Identification of hemizygous and homozygous transgenic rice plants in T₁ generation by DNA blot analysis

Abstract

Southern hybridization-based zygosity analysis was done in a transgenic rice plant (Oryza sativa L. cv Pusa Basmati 1) generated by Agrobacterium-mediated transformation with a rice chitinase (chi11) gene. A T₀ plant with two unlinked T-DNA insertions (A and A'), was chosen for the application of Southern hybridization analysis to study genetic separation of the two loci by segregation and to identify the homozygous and hemizygous plants in T₁-generation. The T₁ plants showed differences in band intensities that reflected the homozygous and hemizygous status of each of the two integration events. The predictions of zygosity of T₁ plants were confirmed by analyzing segregation in T₂ plants. Southern hybridization analysis is demonstrated as a simple and effective method to distinguish hemizygous and homozygous plants in the T₁ generation itself.