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Vol 48(2014) N 6 p. 795-801; DOI 10.1134/S0026893314060120 Full Text

X. Meng1,2, L.M. Li1,2 , G. Gao1 , F.L. Jin1, S.X. Ren1*

The gene expression of the protein SLAWD, mediating the toxic effect of destruxin A on Spodoptera litura larvae, in procaryotic cells: purification and characterization

1Engineering Research Center of Biological Control, Ministry of Education, South China Agricultural University (SCAU), Guangzhou, China
2Guangdong Entomological Institute, Guangzhou, China

*shxren@scau.edu.cn
Received - 2014-02-17; Accepted - 2014-03-12

Spodoptera litura is one of the most destructive phytophagous pest infesting cotton, vegetable, oilseed and fiber crops around the world. Destruxin A (DA), is a one of a kind microbial insecticide, which has potent toxins with bioactivity against S. litura larvae. An abnormal wing disc (AWD) protein was identified as a DA toxic effect protein in S. litura SL-1 cells. To better understand the role of the AWD gene of S. litura (SLAWD) it was purified and characterized. The entire coding region of the SLAWD gene was cloned into a pET-32a (+) expression vector and transformed into competent Escherichia coli BL21 (DE3) cells. SDS-PAGE and western blotting analysis showed that the best induction conditions were 1 mmol mL-1 isopropyl-β-D-thiogalactopyranoside (IPTG) for 6 h at 37°C; the molecular weight of the fusion protein was 35.0 kDa. The production of polyclonal antibodies and an enzyme-linked immunosorbent assay (ELISA) showed that the titer of antiserum was 1 : 25 600; western blotting analysis showed that the recombinant SLAWD was recognized by the anti-SLAWD polyclonal antibody. AWD is a key protein involved in wing development in insects. These tools will assist in the further characterization of SLAWD and studies on the mechanism of action of destruxin A.

destruxin, SLAWD, Spodoptera litura abnormal wing disc protein, prokaryotic expression, purification, polyclonal antibody



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