Cloning, characterisation and expression profiling of the cDNA encoding the ryanodine receptor in diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae)

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Abstract

BACKGROUND: The rynodine receptors (RyRs) are the main targets of diamide insecticides such as chlorantraniliprole. To provide the basis for a good understanding of the molecular mechanisms of diamide insecticide resistance, an RyR gene from Plutella xylostella was cloned and characterised in the present paper.

RESULTS: A full-length cDNA sequence of RyR was cloned from P. xylostella through RT-PCR and rapid amplification of cDNA ends (RACE). The gene (named PxRyR1) is 15 753 bp long, with an open reading frame of 15 354 bp, encoding a predicted RyR of 5117 amino acids. An alternative splicing of the PxRyR1 was also cloned and named PxRyR2. The PxRyR1 shares 77–93% identity with other insect RyRs. Quantitative real-time PCR analysis showed that the PxRyR was expressed at a high level in second-instar larvae and adults, at a low level in prepupae and pupae and abundantly in the body wall muscle and head (respectively 6.00 and 3.12 times the expression in the gut). Western blot analysis with anti-RyR antibodies showed that the RyR was mainly present in the body wall muscle and head, but barely present in the haemocyte and gut.

CONCLUSIONS: There are at least two alternative splices of PxRyR expressed in all developmental stages and tissues in P. xylostella at various levels. The results provided the basis for further understanding of the mechanisms of resistance to diamide insecticides in P. xylostella. Copyright © 2012 Society of Chemical Industry

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