Parameters influencing Agrobacterium-mediated transformation system in safflower genotypes AKS-207 and PKV Pink
2016
Dhumale, DiptiRaghunath | Shingote, PrashantRaghunath | Dudhare, MahendraShankarrao | Jadhav, PravinVishwanath | Kale, PrashantBhaskar
Shoot regeneration in safflower (Carthamus tinctorius ‘AKS 207’ and ‘PKV Pink’) genetically transformed using Agrobacterium was used for assessing various constraints to the efficiency of transformation including infection period, virulence induction medium, co-cultivation period, bacterial titre, selection regime, and the natural phenolic compound acetosyringone. Transformation frequency was promising with 8–10-day-old cotyledonary leaf explants. Therefore, explants of that age cultured on Agrobacterium minimal medium (AB) containing 100 µM acetosyringone were infected with Agrobacterium (cell titre 0.5 OD₆₀₀ₙₘ) for 15 min followed by 48 h of co-cultivation on kanamycin-enriched medium (50 mg/L). Transformation of the shoots was confirmed using β-glucuronidase (GUS) histochemical assay and polymerase chain reaction (PCR). With the transformation protocol thus optimized, the transformation frequency as determined using GUS assays was 54.0 % for AKS 207 and 47.6 % for PKV Pink. The corresponding figures using PCR were 27.0 and 33.3 %. The transformed shoots required 10–14 weeks of culture initiation but produced very few roots.
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