Species-specific COI(SS-COI)primers for rapid identification of Leptocybe invasa cryptic species A & B
Cryptic species A and B are two important globally intrusive cryptic species of Leptocybe invasa,which have caused serious damage to forestry production in many countries.Due to the tiny size and undistinguishable cryptic species in morphology,it makes pest control more difficult.In this study,we researched the rapid molecular identification method of cryptic species of L.invasa,basing on the species-specific COI(SS-COI)PCR method of mitochondrial DNA cytochrome C oxidase subunit I(mtDNA COI)gene sequence.According to the COI sequence,two pairs of cryptic species A and B SS-COI primers(AF/AR and BF/BR)were respectively designed.Using these primers to amplify the DNA of the same L.invasa sample,by which could not only effectively identify the cryptic species,but also mutually verify the identification accuracy.The primer identification sensitivity detection showed that both AF/AR and BF/BR had high identification sensitivity,and their effective identification concentration thresholds for DNA were 11.42 pg/μL and 28.32 pg/μL.The rapid identification method developed for the cryptic species A and B of L.invasa in this study can addresse the challenge of discriminating cryptic species in areas invaded by L.invasa.It significantly shortens identification time,reduces identification costs,and provides technical references for further exploration of the biological differences between cryptic species A and B of L.invasa,as well as their adaptability to different resistant varieties of Eucalyptus.
Leptocybe invasacryptic species identificationSS-COI primersrapid identificationmolecular detection
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