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

Qualitative and quantitative detection method for Xinjiang isolates of apricot chlorotic leaf roll (ACLR) phytoplasma

A quantitative detection method and quantitative detection technology, which are applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc. The qualitative and quantitative detection of Xinjiang isolates has achieved the effect of high sensitivity, improved sensitivity and strong specificity

Active Publication Date: 2020-03-20
XINJIANG AGRI UNIV
View PDF3 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, real-time fluorescent quantitative PCR detection methods have been established for the detection of phytoplasma in apricot chlorotic leaf rolls at home and abroad. Therefore, it is not suitable for the qualitative and quantitative detection of Xinjiang isolates of Phytoplasma chloroticum vol.
In addition, although a real-time fluorescent PCR detection method for members of the phytoplasma elm etiolation group (16SrⅤ) has also been established at home and abroad, this method can only achieve specific detection between phytoplasma groups, and cannot detect phytoplasma in the phytoplasma elm yellow group. distinguish and identify

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
  • Qualitative and quantitative detection method for Xinjiang isolates of apricot chlorotic leaf roll (ACLR) phytoplasma
  • Qualitative and quantitative detection method for Xinjiang isolates of apricot chlorotic leaf roll (ACLR) phytoplasma
  • Qualitative and quantitative detection method for Xinjiang isolates of apricot chlorotic leaf roll (ACLR) phytoplasma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] The preparation of embodiment 1 primer and probe

[0041] 1. Design and synthesis of primers and probes

[0042] The primers and probes used for the real-time fluorescence quantitative PCR detection of the phytoplasma chlorophyllum phytoplasma Xinjiang isolates of apricot chlorosis are designed and synthesized as follows: at first, the 16S rDNA nucleic acid sequences of all phytoplasma and representative bacteria are collected in NCBI, and the Omiga software compared the above sequences and analyzed the consistency to find out the genetic difference sites between the Xinjiang isolate of Phytoplasma chlorophyllum and other phytoplasma, and a pair of primers were screened out with the software Beacon Designer 7.0, and in the primer pair A fluorescent Taqman probe is set in the amplification region, the reporter fluorescent dye is labeled at the 5' end of the probe, and the quencher fluorescent dye is labeled at the 3' end of the probe. After the design of PCR primers and...

Embodiment 2

[0047] Example 2 Establishment of real-time fluorescent PCR detection method for apricot chlorotic phytoplasma Xinjiang isolate

[0048] 1. Extraction of total plant DNA: Plant Genomic DNA Kit (TIANGEN) was used to extract total DNA from phloem or leaf veins of apricot branches, and stored in a -40°C refrigerator for later use.

[0049] 2. PCR Amplification of 16S rDNA from Xinjiang Isolate of Phytoplasma Chlorophyta Curls from Apricot

[0050] The 16S rDNA gene fragment of Phytoplasma chlorophyll rolls was amplified with the universal primer pair R16mF2: 5'-CATGCAAGTCGAACGGA-3', R16mR2: 5'-CTTAACCCCAATCATCGA-3'. PCR reaction system (25µL): 2.0µL DNA template, 1.0µL each primer (10µmol / L), 1.0µL dNTP (10mmol / L), 10×PCR Buffer (2.5mmol / L MgCl 2 ) 2.5 µL, Taq DNA polymerase (2.5U / µL) 0.3µL, supplemented with ddH 2 0 to 25.0 µL. Reaction conditions: pre-denaturation at 94°C for 4min; denaturation at 94°C for 45s, renaturation at 50°C for 45s, extension at 72°C for 1min, 35 cy...

Embodiment 3

[0067] Example 3 Application of real-time fluorescent PCR detection method for apricot chlorotic leaf roll Phytoplasma Xinjiang isolate

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 belongs to the technical field of plant quarantine, and relates to a qualitative and quantitative detection method for Xinjiang isolates of apricot chlorotic leaf roll (ACLR) phytoplasma. According to the qualitative and quantitative detection method, according to the difference of the ACLR phytoplasma and other phytoplasma in sequence of a 16S rDNA gene, a primer pair ACLR-F1 / ACLR-R1 and a probe ACLR-Probe which are used for specific detection of the ACLR phytoplasma are designed, a cycle threshold (Ct) is determined through an established real-time fluorescent quantitative polymerase chain reaction (PCR) detection method for the ACLR phytoplasma, and qualitative detection of the ACLR phytoplasma can be realized according to the positive judgment standard of a real-time fluorescent quantitative PCR detection result; then, the segment copy concentration of the 16S rDNA genes of the ACLR phytoplasma in detected samples is worked out by a pre-established fluorescent quantitative PCR standard curve equation, and thus, the number of the ACLR phytoplasma in the samples is obtained, and thereby, quantitative detection is realized. The qualitative and quantitative detectionmethod has the advantages of high sensitivity, high specificity, high repeatability, high throughput, and the like, and can be widely applied to the fields of plant quarantine, plant protection, scientific research, and the like.

Description

technical field [0001] The invention belongs to the technical field of plant quarantine, and relates to a quantitative detection method for the Xinjiang isolate of apricot chlorotic leaf-curly phytoplasma, in particular to qualitative real-time fluorescent quantitative PCR technology for apricot chlorotic phytoplasma Xinjiang isolate , primers, probes and methods for quantitative detection. Background technique [0002] Apricot chlorotic leaf roll (ACLR) is a highly fatal phytoplasmic disease that harms apricot trees. China, Europe and North America have included it in the list of imported plant quarantine pests. The disease was first discovered on French apricot trees in 1973, and was subsequently found on other European stone fruit tree species such as almonds, European plums, cherries, peaches, and nectarines, and became a disease affecting the commercial cultivation of European stone fruit trees. main disease. Although the symptoms of apricot chlorotic leaf roll disea...

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): C12Q1/689C12Q1/6851C12Q1/06C12N15/11
CPCC12Q1/689C12Q1/6851C12Q2531/113C12Q2561/101C12Q2561/113C12Q2563/107Y02A50/30
Inventor 韩剑罗明艾克热木买买提张祥林唐章虎
Owner XINJIANG AGRI UNIV
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