›› 2014, Vol. 57 ›› Issue (2): 150-160.

• RESEARCH PAPERS • Previous Articles     Next Articles

Molecular identification of laccase 2 gene in the cotton bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae)

CAO Zhi-Shan, LIU Xiao-Ming, AN Shi-Heng, JIANG Jin-Wei*   

  1. (College of Plant Protection, Henan Agricultural University, Zhengzhou 450002, China)
  • Online:2014-02-20 Published:2014-02-20

Abstract: 【Aim】 The melanization plays an important role in process of insect cuticle hardening and the immune defense, in which phenol oxidizing enzymes are the key enzymes. Laccase 2 (LAC2), as one of phenol oxidizing enzymes, plays an important role in metamorphosis and immune system of insects. The aim of this study was to explore its expression pattern and hormonal regulation during the cuticle hardening in the cotton bollworm, Helicoverpa armigera. 【Methods】 The fulllength cDNA sequence of lac2 was amplified from the 5th instar larvae of the cotton bollworm by PCR and RACE methods. The expression patterns and hormonal regulation of lac2 were analyzed by using real-time quantitative PCR, hormone treatment and RNA interference methods. 【Results】 The sequence analysis showed that the full-length cDNA sequence of lac2 is 3 221 bp in length with an open reading frame of 2 268 bp encoding 756 amino acid residues. The developmental expression pattern showed that lac2 transcription was richly expressed in the molting stage of every instar and reached a peak in 96 h-5th instar larvae. The spatial expression manifested that lac2 was richly expressed in the cuticle of larvae and in the ovary and antennae of adults. Further hormone treatment showed that juvenile hormone analogue (methoprene) inhibited lac2 transcription, while molting hormone (20-hydroxyecdysone) attenuated methoprene action and induced the expression of lac2. RNA interference of the molting hormone receptors EcR (ecdysone receptor) and USP (ultraspiracle isoform) significantly inhibited the expression of ecr and usp, and injection of dsRNA of ecr and usp followed by injection of 20-hydroxyecdysone also inhibited the transcription of lac2. 【Conclusion】 Altogether, these results provide a foundation for further research of lac2 in insect cuticle hardening and immune defense.

Key words:  Helicoverpa armigera, melanization, laccase, immune defense, RNA interference, juvenile hormone analogue, molting hormone