Analysis of colchicine effect on anther culture of sunflower (helianthus annuus L.)
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Date
2019-05-30
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Vasantrao Naik Marathwada Krishi Vidyapeeth, Parbhani
Abstract
Sunflower (Helianthus annuus L.), is a multi-purpose crop employed for diverse uses such as dye production, edible oil extraction and for medicinal applications. Production of microspore-derived embryos from cultured anthers is now well established technique for isolation of homozygous lines in sunflower. In this present study different concentrations of colchicine 0.1, 0.5, 1, 1.5 and 2% were used to develop haploid and increase ploidy level of sunflower by direct treatment to anther and treatment to callus in medium.
In direct colchicine treated anthers response in survival percentage were 25, 22.5, 17.5, 12.5and 10%, respectively; whereas, the response of untreated control was 95%. The callus induction frequency was 23.33, 20, 16.66, 11.66 and 8.33, respectively and 88.33 for untreated control. In indirect colchicine treatment, survival percentages of treated callus were 82.5, 72.5, 62.5and47.5% respectively and 95% of untreated control. The callus regeneration percentage of control and treated callus was 96.66, 86.66, 76.66, 66.66 and 55%, respectively.
Ploidy level of regenerated callus and its donor plant were confirmed by SSR marker and flow cytometry. Out of 28 SSR markers, 15 markers viz., ORS-05, ORS-662, ORS-1265, ORS-552, ORS-488, ORS-243, ORS-1159, ORS-928, ORS-58, ORS-337, ORS-959, ORS-154, ORS-780, ORS-423 and ORS-536 were found useful for the identification of haploids in sunflower. Both the regenerated haploid lines had homozygous profiles with a single band, while the donor plant had two bands that indicated allelic sites. The fifteen SSR markers showed 100% homozygosity.
According to SSR markers and Flow cytometry results, calli derived from direct and indirect colchicine treatments are double haploids. The double haploids are also obtained in the control callus in both the cases, indicates spontaneous dihaploidization of the chromosomes.
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