomic DNA of plants such as Bupleurum bupleurum, Atractylodes macrocephala, ginseng, Codonopsis pilosula, habitat, dendrobium, yam, Millae vine, Chuanbei, and wolfberry were used as templates for real-time fluorescent PCR detection to verify the specificity of their primers and probes. The results are shown in Table 4 and Figure 4 with Figure 5 , the results show that the probes and primers designed in this study have strong specificity.

[0073] Table 4. Specificity Verification Tests

[0074]

[0075]

Embodiment 3

[0076] Embodiment 3 Sensitivity experiment

[0077] Quantify the genomic DNA of South and North Bupleurum to 50ng, dilute it in a 5× gradient, and take 2.0 μL for each gradient as the template amount (ie: 10ng, 2ng, 0.4ng, 0.08ng, 0.016ng) for real-time fluorescence quantification PCR detection to assess the detection limit of the present invention. See Image 6 with Figure 7 The result shows that the detection limit of the method is 0.08ng, which shows that the method provided by the invention has high sensitivity.

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 set, a probe composition, a kit and a detection method for simultaneous identification of bupleurum scorzonerifolium and bupleurum chinense and application. According to the invention, specific primers of bupleurum scorzonerifolium and bupleurum chinense are employed for PCR amplification of a DNA extract of a to-be-detected sample, bupleurum universal primers are designed as the quality control, accurate target sites are provided for detection and identification of genuine bupleurum scorzonerifolium and bupleurum chinense, thus achieving fast and accurate identification. The detection method provided by the invention is simple, quick and effective, and can quickly and accurately identify the components of the genuine bupleurum scorzonerifolium and bupleurum chinense.

Description

technical field [0001] The invention relates to a detection method, in particular to a fluorescent PCR detection primer set, a probe composition, a kit, a detection method and an application for simultaneously identifying southern and northern Bupleuri. The invention belongs to the technical field of molecular biology detection. Background technique [0002] Bupleurum, the dried root of Bupleurum L. in the Umbelliferae family, was first recorded in "Shen Nong's Materia Medica", formerly known as Cihu, and listed as top grade. It was renamed Bupleurum in the "Bencao Tujing". It has the effects of resolving the exterior and harmonizing the interior, soothing the liver and relieving stagnation, and elevating yang qi. It is the first-choice essential medicine for the treatment of Shaoyang syndrome in traditional Chinese medicine. The 2015 edition of "Chinese Pharmacopoeia" stipulates that Bupleurum chinense DC. and Bupleurum scorzonerifolium Willd. (South Bupleurum) are genuin...

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): C12Q1/68C12N15/11
CPCC12Q1/6851C12Q1/6895C12Q2537/143C12Q2545/113C12Q2561/101C12Q2563/107
Inventor 步迅刘艳艳胡悦范阳阳谭晴晴张全芳
Owner BIOTECH RES CENT SHANDONG ACADEMY OF AGRI SCI
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