Primer group and kit for detecting whether monochamus alternatus carries Bursaphelenchus xylophilus Nickle and application of primer group
A detection kit and the technology of A. pineapple are applied in the detection of the pine wood nematode carried by the pineapple beetle, and the application field in the detection of the pine wood nematode, which can solve the problem of long time required and poor repeatability of results. , with radioactivity and other problems, to achieve the effect of reducing prevention and control costs, high specificity, and easy operation
Pending Publication Date: 2017-09-01
INST OF FOREST ECOLOGY ENVIRONMENT & PROTECTION CHINESE ACAD OF FORESTRY
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Problems solved by technology
Among them, although DNA probe technology can effectively identify B. xylophilus, it is expensive and radioactive; RAPD technology analyzes the relationship between B. xylophilus and related species, and the result has poor repeatability; RFLP technology distinguishes B. xylophilus from P. Styloid nematodes, endonucleases are expensive and time-consuming; fluorescent PCR is sensitive and fast, but expensive; PCR technology is easy to operate, less expensive, but takes a long time
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Embodiment 1
[0036] Example 1 Establishment of a LAMP rapid detection method for Monochamus alternatus carrying pine xylophilus
[0037] 1. Materials and methods
[0038] 1.1 Extraction of Monochamus alternatus DNA
[0039] Take 0.2 g of head and thorax tissues of Monochamus alternatus adults, put them into a 2 mL EP tube, put them into a steel ball mill and grind them for 5 min (60 Hz); add 600 μL of 3M guanidine isothiocyanate extraction buffer, lyse at room temperature for 1 h, and mix by inversion during the period. Mix well; add 600 μL Tris-phenol chloroform isoamyl alcohol (25:24:1), centrifuge at 12000 r / min for 10 min; absorb the supernatant, add 600 μL chloroform isoamyl alcohol (24:1), centrifuge at 12000 r / min for 10 min; absorb the supernatant Add 600 μL of isopropanol to precipitate at -20°C for 20 minutes, centrifuge at 12,000 r / min for 10 minutes; discard the supernatant, wash the precipitate twice with 70% ethanol, and centrifuge at 12,000 r / min for 3 minutes each time; bl...
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The invention discloses a primer group and a kit for detecting whether monochamus alternatus carries Bursaphelenchus xylophilus Nickle and application of the primer group. The invention first discloses the LAMP primer group for detecting whether the monochamus alternatus carries the Bursaphelenchus xylophilus Nickle. With an SYG-2 gene of the Bursaphelenchus xylophilus Nickle as a target gene, the LAMP primer group with high specificity to the Bursaphelenchus xylophilus Nickle is designed and screened; and the LAMP primer group can specifically distinguish the Bursaphelenchus xylophilus Nickle from Bursaphelenchus mucronatus. The invention further discloses the LAMP detection kit for detecting whether the monochamus alternatus carries the Bursaphelenchus xylophilus Nickle. The LAMP detection kit comprises the LAMP primer group. The invention further establishes an LAMP detection method for detecting whether the monochamus alternatus carries the Bursaphelenchus xylophilus Nickle by using the LAMP primer group. By the LAMP detection method, quick and accurate detection on the Bursaphelenchus xylophilus Nickle carried by the monochamus alternatus can be achieved; the LAMP detection method is high in specificity, easy and convenient to operate and low in cost, and is of a great significance in monitoring occurrence and spread of a pine wilt disease.
Description
technical field [0001] The present invention relates to the detection of pine wood nematodes carried by Monochamus alternatus, in particular to a LAMP primer set and a kit for detecting pine wood nematodes carried by Monochamus alternatus, and the invention also relates to the use of the LAMP primer set and kit in detecting Monochamus alternatus The invention relates to an application in cattle carrying pine wood nematode, belonging to the field of molecular detection of Monochamus alternatus carrying pine wood nematode. Background technique [0002] Pine wilt disease, also known as pine wilt disease and fusarium wilt, is caused by the pine wood nematode [Bursaphelenchus xylophilus (Steiner&Buhrer) Nickle], mainly through insects of the genus Monochamus on pine trees. Natural transmission by feeding and spawning, and long-distance transmission by transportation of diseased wood and diseased wood processed wood. The spread of pine xylophilus in the forest area is mainly by M...
Claims
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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6844C12Q1/6888C12Q2531/119C12Q2537/1376C12Q2563/107
Inventor 王曦茁汪来发王艳娜姚玲朴春根李永薛寒
Owner INST OF FOREST ECOLOGY ENVIRONMENT & PROTECTION CHINESE ACAD OF FORESTRY
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