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In vitro double haploid production through anther culture in niger (Guizotia abyssinica L.F. Cass)

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Abstract  

To keep pace with the skyrocketing demand of vegetable oil and reduced dependency on import, there is a need to focus on indigenous oilseed crops, such as niger (Guizotia abyssinica L.F. Cass). Niger is a minor oilseed crop that consists of 32 to 40% quality oil and 18 to 24% protein in the seeds. Despite its considerable importance, self-incompatibility and high outcrossing pose difficulty in breeding through conventional methods. Double haploid (DH) technology is one of the alternatives to overcome the problem and develop inbred lines in a shorter time. In context to this, a highly efficient protocol for DH production through anther culture in Indian niger varieties JNS 9 and JNS 28 was developed. The investigation was conducted to identify the suitable microspore stage for anther culture, cold pre-treatment, hormonal combinations for callus induction (T1–T4), shooting (S1–S26), rooting (R1–R3), hardening, ploidy index, and the effect of different colchicine treatments on the efficiency of the developed protocol. Anthers in disc florets of light green to greenish yellow color have uninucleate microspores (7 d of bud initiation). For callus initiation, Murashige and Skoog (MS) medium containing 2.0 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.3 mg L−1 kinetin (KIN) showed 100% efficiency. For shoot initiation, 37 to 40% greening was observed on MS medium with 0.5 mg L−1 6-benzylaminopurine (BAP) and 0.5 mg L−1 activated charcoal and, subsequently, 19% shoot emergence on MS medium containing 0.5 mg L−1 BAP and 0.5 mg L−1 KIN. Low efficiency of shoot regeneration was addressed by adding 0.2% colchicine for 30 h at the callusing stage which resulted in 90 to 95% increased efficiency in shooting. For rooting, MS medium containing 2.0 mg L−1 indole-3-butyric acid (IBA) was efficient with a frequency of 85 to 90% followed by hardening in 1:1:1 ratio of vermiculite:sand:soil in pots. Ploidy indexing was done using cell size analysis followed by chromosome count using a microscope which confirmed the ploidy level of regenerants as compared to diploid control. Thus, the procedure developed in the study demonstrates the successful development of DH which in turn facilitates the acceleration of the breeding program through the development of homozygous inbreds and new cultivars.

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Acknowledgements

The authors acknowledge the funding support of Department of Biotechnology, Ministry of Science and Technology, Government of India, New Delhi, vides no. BT/Ag/Network/Niger/ 2019-20 dated 29-02-2020 for the Network Project entitled “Exploitation of genetic and genomic resources for improvement of niger (Guizotia abyssinica L.F. Cass) through breeding and biotechnological tools”. The authors are grateful to Dr. S.B. Verulkar (Head of the Department, Department of Plant Molecular Biology and Biotechnology, Indira Gandhi Krishi Vishwavidyalaya, Raipur, Chhattisgarh) for his constant guidance and support.

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  1. Department of Plant Molecular Biology and Biotechnology, Indira Gandhi Krishi Vishwavidyalaya, Raipur, C.G, 492 012, India

    Shrinkhla Maurya, Bhumika Sharma, Kajal Thakur, Suman Rawte, Nirmala Bharti Patel & Zenu Jha

  2. Department of Genetics and Plant Breeding, Jawaharlal Nehru Krishi VishwavidyalayaM.P. 482 004, Jabalpur, India

    Rajani Bisen

  3. Division of Genomic Resources, National Bureau of Plant Genetic Resources, Indian Council of Agricultural Research, New Delhi, 110012, India

    S. Rajkumar

  4. Indian Institute of Oilseeds Research, Rajendranagar, Hyderabad, 500 030, India

    M. Sujatha

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  1. Shrinkhla Maurya
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Conception and research design by SU and ZJ; research performed by SM, BS, SR, KT, NBP, and RS; data compilation by SM, BS, SR, KT, NBP, and RS; data analysis by SM and ZJ; material provided for experiment by RB and SU; paper writing and editing by SM, ZJ, and BS; and final editing, improvement, and suggestions by SU.

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Maurya, S., Sharma, B., Thakur, K. et al. In vitro double haploid production through anther culture in niger (Guizotia abyssinica L.F. Cass). In Vitro Cell.Dev.Biol.-Plant 60, 50–66 (2024). https://doi.org/10.1007/s11627-023-10391-z

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