PJB-2025-153
AN EFFICIENT METHOD OF ROOTING FROM IN VITRO CULTURE OF ZYGOTIC EMBRYOS OF PINUS ROXBURGHII SARG.
AROOSHA ASHRAF
Abstract
This study was conducted to explore the possible rooting efficacy of the mature zygotic embryo of Pinus roxburghii. Megagametophytes with zygotic embryos were aseptically removed from surface-sterilized mature cones. After aseptic extraction, mature zygotic embryos (MZE) and megagametophytes (Mgt) were individually cultured on modified LP 505 media + 0.45 mg/l BAP + 2 mg/l NAA + 0.43 mg/l Kinetin for initiation and extrusion of cultures in the dark for 20 days before switching to 16 h photoperiod for 20 more days. In light (40-day-old cultures), embryos extruded 75% and Mgt 19.04%. The shoot length was 2cm after 30 days of cultivation, while Mgt produced 3cm shoots. Developing shoots (40 days old) were moved to DCR or LP semi-solid media enriched with 10, 20, 30, or 40 µM of indole-3-butyric acid (IBA) + α-naphthalene acetic acid (NAA) in 16 h photoperiod. Rooting was unsuccessful on these media; however, shoots developed up to 4 cm on LP + 30 µM IBA + NAA after 20 days (2-month-old cultures). Both MS and DCR cultures had 50% rooting with 2cm long roots in half DCR, LP, or whole MS media. After a month, rooted shoots were transferred to 8 × 15 cm plastic pots with 3:1 peat moss and sand potting media. Pots were wrapped in plastic bags to constrain moisture and kept under the culture room conditions. For one month, irrigation with ¼ DCR was done every 10 days. Plants from MS lasted 35 days, whereas DCR plants stayed green in the potting mix for 15 days and successfully grew under field conditions.
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