Radiation induced alterations in Vigna radiata during in vitro somatic embryogenesis

Abstract

Purpose: Tissue culture has been exploited to understand molecular aspects of regeneration potential of the plants in normal and in stressed conditions. The present study describes ionizing radiation from ⁶⁰Co source as the stress stimulator to assess in vitro development of somatic embryo of Vigna radiata, a protein-rich pulse.

Materials and methods: Callus culture was established, using leaves of V. radiata. Somatic embryogenesis was induced by manipulating plant hormones. Calli were exposed to gamma rays. Genomic DNA isolated from gamma-irradiated callus samples were subjected to random amplified polymorphic DNA analysis. A band of molecular weight 1440 bp was used as a probe and Southern hybridization was carried out. To determine alterations in DNA following irradiation, RAPD bands were cloned and sequenced from control and irradiated samples. Embryogenic calli were exposed to gamma irradiation and the effects were assessed immediately and after seven days of exposure. Phenotypic alterations were observed using scanning electron microscopy.

Results: Exposed calli revealed altered frequency of somatic embryo formation. Results showed that the 1440 bp molecular weight probe hybridized with bands of low molecular weight. DNA sequences from irradiated samples showed recombination when compared to control. Scanning electron micrography illustrated presence of transient pores on the exposed embryos. BLAST search of the DNA sequences showed partial homology with some sequences from Arabidopsis thaliana.

Conclusion: The present report might help in designing a breeding program, where both radiation coupled with somatic embryogenesis could be employed to build up the desired variants.

Keywords:

• Gamma rays
• somatic embryo
• scanning electron microscopy
• random amplified polymorphic DNA
• southern hybridization
• cloning
• sequencing