Primary Culture of Lepidopteran Adherent and Suspension Cells from Larval Testes of Bombyx mori

Abstract

In this study, first aim is to establish primary insect cell culture from larval testes of Bombyx mori. The other aim this to investigate the appropriate cells for establishing continuous cell lines. Testes obtained from 5 instar larvae of B. mori were cultured in Grace’s insect medium. Consequently, it was seen that primary cell culture from testes of B. mori resulted in suspension and adherent cell lines. In addition, this study showed that while adherent cells were not appropriate choice to initiate continuous cell line, suspension cells were suitable to establish continuous cell lines from Bombyx mori testes.

Keywords

• Insect cell culture,larval testes,Bombyx mori,primary cell culture

Bombyx mori’nin Larval Testislerinden Elde Edilen Yüzeye Bağımlı ve Süspanse Lepidopter Hücrelerinin Primer Kültürü

Abstract

Ç alışmada öncelikli olarak Bombyx mori’nin larval testislerinden primer böcek hücre kültürü oluşturulması amaçlanmıştır. Diğer bir yandan yetiştirilen bu hücrelerin sürdürülebilir hücre hatları oluşturma potansiyelleri araştırılmıştır. Testisler Bombyx mori’nin 5.evre larvalarından elde edilmiş ve Grace insect medium’da kültüre edilmiştir. Sonuç olarak, Bombyx mori 5.evre larva testislerinden elde edilmiş primer böcek hücre kültüründe hücrelerin süspanse ve yüzeye bağımlı olmak üzere iki şekilde bulunduğu gözlenmiştir. Ek olarak, bu çalışmada Bombyx mori testislerinden sürdürülebilir hücre hatları oluşturmak için yapışkan hücrelerin değil, süspanse hücrelerin daha uygun oldukları tespit edilmiştir

Keywords

• Böcek hücre kültürü,larval testis,Bombyx mori,primer hücre kültürü

References

1. G. Smagghe, C.L. Goodman, D. Stanley, Insect cell culture and applications to research and pest management, In Vitro Cell Dev. Biol. Anim., 45 (2009) 93-105.
2. D. Gundersen-Rindal, E.M. Dougherty, Evidence for integration of Glyptapanteles indiensis polydnavirus DNA into the chromosome of Lymantria dispar in vitro, Virus Res., 66 (2000) 27–37.
3. U. Mudiganti, R. Hernandez, D. Ferreira, D.R. Brown, Sindbis virus infection of two model insect cell systems: A comparative study, Virus Res., 122 (2006) 28–34.
4. S. Schütz, P. Sarnow, Interaction of viruses with the mammalian RNA interference pathway, Virology., 344 (2006) 151–157.
5. E. Lennan, R. Vandergaast, P.D. Friesen, Baculovirus caspase inhibitors P49 and P35 block virus-induced apoptosis downstream of effector caspase DrICE activation in Drosophila melanogaster cells, J. Virol., 81 (2007) 9319–9330.
6. H. Li, B.C. Bonning, Evaluation of the insecticidal efficacy of wild type and recombinant baculoviruses, In: Murhammer DW, editor. Methods in molecular biology series, Baculovirus and insect cell expression protocols, 2007, pp 379-404. Springer, New York.
7. D.E. Lynn, Lepidopteran insect cell line isolation from insect tissue, Methods Mol Biol., 388 (2007) 139-154.
8. M. Beckmann, K.J. Haack, Chemical pest control— insecticides for agriculture, Chem. Unserer. Zeit., 37 (2003) 88–97.

9. B. Lemaitre, J. Hoffmann, The host defense of Drosophila melanogaster, Annu. Rev. Immunol., 25 (2007) 697–743.
10. K. Iwabuchi, A continuous cell line derived from larval fat bodies of Thysanoplusia intermixta (Lepidoptera: Noctuidae), Appl. Entomol. Zool., 35 (2000) 245–249.
11. M.M. van Oers, D.E. Lynn, Insect cell culture. Encyclopedia of Life Sciences, John Wiley Sons Ltd., Chichester., (2010) 1-5.
12. D.E. Lynn, Novel techniques to establish new insect cell lines, In Vitro Cell. Dev. Biol. Anim., 37 (2001) 319- 321.
13. S. Soya, H. Can, M.S. Yıkılmaz, Primary insect cell culture from total embryo and embryonic brain tissue of Periplaneta americana: A preliminary study, Archives of Biological Sciences, 67 (2015) 1203-1208.
14. R. Goldschmidt, Some experiments on spermatogenesis in vitro, Proc. Natl. Acad. Sci., USA, 1 (1915) 220-222.
15. C. Stern, The growth of testes in Drosophila. I. The relation between vas deferens and testis within various species, J. Exp. Zool., 87 (1941) 113– 158.
16. E.L. Schmidt, C.M. Williams, Physıology of insect diapause. Assay for growth and differentiation 25. C.M. Williams, M.P. Kambysellis, In vitro action of hormone of Lepidoptera by the method of tissue culture, Anat. Rec. Suppl., 105 (1953) 174-187.
17. J. Mitsuhashi, Tissue Culture of the Rice Stem Borer, Chilo suppressalis WALKER (Lepidoptera: Pyralidae) I. Cell Migration from the Explanted Tissues of Diapausing Larvae, Japanese Journal of Applied Entomology and Zoology, 9 (1965) 217-224.
18. M. Kambysellis, C. Williams, In vitro development of insect tissues. II. The role of ecdysone in the spermatogenesis of silkworms, Biol. Bull., 141 (1971) 541–552.
19. Y. Kuroda, Studies on Drosophila embryonic cells in vitro. I. Characteristics of cell types in culture, Develop. Growth. Differ., 16 (1974) 55-66.
20. S. Yagi, T. Fukushima, Hormonal effect on cultivated insect tissues. II. Effects of juvenile hormone on spermatogenesis of the silkworm, Bombyx mori L. in vitro (Lepidoptera: Bombycidae), Appl. Entomol. Zool., 10 (1975) 77–83.
21. N. Kawamura, K. Sahara, In vitro cultivation of spermatocysts to matured sperm in the silkworm Bombyx mori, Dev. Growth. Differ., 44 (2002) 273–280.
22. H. Hamada, H. Fugo, Effect of fetal bovine serum on the enhancement of in-vitro cultivation of spermatocysts of the silkworm, Bombyx mori L. (Lepidoptera: Bombycidae), Zoolog. Sci., 24 (2007) 1251-1258.
23. M. Njogu, P.G. Ricketts, A.V. Klaus, Spermatogenic cyst and organ culture in Drosophila pseudoobscura, Cell. Tissue. Res., 341 (2010) 453-464.
24. S.M. Gärtner, C. Rathke, R. Renkawitz-Pohl, S. Awe, Ex vivo culture of Drosophila pupal testis and single male germ-line cysts: dissection, imaging, and pharmacological treatment, J. Vis. Exp., 11 (2014) 51868. Ecdysone. Proc. Natl. Acad. Sci., 63 (1969) 231.
25. J. Mitsuhashi, Determination of Essential Amino Acids for Insect Cell Lines. In: Maramorosch K, Mitsuhashi J, editors. Invertebrate Cell Culture Applications., Academic Press, INC; 1982. p. 9-53.