(242) Callus Induction and Regeneration in Hydrangea macrophylla (Thunb.) Ser.

Abstract

Development of a reliable method for Hydrangea macrophylla (Thunb.) Ser. organogenesis is critical for developing an in-vitro mutagenesis protocol. Container-grown (11.8 L) H. macrophylla `Nikko Blue' plants were maintained in a controlled environment greenhouse, with supplemental lighting (1600 hr to 2400 hr mercury vapor lamp), fertilized with 65 g Nutricote total (18N–2.6P–6.6K, Agrivert, Inc., New York, N.Y.) and hand-watered. To reduce fungal contamination, stock plants were sprayed to run-off biweekly with Alliette WDG (375 mg·L-1, aluminum tris), Bayleton (250 mg·L-1, triadimefon), and Heritage (25 mg·L-1, azoxystrobin). Leaf explants were sterilized with 0%, 10%, 15%, or 20% bleach (5.25% sodium hypochlorite) (by volume) for 10 or 15 min, and stem explants were sterilized with 0%, 10%, 25%, or 50% bleach (5.25% sodium hypochlorite) for 10 or 15 min. About 97% of fungal contaminates were eliminated from leaf and stem explants when treated with 10% bleach for either 10 or 15 min. Leaves were plated on Gamborg B5 media at pH 5.7 containing 0, 2.5, 5, or 10 μM 2,4-D and 0, 0.25, 0.5, or 1.0 μM BAP and placed in a controlled environment growth room under a 14-h photoperiod or in a dark growth chamber. Callogenesis followed by root organogenesis was observed on explants treated with a variety of concentrations and combinations of 2,4-D and BAP. Strongest callogenesis was observed on media supplemented with 10 μM 2,4-D. A greater callus concentration was observed along the edges of dark cultured leaf discs. Indirect root induction was greatest on 10 μM 2,4-D.