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Development of a highly accurate and sensitive diagnostic tool for pyrethroid‐resistant chimeric P450 CYP337B3 of Helicoverpa armigera using loop‐mediated isothermal amplification

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Heckel,  David G.
Department of Entomology, Prof. D. G. Heckel, MPI for Chemical Ecology, Max Planck Society;

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Kim,  Juil
Department of Entomology, Prof. D. G. Heckel, MPI for Chemical Ecology, Max Planck Society;

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Citation

Choi, B. H., Hur, J. H., Heckel, D. G., Kim, J., & Koh, Y. H. (2018). Development of a highly accurate and sensitive diagnostic tool for pyrethroid‐resistant chimeric P450 CYP337B3 of Helicoverpa armigera using loop‐mediated isothermal amplification. Archives of Insect Biochemistry and Physiology, 99(3): e21504. doi:10.1002/arch.21504.


Cite as: https://hdl.handle.net/21.11116/0000-0003-B33D-F
Abstract
Recent studies have shown that pyrethroid resistance in the
cotton bollworm (CBW) Helicoverpa armigera is conferred by
the generation of a chimeric CYP337B3 gene, which resulted
from unequal crossing‐over between the CYP337B1 and
CYP337B2 genes. In this study, we developed a diagnostic
protocol based on the loop‐mediated isothermal amplification
(LAMP) assay for the detection of chimeric CYP337B3. The
CYP337B3 LAMP assay utilized six primers and generated
strong fluorescence signals visible to the naked eye under
normal or ultraviolet light. The primers were designed based
on CYP337B3v1 (JQ995292), the major allele detected in
Australia. The detection limit of this LAMP assay was 10 fg
genomic DNA in a 25‐μl reaction mixture. Compared with
CYP337B2v1, the Korean CYP337B3v2 allele had two
nucleotide mismatches within the amplifying regions of this
LAMP assay; therefore, we confirmed that polymerase chain
reaction‐synthesized CYP337B3v2 was well amplified using
this LAMP assay. In addition, we determined that the presence
of CYP337B3 from H. armigera collected by pheromone traps
from Korean fields could be confirmed using this LAMP assay.
This assay could detect CYP337B3 even in heterozygotes,
which is relevant because CYP337B3 is dominant, and
heterozygotes are pyrethroid resistant. Therefore, the newly developed CYP337B3 LAMP assay could detect the presence
of pyrethroid resistance in H. armigera that were captured by
pheromone traps during the early season and provide information on whether pyrethroids could be used to control H. armigera.