Efficient Microspore Embryogenesis for the Development of Doubled Haploid Plants in Broccoli

The most acceptable strategic tool for creating homozygous inbred lines is the development of doubled haploids (DH's) by microspore culture technique. The current research focuses on development of DH's by microspore culture through the optimization of important factor limiting its efficiency. The effect of carbon sources, temperature shock for embryogenic response and effect of different plant growth regulators (PGRs) on effective regeneration was studied in four commercial varieties/hybrids of broccoli viz., Palam Vichitra, Lucky, Palam Samridhi and Palam Kanchan. Interaction between variety and carbon source (V × C) revealed maximum numbers of embryos per plate (330.00) in genotype V3 (Palam Samridhi) when supplemented with 13% sucrose (C2). While interaction between varieties and temperature (V × T) revealed V3 × T3 with significantly highest embryo yield (93.11). Interaction between temperature and carbon sources (T × C) revealed the maximum number of embryos per plate with treatment combination T3 × C2 (382.67). Overall, the highest embryogenic response (715.00) was obtained in Palam Samridhi (V3) genotype when supplemented with 13% sucrose (C2) at temperature treatment T3 (4°C-48 hr + 32°C-24 hr + 25°C-Cont). Further, maximum regeneration of embryos (75.00%) was recorded in genotype V3 when supplemented with 1.0 mg/l BAP (H5). Ploidy analysis of 952 microspore-derived plants from four broccoli genotypes revealed 52.31% plants were haploids, 43.17% plants were doubled haploids, 1.05% of plants were triploids, 2.84% were tetraploids and 1.20% were mixoploid. The present findings will accelerate doubled haploid based hybrid breeding in broccoli.