Activated charcoal induced high frequency microspore embryogenesis and efficient doubled haploid production in Brassica juncea |
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Authors: | Deepak Prem Kadambari Gupta Gautam Sarkar Abha Agnihotri |
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Affiliation: | (1) Center for Bioresources and Biotechnology, TERI University, Darbari Seth Block, Habitat Place, Lodhi Road, New Delhi, 110 003, India;(2) Present address: Monsanto India Ltd., Ahura Centre, 5th Floor, 96, Mahakali Caves Road, Andheri (East), Mumbai, 400093, India;(3) Present address: Financial Technologies (India) Limited, 406, New Delhi House, 27, Barakhamba Road, Connaught Place, New Delhi, 110 001, India;(4) Environmental and Industrial Biotechnology Division, TERI, Darbari Seth Block, Habitat Place, Lodhi Road, New Delhi, 110 003, India |
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Abstract: | Three Indian Brassica juncea cultivars were studied for embryogenic response of microspores, microspore embryo regeneration, ploidy assessment of microspore-derived plants and their diploidization. Genotype dependence for microspore totipotency was observed and a significant effect of genotype by bud size selection was established. The addition of activated charcoal in NLN medium containing 13% (w/v) sucrose and 10 μM silver nitrate resulted in a fourfold increase in microspore embryogenesis, ranging from 100 to 405 embryos per Petri dish corresponding to 2,700–10,935 embryos per 100 buds. Conversion/germination of embryos produced in presence or absence of activated charcoal was similar but air-drying of microspore embryos was essential. Incubation of microspore embryos at 4 ± 1°C for 10 days in dark resulted in 82.3% conversion. The majority of plants produced from these embryos was haploid. Treating microspore-derived plants at the 3–4 leaf growth stage with 0.34% colchicine for 2–3 h resulted in greatest survival (70%) and chromosome doubling (75%) frequencies. Doubled haploid plants were self-pollinated and grown to maturity under field conditions. |
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Keywords: | Microspore embryo induction Androgenesis Embryo conversion Colchicine treatment DH production |
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