Efficient induction of parthenogenetic haploids in African marigold (Tagetes erecta) L.) using Co60 gamma-irradiated pollen of French Marigold (Tagetes patula L.)

Haploids and doubled haploids play a crucial role in crop improvement programs, including hybridisation, mutation breeding, and genetic and genomic studies. Induction of parthenogenesis using gamma-irradiated pollens is an efficient approach for inducing haploids and developing homozygous lines in horticultural crops. However, no research has been conducted using the induced parthenogenesis approach for haploid induction in the genus Tagetes. This study aims to assess the effectiveness of gamma irradiation at doses of 100, 200, 300, 400, and 500 Gy, using a Co60 (Cobalt-60) source, for haploid induction. The irradiated pollen of a distant tetraploid species, Tagetes patula genotype ‘Pusa Deep’, was used to induce haploids in the diploid marigold species, T. erecta genotypes ‘DAMH-24’ and ‘DAMH-55’. The pseudo-fertilized ovaries were cultured on modified MS medium supplemented with 0.75 mgl-1 TDZ, 0.2 mgl-1 NAA, and 0.5 mgl-1 GA3. To better understand the effect of γ irradiation doses on haploid induction rate, a correlation analysis was performed. Out of 50 regenerants, 20 were selected to determine ploidy levels using morphological traits, chloroplast count, chromosomal count, and flow cytometry analysis. A plant with 3-7 chloroplasts in stomatal guard cells, 2n=x=12 chromosomes, and 1C DNA (~54K mean Pi) was identified as haploid; the plant with 10-14 chloroplasts, 2n=2x=24 chromosomes, and 2C DNA (~108K mean Pi) was diploid, while the plant with 16-20 chloroplasts, 2n=2x=24 chromosomes, and 3C DNA (~154K mean Pi) was triploid. For efficient pollen inactivation and high-throughput haploid induction in Tagetes erecta, the 400 Gy treatment was found to be the most effective.