Peronophythora litchii LAMP (loop-mediated isothermal amplification) primer and rapid detection method thereof
A technique for detection of Pythophthora litchie and its detection method, which is applied in the field of LAMP primers and rapid detection of Pythora litchie, can solve the problems of poor specificity, long period, and low sensitivity of the detection method, and achieve strong specificity, reliable results, and high sensitivity. high effect
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0049] Embodiment 1: LAMP primer is to the specific amplification of litchi peronosophthora
[0050] 1. Specific detection of LAMP on Lychee peronosophthora
[0051] ①LAMP reaction system: 25.0 μL: including 0.2 μM each of F3 and B3, 1.6 μM each of FIP and BIP, 1× ThermopolBuffer, 1.0 mM dNTPs, 0.8 M betaine, 6.0 mM magnesium sulfate, 0.1% Tween-20,
[0052] 8U Bst The DNA polymerase is 50ng template DNA, 50μM calcein, 500μM manganese chloride, and the insufficient part is made up with sterile double distilled water.
[0053] The LAMP reaction condition is to incubate at 63° C. for 60 minutes, and then incubate at 80° C. for 5 minutes.
[0054] ②After the LAMP reaction is completed, if green fluorescence is observed in the color development results, it is judged as positive, and orange is judged as negative; or take 2 μL of the amplification product and use 2.0% agarose gel electrophoresis to detect it. If the characteristic trapezoidal band of LAMP appears, it is judged...
Embodiment 2
[0057] Embodiment 2: Sensitivity detection of LAMP primers to Lychee peronosophthora
[0058] 1. Sensitivity Detection of LAMP on Lychee Peronosporae
[0059] Using 10-fold concentration serial dilution method, the extracted DNA of Peronospora litchie was diluted to 1ng / μL, 100pg / μL, 10pg / μL, 1pg / μL, 100fg / μL, 10fg / μL, 1fg / μL, 100ag / μL, a total of 8 different concentration gradients.
[0060] ①LAMP reaction system is 25uL: including 0.2μM each of F3 and B3, 1.6μM each of FIP and BIP, 1×ThermopolBuffer, 1.0mMdNTPs, 0.8M betaine, 6.0mM magnesium sulfate, 0.1%Tween-20,
[0061] 8U Bst The DNA polymerase is 1 μL template DNA, 50 μM calcein, 500 μM manganese chloride, and the insufficient part is made up with sterile double distilled water.
[0062] The LAMP reaction condition is to incubate at 63° C. for 60 minutes, and then incubate at 80° C. for 5 minutes.
[0063] ②After the LAMP reaction is completed, if green fluorescence is observed in the color development results, ...
Embodiment 3
[0065] Embodiment 3: detection of litchi downy mildew in pathogenic tissue
[0066] 1. Sample collection: Plant tissue samples were collected from the litchi production base in Fuzhou, Fujian.
[0067] 2. DNA extraction and detection
[0068] The DNA of P. litchi was extracted from the diseased plant tissue by NaOH rapid lysis method.
[0069] Perform LAMP detection as follows:
[0070] ①LAMP reaction system is 25 μL: including 0.2 μM each of F3 and B3, 1.6 μM each of FIP and BIP, 1× ThermopolBuffer, 1.0 mM dNTPs, 0.8 M betaine, 6.0 mM magnesium sulfate, 0.1% Tween-20,
[0071] 8U Bst The DNA polymerase is 1 μL template DNA, 50 μM calcein, 500 μM manganese chloride, and the insufficient part is made up with sterile double distilled water.
[0072] The LAMP reaction condition is to incubate at 63° C. for 60 minutes, and then incubate at 80° C. for 5 minutes.
[0073] ② After the LAMP reaction is completed, if green fluorescence is observed in the color development res...
PUM

Abstract
Description
Claims
Application Information

- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
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