Pine wood nematode pcr detection kit and its detection method

Abstract

The invention discloses a pine wood nematode PCR (polymerase chain reaction) detection kit. The detection kit comprises a PCR reaction solution containing a primer A and a fluorescent probe A, wherein the primer A is divided into an upstream primer A and a downstream primer A; the nucleotide sequence of the upstream primer A is 5'-CCGTTTGGTGATGTTGTTTCA-3'; the nucleotide sequence of the downstream primer A is 5'-GCAAGGCTTCACGACGAA-3'; and the nucleotide sequence of the fluorescent probe A is 5'-FAM-CTCAACAAACCGCAGCCAGGACTC-BHQ-3'. The invention further discloses a pine wood nematode PCR detection method. The detection kit disclosed by the invention has the advantages of good DNA (deoxyribonucleic acid) extraction effect, simple operation, short consumed time, objective and reliable results, high sensitivity, high specificity and the like.

Description

technical field [0001] The invention relates to the technical field of microbial detection, in particular to a PCR detection kit for pine xylophilus and a detection method thereof. Background technique [0002] Pine wood nematode (North American pinewood nematode; Bursaphelenchus xylophilus), the nematode belongs to the phylum Vermiformes, class Nematodes, orders Osmocylinda, Superfamily Slipperidae, and the genus Umbris. The body of nematode adults is about 1 mm long, the tail of the female is nearly conical, with a rounded end; the tail of the male is like a bird's claw, curved towards the ventral surface. Pine wilt, also known as pine blight, is a devastating pest. It is spread in pine trees by vector insects such as Monochamus alternatus, thus causing pine tree diseases. After being infected by pine wood nematode, the needle leaves are yellowish brown or reddish brown, wilting and drooping, the resin secretion stops, and the long beetle intrusion holes or traces of spa...

AI Technical Summary

Problems solved by technology

The first category is common PCR detection methods, such as the "pine wood nematode detection kit and its detection method" with the patent application number 200310106109.5. In actual production; the second type is real-time fluorescent PCR detection method, such as "a method for detecting pine wood nematode and its special primers and probes" with the patent application number 200310109374.9, etc., these detection methods more or less exist to detect The specificity is not strong, and the detection accuracy needs to be improved, so it has not been applied in production practice; the third category is the constant temperature amplification detection technology, such as the patent No. 201210303532.3 "A kit for qualitative detection of pine wood nematode and Its detection method”, this detection technology has yet to be verified in production

[0004] In summary, the above-mentioned molecular detection methods of pine wood nematodes are difficult to meet the rapid quarantine requirements of various forest plant quarantine inspection stations

From the perspective of rapid quarantine, the above-mentioned pine wood nematode detection technologies and methods still have a certain gap from rapid quarantine. In order to meet the rapid identification requirements of quarantine practice, it is necessary to develop more simple, rapid and efficient detection and identification technologies and methods.

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