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A Novel Approach of Transducing Recombinant Baculovirus into Primary Lymphoid Cells of Penaeus monodon for Developing Continuous Cell Line

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Abstract

Cell line development from shrimp is not a novel venture as researchers across the globe have been trying to have crustacean cell lines over 30 years. The reason for not attaining a crustacean or precisely a shrimp cell line is believed to be the replicative senescence and the inability to maintain telomere length in vitro. Moreover, spontaneous in vitro transformations do not happen in shrimp cells. Oncogenic induction in primary cell culture is one of the ways to attain in vitro transformation by way of disrupting the mechanisms which involve cellular senescence. In this context, a recombinant baculovirus with shrimp viral promoter IHHNV-P2 was used for the transduction aimed at immortalization. An oncogene, H-ras, was successfully amplified and cloned in to the baculoviral vector, downstream to shrimp viral promoter IHHNV-P2 and upstream to GFP. Recombinant baculovirus with H-ras was generated and used for transduction into shrimp lymphoid cells during early dividing stage. Accordingly, fibroblast-like primary cell culture got developed, and H-ras and GFP expression could be confirmed. The study suggests that the simple method of incubating recombinant baculovirus with minced tissue enables in vitro transduction during early dividing stage of the cells, and the transduction efficiency gets enhanced by adding 5 mM sodium butyrate to the culture medium.

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Funding

This research was supported by the Department of Biotechnology, Government of India (BT/PR5126/AAQ/3/591/2012). University Grants Commission, Government of India, offered BSR fellowship subsequent to the project period.

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Authors and Affiliations

  1. National Centre for Aquatic Animal Health, Cochin University of Science and Technology, Fine Arts Avenue, Kochi, Kerala, 682016, India

    Anoop B.S., Jayesh Puthumana, Vrinda Sukumaran, Cijo George Vazhappilly, Salini Kombiyil & Isaac Sarojini Bright Singh

  2. CSIR-National Institute of Oceanography, Regional Centre, Cochin, 682019, India

    Vrinda Sukumaran

  3. Department of Biotechnology, American University of Ras Al Khaimah, Ras Al Khaimah, UAE

    Cijo George Vazhappilly

  4. Department of Marine Biology, Microbiology and Biochemistry, School of Marine Sciences, Cochin University of Science and Technology, Fine Arts Avenue, Kochi, Kerala, 682016, India

    Rosamma Philip

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  1. Anoop B.S.
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  2. Jayesh Puthumana
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  3. Vrinda Sukumaran
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  4. Cijo George Vazhappilly
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  5. Salini Kombiyil
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  6. Rosamma Philip
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  7. Isaac Sarojini Bright Singh
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Contributions

ABS—carried out work at practical level and prepared the draft manuscript; JP—initiated the program and associated with the first author; VS, CGV, and SK—associated with the first author in executing the work at practical level; RP—assisted the corresponding author as co-investigator of the project; ISBS—conceptualized the work, generated funds, implemented the project, supervised the work, and prepared the manuscript in final form.

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Correspondence to Isaac Sarojini Bright Singh.

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B.S., A., Puthumana, J., Sukumaran, V. et al. A Novel Approach of Transducing Recombinant Baculovirus into Primary Lymphoid Cells of Penaeus monodon for Developing Continuous Cell Line. Mar Biotechnol 23, 517–528 (2021). https://doi.org/10.1007/s10126-021-10043-6

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