Robust genetic transformation of sorghum ( Sorghum bicolor L.) using differentiating embryogenic callus induced from immature embryos

• Published in: Plant methods 2017-12, Vol.13 (1), p.109-109, Article 109
• Main Authors: Belide, Srinivas, Vanhercke, Thomas, Petrie, James Robertson, Singh, Surinder Pal
• Format: Article
• Language: English
• Subjects: Analysis; Embryonic development; Flow cytometry; Genetic aspects; Genetic transformation; Geneticin; Lipoic acid; Methodology; Nodular structures; Particle bombardment; Physiological aspects; Ploidy analysis; Sorghum; Technology application
• ISSN: 1746-4811; 1746-4811
• DOI: 10.1186/s13007-017-0260-9

Sorghum ( L.) is one of the world's most important cereal crops grown for multiple applications and has been identified as a potential biofuel crop. Despite several decades of study, sorghum has been widely considered as a recalcitrant major crop for transformation due to accumulation of phenolic compounds, lack of model genotypes, low regeneration frequency and loss of regeneration potential through sub-cultures. Among different explants used for genetic transformation of sorghum, immature embryos are ideal over other explants. However, the continuous supply of quality immature embryos for transformation is labour intensive and expensive. In addition, transformation efficiencies are also influenced by environmental conditions (light and temperature). Despite these challenges, immature embryos remain the predominant choice because of their success rate and also due to non-availability of other dependable explants without compromising the transformation efficiency. We report here a robust genetic transformation method for sorghum (Tx430) using differentiating embryogenic calli (DEC) with nodular structures induced from immature embryos and maintained for more than a year without losing regeneration potential on modified MS media. The addition of lipoic acid (LA) to callus induction media along with optimized growth regulators increased callus induction frequency from 61.3 ± 3.2 to 79 ± 6.5% from immature embryos (1.5-2.0 mm in length) isolated 12-15 days after pollination. Similarly, the regeneration efficiency and the number of shoots from DEC tissue was enhanced by LA. The optimized regeneration system in combination with particle bombardment resulted in an average transformation efficiency (TE) of 27.2 or 46.6% based on the selection strategy, 25% to twofold higher TE than published reports in Tx430. Up to 100% putative transgenic shoots were positive for - by PCR and 48% of events had