6-Desaturase-Like Encoding Gene Introduction in Catfish (Clarias gariepinus)

Anny Hary Ayu, Alimuddin Alimuddin, Dedi Jusadi

Abstract


African catfish (Clarias gariepinus) is one of the economically valuable aquaculture fish species in Indonesia. This research was aimed to produce F0 transgenic catfish carrying masou salmon Δ6-desaturase-like (OmΔ6FAD) gene. The Δ6-desaturase enzyme is involved in highly unsaturated fatty acid biosynthesis. Transgenic catfish was produced by sperm-mediated gene transfer using electroporation method. In this study, as the first step, sperms were electroporated with three different OmΔ6FAD concentration (25, 50, and 100 µg mL-1) to have the highest sperm viability after electroporation (125 V/cm, pulse frequency 5 times, pulse length 30 millisecond, pulse interval 0.1 second). The highest sperm viability and sperm carrying OmΔ6FAD were obtain at 100 µg mL-1. This concentration was then used to produce F0 transgenic catfish in the second step. Sperm motility, sperm viability, fertilization rate, hatching rate, and larval survival at 14 days after hatching were the same as the controls (p>0.05). Genomic DNA was extracted from caudal fin and then used as template to identify transgenic F0 by PCR method using specific primer for OmΔ6FAD gene. The PCR result showed that 53.84% of F0 carried OmΔ6FAD gene. The result of fatty acid analysis showed that EPA and DHA contents of F0 transgenic fish and non-transgenic fish were similar.

 

Keywords: catfish, Δ6-desaturase-like gene, fatty acids, electroporation


Full Text:

73-79

References


Ackman, R.G. 1982. Fatty acid composition of fish oil. London: Press Ltd.

Alimuddin, Yoshizaki G, Viswanath K, Satoh S, Takeuchi T. 2005. Enhancement of EPA and DHA biosynthesis by over-expression of masu salmon Δ6-desaturase-like gene in zebrafish. Transgenic Research. 14: 159-165.

Chen TT, Chen MJ, Lu JK. 2009. Transfer of foreign DNA into aquatic animal by electroporation. Dalam Nakamura H (ed.). Electroporation and sonoporation in developmental biology. Springer. 229-237.

Chou CY, Hong LS, Tsai HJ. 2001. Uniform GFP-expression in transgenic medaka (Oryzias latipes) at the F0 generation. Transgenic Research. 10: 303-315.

De la Cueva CFI, Rigau T, Bonet S, Mirό J, Briz M, Rodrίguez-Gil JE. 1997. Subjecting horse spermatozoa to hypoosmotic incubation: effects of ouabain. Theriogenology. 47:84-765.

Guest WC, Avault JW, Rousel JA. 1976. Spermatogeny study of channel catfish (Ictalurus punctatus). Transactions of the American Fisheries Society. 105: 463-468.

Gusrina. 2011. Introduksi dan Ekspresi Gen Hormone Pertumbuhan Ikan Nila (Oreochromis niloticus) pada Ikan Lele (Clarias sp.) [disertasi]. Bogor (ID): Institut Pertanian Bogor.

Gunawan ER, Suhendra D, Handayani SS, Kurniawati L, Murniati, Nurhidayanti. 2014. Analisis Kandungan Asam Lemak Omega-3 dan 6 Pada Bagian Kepala Dan Badan Ikan Lele (Clarias Sp) Melalui Reaksi Enzimatis. Prosiding Seminar Nasional Kimia. Jurusan Kimia FMIPA, Universitas Negeri Surabaya.

Holland B, Brown J, Buss DH. 1993. Fish and Fish Products. The third supplement to McCance & Widdowson’s The Composition of Foods (5th edition), London. HMSO.

Lauritzen L, Hansen H S, Jorgensen MH, Michaelsen KF, 2001. The essentiality of long chain n-3 fatty acids in relation to development and function of the brain and retina. Progress in Lipid Research. 40: 1-94.

Sambrook J, Fritssch EF, Maniatis T. 1989. Molecular Cloning. A laboratory manual, 2nd ed. USA: Cold spring harbor laboratory press.

Sarmasik A, Jang IK, Chung CZ, Lu JK, Chen TT, 2001. Transgenic live-bearing fish and crustacean produced by transforming immature gonads with replication-defective pantropic retrovial victor. Marine Biotechnology. 3: 470-477.

Simopoulos AP. 1991. Omega-3 fatty acids in health and disease and in growth and development. The American Journal of Clinical Nutrition. 54:438–463.

Sprecher H. 2000. Metabolism of highly unsaturated n-3 and n-6 fatty acids. Biochimica et Biophysica Acta. 1486:219-231.

Symonds JE, Walker SP, Sin FYT. 1994. Development of mass gene transfer method in Chinook salmon: optimization of gene transfer by electroporated sperm. Clinical Biochemistry. 40:343-350.

Tocher DR, Leaver MJ, Hodgson PA. 1998. Recent advances in the biochemistry and molecular biology of fatty acyl desaturases. Lipid Research, 37:73-117.

Tsai HJ. 2000. Electroporated sperm mediation of gene transfer system for finfish and shellfish. Molecular Reproduction and Development. 56: 281-284.




DOI: http://dx.doi.org/10.20884/1.oa.2017.13.2.188

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.

 

Lisensi Creative Commons

Omni-Akuatika de Fisheries and Marine Science Faculty - Jenderal Soedirman University est mis à disposition selon les termes de la licence Creative Commons Attribution 4.0 International.

Fondé(e) sur une œuvre à www.ojs.omniakuatika.net.
Les autorisations au-delà du champ de cette licence peuvent être obtenues à www.ojs.omniakuatika.net.