Carambola bacterial spot pathogenic bacteria molecule detection primer and detection method thereof

A technology for pathogenic bacteria and molecular detection, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc.

Inactive Publication Date: 2012-04-11
INST OF PLANT PROTECTION FAAS
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The present invention is aimed at the long period required for the biological detection method of carambola bacterial spot bacterium in the prior art, and the specificity of current carambola bacterial spot bacterium molecular detection method is poor, For the problem of low sensitivity, a specific molecular detection primer for carambola bacterial spot and a rapid molecular detection and diagnosis method for carambola bacterial spot with reliable results, easy operation, strong specificity and high sensitivity are provided

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
  • Carambola bacterial spot pathogenic bacteria molecule detection primer and detection method thereof
  • Carambola bacterial spot pathogenic bacteria molecule detection primer and detection method thereof
  • Carambola bacterial spot pathogenic bacteria molecule detection primer and detection method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1: Primer is to the specific amplification of carambola bacterial spot pathogen

[0031] 1. DNA extraction

[0032] Taking 13 strains of carambola bacterial spot collected from various places in Fujian Province (5 strains from Xindian Town, Fuzhou City, Fujian Province, 4 strains from Dongzhang Town, Fuqing City, Fujian Province, and 4 strains from Zini Town, Longhai City, Fujian Province) and 9 common pathogenic bacteria (Pseudomonas syringae pv. syringae, rice bacterial leaf sheath brown rot, cruciferous black rot, tomato bacterial pith necrosis, Xanthomonas, R. solanacearum, citrus canker Tomato solanacearum, Cucurbitaceae bacterial leaf spot) were used as test materials, and these strains were all preserved in the Molecular Pathology Laboratory of Plant Protection Research Institute, Fujian Academy of Agricultural Sciences; the genomic DNA of the tested strains was extracted by phenol and chloroform methods ;

[0033] 2. Specific Detection of Caram...

Embodiment 2

[0038] Embodiment 2: Sensitivity detection of primers to carambola bacterial spot pathogen

[0039] 1. Dilution of DNA concentration: according to the DNA extraction method of Example 1, the genomic DNA of carambola bacterial spot bacterium was extracted, and after the concentration was measured by a spectrophotometer, serial concentration dilution was adopted.

[0040] 2. Sensitive Detection of Carambola Bacterial Spot

[0041] PCR reaction system 25ul, including 2.5ul 10×PCR reaction buffer, 25mM Mg 2+ 2ul, 2.5mM dNTPs 2ul, 0.5U Taq DNA polymerase, 10uM primer PSaveF / PSaveR 0.4ul each and 10ng template DNA, d.d.H 2 O make up 25ul. The PCR reaction conditions were: pre-denaturation at 95°C for 3 min; denaturation at 94°C for 1 min, annealing at 60°C for 30 sec, and extension at 72°C for 45 sec; a total of 30 cycles; extension at 72°C for 10 min. Amplified products were detected by electrophoresis.

[0042] 3. Test result: if figure 2 As shown, in the 25ul reaction...

Embodiment 3

[0043] Example 3: Detection of carambola bacterial spot pathogen in diseased plant tissues.

[0044] 1. Sample collection: Plant tissue samples were collected from the base of Plant Protection Institute of Academy of Agricultural Sciences, Xindian Town, Fuzhou City, Fujian Province

[0045] 2. DNA extraction and detection

[0046] Collect carambola leaf spot with symptoms of carambola bacterial spot in the field and carambola leaf plant tissues without symptoms, use CTAB method to extract DNA, and perform PCR amplification according to the method implemented in the kit: PCR reaction system 25ul, including 2.5ul 10×PCR reaction buffer , 25mM Mg 2+ 2ul, 2.5mM dNTPs 2ul, 0.5U Taq DNA polymerase, 10uM primer PSaveF / PSaveR 0.4ul each and 10ng template DNA, d.d.H 2 O make up 25ul. The PCR reaction conditions were: pre-denaturation at 95°C for 3 min; denaturation at 94°C for 1 min, annealing at 60°C for 30 sec, and extension at 72°C for 45 sec for a total of 30 cycles; extens...

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 provides a molecule detection primer for carambola bacterial spot pathogenic bacteria and a detection method thereof. The detection primer comprises an upstream primer PSaveF: 5'----CTTATCGACGACTCAGCTGCG----3' and a downstreame primer PSaveR: 5'----TCATGCGTTGATCGTCAGGATC----3'. After being subject to PCR amplification and agarose gel electrophoresis, a specific amplification product with the fragment length of 373bp can be amplificated in a specific mode on carambola bacterial spot bacteria pure DNAs and diseased tissue with bacteria. The detection primer and the use method thereof in the invention can be used for detecting plant tissues infected by the carambola bacterial spot bacteria in a rapid, sensitive and specific mode in production practice, and can be used for early diagnosis of field diseases and monitoring and identification of bacteria, thereby providing reliable technical and theoretical basis for preventing and curing diseases caused by the carambola bacterial spot bacteria.

Description

technical field [0001] The invention relates to a primer for molecular detection of carambola bacterial spot pathogenic bacteria and a detection method thereof, and belongs to the field of detection, identification and control technologies of crop diseases. Background technique [0002] Carambola bacterial spot is caused by carambola bacterial spot pathogenic bacteria ( Pseudomonas syringae PV . averrhoi ) is a disease caused by infection. Starfruit (Carambola) belongs to the Oxalis family and belongs to tropical and subtropical evergreen tree fruit trees. It is widely planted in Guangdong, Hainan and other provinces of my country. In recent years, some carambola plantations or nurseries in Guangdong and Hainan provinces have discovered a leaf spot disease that causes severe leaf defoliation. Leaf spot tissue was cut out for microscopic examination, and bacterial overflow was found, which was considered to be a bacterial disease. It has not been reported at home and abr...

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 & AuthorityPatents(China)
IPC IPC(8): C12Q1/68C12Q1/04C12N15/11
Inventor陈庆河翁启勇林静李本金兰成忠
OwnerINST OF PLANT PROTECTION FAAS