Invasive Species Compendium

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Abstract Full Text

Cloning and functional characterization of the NanosO gene promoter in Plutella xylostella (Lepidoptera: Noctuidae).

Abstract

Aim: To clone the NanosO gene (PxnosO) promoter from the diamondback moth, Plutella xylostella, and to verify its gonad-specific activity, so as to further study the gene functions and develop transgenic insects for integrated management of P. xylostella and other agricultural pests. Methods: According to the sequence information of NanosO gene in the P. xylostella genome, the NanosO gene promoter was cloned by PCR and subjected to sequence analysis. The PxnosO-EGFP expression plasmid was constructed, and then the PxnosO-EGFP and IE1-EGFP expression plasmids were transfected into P. xylostella embryonic cell line (Px-6) and Spodoptera frugiperda ovary cell line (Sf9) by lipofection. The activity of the NanosO promoter in P. xylostella was confirmed by the qualitative and quantitative analysis of EGFP expression by laser confocal fluorescence observation and qRT-PCR, respectively. Results: We cloned the PxnosO (Px004767) promoter (with the length of 1 743 bp) of P. xylostella. Sequence analysis showed that this promoter not only contains common core promoter elements like TATA box, and upstream promoter domains TATA box and GC box, but also contains dozens of transcription factor binding sites. Using cell transfection technology, we successfully expressed exogenous gene EGFP driven by this promoter in the Px-6 and Sf9 cell lines. Conclusion: We cloned the PxnosO promoter of P. xylostella and verified that it can drive the expression of EGFP gene at the cellular level. This study provides a foundation for analyzing the expression pattern of PxnosO in different developmental stages of P. xylostella and the potential functions of the PxnosO promoter in vivo.