Factors affecting genetic transformation by particle bombardment of the prickly pear cactus (O. ficus-indica)

摘要

In the present study, a novel transformation protocol for Opuntia ficus-indica was generated by means of particle bombardment. The best conditions obtained were: 900 psi rupture disk pressure, 8 cm microprojectile travel distance, and 4 h of exposition to 0.2 M mannitol. For all experiments, gold particles coated with 1.0 µg/µL of pBI426 plasmid DNA were used. With all these conditions, a 23% of transformation efficiency in terms of regeneration in selection media (100 mg/L kanamycin) was obtained. Interestingly, the presence of both transgenes: nptII and uidA, by means of PCR and RT-PCR assays was detected. The regeneration percentage achieved in terms of stable integration for both genes was 10%. In addition, we also detected adequate amounts of β-glucuronidase activity by means of the GUS fluorometric assay. The procedure described in the present investigation reveals the feasibility of using nopal for the introduction, expression, and possible production of heterologous proteins.