Engineering of insecticidal hybrid gene into potato chloroplast genome exhibits promising control of Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae)


Hossain M. J., Bakhsh A., Joyia F. A., AKSOY E., Gökçe N. Z. Ö., Khan M. S.

Transgenic Research, vol.32, no.6, pp.497-512, 2023 (SCI-Expanded) identifier identifier identifier

  • Publication Type: Article / Article
  • Volume: 32 Issue: 6
  • Publication Date: 2023
  • Doi Number: 10.1007/s11248-023-00366-6
  • Journal Name: Transgenic Research
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus, BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, Chemical Abstracts Core, Environment Index, MEDLINE, Veterinary Science Database
  • Page Numbers: pp.497-512
  • Keywords: Bacillus thuringiensis, CPB, Insect resistance, Transplastomic approach
  • Middle East Technical University Affiliated: Yes

Abstract

The potato chloroplast was transformed with codon optimized synthetic hybrid cry gene (SN19) to mitigate crop losses by Colorado potato beetle (CPB). The bombarded explants (leaves and internode) were cultured on MS medium supplemented with BAP (2.0 mg/l), NAA (0.2 mg/l), TDZ (2.0 mg/l) and GA3 (0.1 mg/l); spectinomycin 50 mg/l was used as a selection agent in the medium. Leaf explants of cultivar Kuroda induced highest percentage (92%) of callus where cultivar Santae produced the highest percentage (85.7%) of transplastomic shoots. Sante and Challenger showed 9.6% shoot regeneration efficiency followed by cultivar Simply Red (8.8%). PCR amplification yielded 16 postive transplastomic plantlets out of 21 spectinomycin resistant ones. Target gene integration was confirmed by PCR and Southern blot, whereas RT-qPCR was used to assess the expression level of transgene. The localization of visual marker gene gfp was tracked by laser scanning confocal microscopy which confirmed its expression in chloroplasts of leaf cells. The transplastomic plants ensured high mortality to both larvae and adult CPB. Foliage consumption and weight gain of CPB fed on transplastomic leaves were lower compared to the control plants. Sucessful implementation of current research findings can lead to a viable solution to CPB mediated potato losses globally.