Method for increasing the efficiency of in vitro experimental androgenesis induction in sugar beet

Abstract

Purpose and objectives. To develop a method for increasing the efficiency of in vitro experimental androgenesis induction in sugar beet.

Material and methods. The study was conducted in 2014–2017. The breeding material of Bila Tserkva Experimental Breeding Station, sugar beet tetraploid pollinators and diploid seed bearers (breeding genotypes BC 45 K1 - K15, 1475 K1-25) were taken as the test material. Biotechnological, breeding, physiological and statistical methods were used.

Results and discussion. The influence of temperature regime and composition of nutrient media used for pre-treatment of explants, stalks with buds, was investigated. Sugar beet tetraploid pollinators and diploid seed bearers grown in the field and under the laboratory conditions were used in the study. The explants were cultured in vitro at various temperature – from 4–10 to 20–25ºС on modified agarized nutrient media. The best results were obtained, when explants were cultured on Murashigе–Skооg medium containing ½ doses of macronutrients and full doses of micronutrients supplemented with Gamborg’s vitamin solution, ascorbic acid (1 mg/L), amino acids (glutamine –250 mg/L, asparagine – 30.0 mg/L, tyrosine – 5 mg/L, arginine – 3 mg/L, hydroxyproline – 2 mg/L), and growth regulators (2,4–D – 2.0 mg/L, 6–BAP – 0.6 mg/L and ABA – 0.3 mg/L) in a refrigerator at 4–10 ºС, with 16-hour illumination of 1.0–2.0 kilolux for 7–30 days. Such conditions increased the numbers of calluses from anthers from 1.8–2.1% to 4.9–5.1%, of embryoids from 0.9–1.2% to 6.1–6.3%.

Conclusions. The method for increasing the efficiency of in vitro experimental androgenesis induction in sugar beet, which includes low-temperature pre-treatment of explants - stalks with buds on Murashige-Skoog modified medium supplemented with vitamins, amino acids and growth regulators, in particular, ABA (0.3 mg/L), was developed. Application of the method allows over twice increase in the calluse number and 5–6 time increase in the number of embryoids obtained from sugar beet anthers in vitro, which will ensure the production of haploid regenerants for breeding programs to create homozygous lines.