Primers for detection of Phytophthora cowpea lamp and its detection method

A technology for detecting primers and Phytophthora, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problems of long cycle, poor specificity and low sensitivity of detection methods, and achieve reliable results, strong specificity, The effect of high sensitivity

Active Publication Date: 2018-10-02
INST OF PLANT PROTECTION FAAS
View PDF3 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The purpose of the present invention is to provide a kind of Phytophthora cowpea in the prior art for the traditional detection method of Phytophthora cowpea required period is long, detection method specificity is poor, PCR detection needs to rely on equipment such as amplification instrument, and sensitivity is low, provides a kind of Phytophthora cowpea LAMP detection primers and a detection method with reliable results, easy operation, strong specificity and high sensitivity

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 detection of Phytophthora cowpea lamp and its detection method
  • Primers for detection of Phytophthora cowpea lamp and its detection method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Specific amplification of LAMP primers to Phytophthora cowpea

[0043] 1. LAMP Specific Detection of Phytophthora vignacea

[0044] ①LAMP reaction system: In the 25 μl reaction system, the concentration of F3 and B3 primers is 0.2 mmol / L, the concentration of FIP and BIP primers is 1.6 mmol / L, 20mM Tris-HCl, 10mM (NH 4 ) 2 SO 4 , 10mM KCl, 8mM MgSO 4 , betaine 0.8mol / L, Bst 8 U of polymerase, 1.0 mmol / L of dNTPs, 50 μmol / L of calcein, 500 μmol / L of manganese chloride, 0.1% of TWeen-20, 50 ng of template DNA, and make up the deficiency with sterile double distilled water; LAMP reaction The conditions were incubation at 65°C for 50 min and inactivation at 85°C for 10 min.

[0045] ②After the LAMP reaction, green fluorescence was observed as a positive color, and orange was negative. Alternatively, take 2 μl of the amplification product and use 2% agarose gel electrophoresis to detect it. If the characteristic trapezoidal band of LAMP appears, it is jud...

Embodiment 2

[0048] Embodiment 2: Sensitivity detection of LAMP primers to Phytophthora cowpea

[0049] 1. LAMP Sensitive Detection of Phytophthora vignacea

[0050] The extracted P. cowpea DNA was diluted to 100 ng, 10 ng, 1 ng, 100 pg, 10 pg, 1 pg, 100 fg, 10 fg, 1 fg / μL by 10-fold concentration serial dilution method, a total of 9 different concentrations gradient.

[0051] ①LAMP reaction system: In the 25 μl reaction system, the concentration of F3 and B3 primers is 0.2 mmol / L, the concentration of FIP and BIP primers is 1.6 mmol / L, 20mM Tris-HCl, 10mM (NH 4 ) 2 SO 4 , 10mM KCl, 8mM MgSO 4 , betaine 0.8mol / L, Bst 8 U of polymerase, 1.0 mmol / L of dNTPs, 50 μmol / L of calcein, 500 μmol / L of manganese chloride, 0.1% of TWeen-20, 100 ng of template DNA, and make up the deficiency with sterile double distilled water; LAMP reaction conditions Incubate at 65°C for 60 minutes and inactivate at 85°C for 5 minutes.

[0052] ②After the LAMP reaction, green fluorescence was observed as a...

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 phytophthora vignae LAMP (loop-mediated isothermal amplification) detection primers and a phytophthora vignae LAMP detection method which are specially applied to specific detection of phytophthora vignae. The phytophthora vignae LAMP primers are mainly adopted and designed, and green fluorescence can be observed or an LAMP characteristic ladder can appear through the chromogenic reaction or agarose gel electrophoresis detection after LAMP. The LAMP primers and a usage method thereof can be applied to rapid, sensitive and accurate detection of the cowpea diseases in production practice, can also be applied to early diagnosis of diseases in fields as well as monitoring and identification of germs, and provide reliable technical and theoretical bases for control of the cowpea diseases.

Description

technical field [0001] The primers for detection of Phytophthora cowpea LAMP and the detection method thereof of the present invention are specially used for high-sensitivity and fast LAMP detection of Phytophthora cowpea, and can be used for early diagnosis of cowpea blight in the field and monitoring and identification of pathogens, and belong to the detection, identification and control technology of crop diseases. field. Background technique [0002] by Phytophthora vigna ( Phytophthora vignae ) infection of cowpea blight is the main disease of cowpea adult plants, the suitable temperature for onset is 25-28 ℃, and with the increase of rainwater and humidity, the disease will become more serious, so the period of continuous cloudy rain and plum rain in spring is the high incidence of disease Expect. Secondly, if the ventilation is not good, the drainage is poor, the application of immature basal fertilizer or continuous cropping, the disease will be more serious. The ...

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 Patents(China)
IPC IPC(8): C12Q1/6895C12Q1/04C12N15/11C12R1/645
CPCC12Q1/6844C12Q1/6895C12Q2531/119
Inventor 陈庆河翁启勇李本金刘裴清刘小丽
Owner INST OF PLANT PROTECTION FAAS
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap