Involvement of exogenous polyamines enhances regeneration and Agrobacterium-mediated genetic transformation in half-seeds of soybean
2016
Arun, Muthukrishnan | Chinnathambi, Arunachalam | Subramanyam, Kondeti | Karthik, Sivabalan | Sivanandhan, Ganeshan | Theboral, Jeevaraj | Alharbi, SulaimanAli | Kim, ChangKil | Ganapathi, Andy
The present work demonstrates the participation of polyamines (PAs) to improve direct regeneration and Agrobacterium-mediated transformation in soybean half-seeds. The inclusion of PAs to culture medium along with optimal plant growth regulators (PGRs) enhanced shoot induction [98.3 %; 4.44 µM N⁶-benzyladenine (BA) and 103.27 µM spermidine] and elongation [90.0 %; 1.45 µM gibberellic acid (GA₃) and 49.42 µM spermine]. The polyamine putrescine (62.08 µM) alone greatly enriched root induction (96.3 %). The influence of PAs on transformed plant production was assessed by comparing optimized protocol (comprising PAs and PGRs) with a regeneration system involving only PGRs. Plant transformation was performed in half-seeds of cultivar DS 97–12 using strain EHA105 harboring pCAMBIA1301. Transgene expression and integration was confirmed by GUS staining, PCR, and Southern hybridization. The transformed explants/materials successively cultured on co-cultivation (BA and spermidine), shoot induction (BA and spermidine), shoot elongation (GA₃ and spermine), and rooting medium (putrescine) showed enhanced transformation efficiency (29.3 %) compared with its counterparts (14.6 %) with respective PGR alone [BA, GA₃, or indole-3-butyric acid (IBA)]. Overall findings of the study suggest that involvement of PAs improved T-DNA transfer during co-cultivation, and delivered most suitable condition for efficient regeneration/survival, which led to enhanced transformation efficiency in soybean.
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