OPTIMIZED REGENERATION OF Kolanchoe (Kalanchoe blossfeldiana R.) AND Agrobacterium tumefaciense-MEDIATED Gus GENE TRANSFORMATION
MARYAM MEHDIZADEH HAKKAK *
Department of Biotechnology and Plant Breeding, Ferdowsi University of Mashhad, Iran.
NASRIN MOSHTAGHI
Department of Biotechnology and Plant Breeding, Faculty of Agricultural Biotechnology, Ferdowsi University of Mashhad, Iran
ABDOLREZA BAGHERI
Department of Biotechnology and Plant Breeding, Faculty of Agricultural Biotechnology, Ferdowsi University of Mashhad, Iran.
*Author to whom correspondence should be addressed.
Abstract
The effects of plant growth regulators were investigated on Kalanchoe blossfeldiana regeneration and then gene transferring was optimized. Three types of explants: leaf surface, vein and petiole and different concentrations of Benzyl Adenine (BA) and Kinetin (Kin) alone or in combination with Naphthalene Acetic Acid (NAA) were used for regeneration. Results showed that the highest number of shoots (47.33) and leaves (330.33) were obtained on Murashige-Skoog (MS) medium + 1.5 mg/L BA and 0.7 mg/L NAA. Maximum length of shoots (1.7 cm) was obtained from petiole explants on MS medium + 1.5 mg/L BA and 0.7 mg/L NAA. In medium containing Kin, adventitious root regeneration obtained from leaf explants. The most root number (21.12) and root length (1.56 cm) produced in MS+1 mg/L Indole Butyric Acid (IBA). For acclimation, the most length and leaf number increase percentage were obtained in peat-moss substrate. For optimization of β-glucuronidase-gene transferring, leaf sections and stems of in-vitro plantlets were used as explants for co-culturing in 10 and 30 min by Agrobacterium tumefaciense strain LBA4404 containing gus gene. The highest percentage of gene transferring and its expression (38.46%) was observed in leaf explants by 10 min co-culturing. So this protocol can be used for efficient gene transferring to this plant.
Keywords: Gene expression, gene transferring, Kalanchoe, regeneration, tissue culture.
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