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

• Published in: Marine biotechnology (New York, N.Y.) N.Y.), 2021-08, Vol.23 (4), p.517-528
• Main Authors: B.S., Anoop, Puthumana, Jayesh, Sukumaran, Vrinda, Vazhappilly, Cijo George, Kombiyil, Salini, Philip, Rosamma, Singh, Isaac Sarojini Bright
• Format: Article
• Language: English
• Subjects: Aquatic crustaceans; Baculovirus; Biomedical and Life Sciences; Cell culture; Cell lines; Crustaceans; Culture media; Engineering; Freshwater & Marine Ecology; H-Ras protein; Immortalization; Life Sciences; Lymphoid cells; Marine crustaceans; Microbiology; Original Article; Penaeus monodon; Promoters; Recombinants; Senescence; Sodium; Sodium butyrate; Telomeres; Transduction; Zoology
• ISSN: 1436-2228; 1436-2236
• DOI: 10.1007/s10126-021-10043-6

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.