Transgenic cassava lines carrying heterologous alternative oxidase (AtAOX1a) showed impaired quantitative and qualitative response to embryogenesis

Abstract

In our approach to control reactive oxygen species produced as a result of oxidative stress experienced by cassava roots during harvesting, a phenomena which causes postharvest physiological deterioration in the roots, we transformed cassava variety TMS 60444 with the AtAOX1a gene, driven by the 35S promoter, using agrobacterium-mediated approach. Extracted genomic DNAs of putative transgenic lines were screened using polymerase chain reaction technique (PCR). Messenger RNA was extracted from selected PCR-positive lines for reverse transcription-PCR analysis for gene expression. To screen positive lines for gene function, leaf lobes from two transgenic lines with a line carrying an empty vector and the wild type were subjected to somatic embryogenesis (SE), a known oxidative stress process. The results show that the wild type, at 16 days after initiation (DAI) of the leaf lobes on callus initiation medium, had the highest (100%) number of leaf lobes that produced at least one observed organised embryogenic structure (OES). This was followed by PEV-3, the empty vector plant with 50% OES production, while PB-3 had the least percent (20%) of leaf lobes with OES. PB-3 line also had no OES at all in five out of the seven periods of data collection. During the period, the wild type recorded the highest attainable OES quality score of 2.0 (on a scale of 1-5 where 1=bad and 5=excellent) at the first initiation cycle. Both the transgenic lines and the empty-vector plantlet recorded quality score of 1.0. It seems AtAOX1a only hinders OES development, but exerts little effect on the quality, if OES does not degenerate after development.

Keywords: Genomic DNAs, reverse transcription-PCR, somatic embryogenesis (SE)

African Journal of Biotechnology Vol. 12(27), pp. 4303-4309