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CRISPR-Cas Detection Coupled with Isothermal Amplification of Bursaphelenchus xylophilus-武 瑾(1)
The pine wood nematode (PWN), Bursaphelenchus xylophilus, causessignificant damage to pine trees and, thus, poses a serious threat to pineforests worldwide, particularly in China, Korea, and Japan. A fast, af-fordable, and ultrasensitive detection of B. xylophilus is urgently neededfor disease diagnosis. Recently, clustered regularly interspaced shortpalindromic repeats (CRISPR)-based diagnostics have reshaped molec-ular diagnosis, with high speed, precision, specificity, strength, effi.ciency, and versatility. Herein, we established two isothermal diagnosticsmethods based on CRISPR-based platforms(CRISPR/Cas12a andCRISPR/Cas13a) for B, xylophilus-specific detection via fluorescence orlateral-flow strip readout. The guide RNA and CRISPR RNA weredesigned to target the 5S ribosomal DNA intergenic spacer sequencesregion of B, xylophilus. Recombinase-aided amplification was used forpreamplification whose reaction condition was 37°C for 15 min.
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