Molecular marker assisted confirmation of a hybrid between Ascocentrum ampullaceum var. auranticum and Rhynchostylis retusa.

Two rare and endangered orchids, Rhynchostylis retusa, and Ascocentrum ampullaceum var. auranticum, a narrowly endemic orchid from Manipur, were used for the present study. These two orchids were selected as parents because they have desirable traits like dense and floriferous, colorful, and long-lasting flowers. Our goal is to develop a hybrid species that is intermediate and improved over its parents in terms of floral characteristics by the F1 generation. When A. ampullaceum var. auranticum was the female parent, a 90 % crossability success rate was attained. Half-strength Murashige and Skoog (MS) basal medium with no phytohormones was used for seed germination in vitro and protocorm development of the putative hybrid. The basal medium containing 1 mg/L BAP and 0.5 mg/L NAA showed the highest growth response with 7.57 shoots/explant, 5.90 leaves/explant, and 57.19 PLBs/explant, while the medium enriched with 1 mg/L NAA produced the maximum number of roots (4.95) after 150 days of inoculation. The assessment for rapid determination of genetic purity of the developed hybrid (F1 plants) between A. ampullaceum var. auranticum and R. retusa was carried out by using the molecular markers, SCoT and SSR. The F1 plants' complementing banding patterns, which they inherited from their parents, confirmed that they were pure hybrids. These PCR-based molecular markers could be used for the early assessment of hybridity.