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

Primers for detecting bacillus subtilis through loop-mediated isothermal amplification method and application thereof

A Bacillus subtilis, ring-mediated isothermal technology, applied in the direction of microorganisms, microorganism-based methods, biochemical equipment and methods, etc., can solve the problems of less development and application reports, achieve short detection cycle, strong specificity, and high results reliable effect

Active Publication Date: 2019-09-24
SHANDONG INST FOR PROD QUALITY INSPECTION +1
View PDF6 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are few reports on the development and application of Bacillus subtilis-related LAMP technology

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
  • Primers for detecting bacillus subtilis through loop-mediated isothermal amplification method and application thereof
  • Primers for detecting bacillus subtilis through loop-mediated isothermal amplification method and application thereof
  • Primers for detecting bacillus subtilis through loop-mediated isothermal amplification method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Extract the genomic DNA of the sample to be tested, use the extracted DNA as a template, add outer primer F3 / B3, inner primer FIP / BIP and loop primer LF / LB for LAMP amplification, the total volume of the LAMP detection reaction system is 20 μL, including 0.2 μM outer primer F3 / B3, 1.6 μM inner primer FIP / BIP and 0.4 μM loop primer LF / LB, LAMP reaction mixture 10 μL, DNA template 2.0 μL, make up to 20 μL with sterilized ultrapure water. React at 65°C for 35 minutes, and then detect the reaction result.

Embodiment 2

[0049] Extract the genomic DNA of the sample to be tested, use the extracted DNA as a template, add outer primer F3 / B3, inner primer FIP / BIP and loop primer LF / LB for LAMP amplification, the total volume of the LAMP detection reaction system is 20 μL, including 0.2 μM Outer primer F3 / B3, 1.6 μM inner primer FIP / BIP and 0.4 μM loop primer LF / LB, LAMP reaction mixture 10 μL, DNA template 2.0 μL, make up to 20 μL with sterilized ultrapure water. React at 61°C for 30 minutes, and then detect the reaction result.

Embodiment 3

[0051] Extract the genomic DNA of the sample to be tested, use the extracted DNA as a template, add outer primer F3 / B3, inner primer FIP / BIP and loop primer LF / LB for LAMP amplification, the total volume of the LAMP detection reaction system is 20 μL, including 0.2 μM outer primer F3 / B3, 1.6 μM inner primer FIP / BIP and 0.4 μM loop primer LF / LB, LAMP reaction mixture 10 μL, DNA template 2.0 μL, make up to 20 μL with sterilized ultrapure water. React at 63°C for 60 minutes, and then detect the reaction result.

[0052] The primers of Bacillus subtilis detected by the loop-mediated isothermal amplification technique in the above-mentioned embodiments 1-3, i.e. the nucleotide sequences of the outer primer F3 / B3, the inner primer FIP / BIP and the loop primer LF / LB are as follows:

[0053] F3: 5'-GCGATTCCAGTCACGG-3';

[0054] B3: 5'-CGTTTTTCGTTCCAATGATGA-3';

[0055] FIP: 5'-CGAATTGAGATAGCGATGTTCGTTTATGCGGAGCTTTACCCTCT-3';

[0056] BIP: 5'-ATATCCGCAACAATGGTCTAATTGCTGTTTTGTGCCGTCAG...

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 primers for detecting bacillus subtilis through a loop-mediated isothermal amplification method. The primers are designed and obtained with the gyrA gene and gyrB gene of the bacillus subtilis as target sequences and include a group of external primers F3 / B3, a group of internal primers FIP / BIP and a group of loop primers LF / LB. The invention further discloses application of the primers to detecting the bacillus subtilis. It is proved through experiments that the three groups of LAMP primers have quite high specificity and has the sensitivity at least one magnitude order higher than that of an existing method in a contrast test. The detecting process only needs to be conducted under the constant-temperature water bath conditions without the aid of expensive variable-temperature instruments, the result can be judged with naked eyes or by adding a simple indication agent, the reaction time is 40 minutes or shorter, the detection cycle is shorter, and the experiment cost is lower.

Description

technical field [0001] The invention belongs to the technical field of detection or identification of Bacillus subtilis in microbial fertilizers, and relates to a primer for rapid detection of Bacillus subtilis using loop-mediated isothermal amplification (LAMP) technology and an application thereof. Background technique [0002] At present, the detection methods for bacteria mainly include bacterial isolation and culture identification methods, immunological detection methods and molecular biology methods. [0003] The identification method of bacteria isolation and culture is to use liquid medium, semi-solid medium or solid medium to cultivate the isolated bacteria, and to diagnose the microorganisms through physiological, biochemical and staining microscopy. The disadvantage of this identification method is that the test cycle is very long. Generally, it takes 2 days to isolate and cultivate microorganisms, and the time required for identification is about one week. Ther...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/6844C12Q1/04C12N15/11C12R1/125
CPCC12Q1/6844C12Q1/689C12Q2531/119
Inventor 张娟杨春玉吕佩文程彬
Owner SHANDONG INST FOR PROD QUALITY INSPECTION
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