OPEN ACCESS PEER-REVIEWED | RESEARCH ARTICLE

Main Article Content

Authors

Wartono Hadie
nFN Sularto
Lies Emmawati Hadie
Angela Mariana Lusiastuti
nFN Alimuddin
Evi Tahapari
Huria Marnis

Abstract

Bacterial disease in catfish has many detrimental to farmers, especially in the larvae stadium in the hatchery up to ready size of stocking seed. Therefore we need technology that can produce disease resistant of catfish. In that regard, one of the antimicrobial enzyme namely lysozyme which plays an important role in the maternal immunity can be introduced into the catfish genome. This study aimed to evaluate the success of the lysozyme gene insertion into the genome of striped catfish to generate maternal disease resistance. Gene transfer method was done by using electroporation on striped catfish spermatozoa. Lysozyme plasmid electroporation performed with a dose 100^g ml-1 and 125V.cm-1, pulse length 30 milliseconds with a pulse interval 0.1 milliseconds and pulse number 5 times. Electroporated spermatozoa were used to fertilize the egg. Lysozyme gene insertion success was indicated by tests performed on embryogenesis, larval, and seed stage. The test results at both DNA and RNA level showed a positive result. Individual that brings the lysozyme gene will be used as a candidate for breed line maternal immune as other ways to formation of resistant disease varieties. AbstrakSerangan penyakit bakterial pada ikan patin telah banyak merugikan para pembudi daya ikan patin terutama pada seg-men perbenihan hingga ukuran siap tebar. Oleh karena itu diperlukan teknologi yang mampu menghasilkan ikan patin yang tahan penyakit. Berkaitan dengan hal itu, salah satu enzim antimikroba yaitu lisozim yang memainkan peranan penting dalam imunitas bawaan dapat diintroduksikan ke dalam genom ikan. Penelitian ini bertujuan untuk mengevalu-asi keberhasilan insersi gen lisozim ke dalam genom ikan patin siam sebagai galur ikan patin tahan penyakit. Transfer gen dilakukan dengan menggunakan teknik elektroporasi pada spermatozoa ikan patin siam. Elektroporasi dilakukan dengan konstruksi gen lisozim berupa plasmid DNA dosis 100 ^g.ml-1 dengan voltase 125 V. cm-1, panjang kejutan 30 milidetik dengan interval kejutan 0.1 milidetik dan jumlah kejutan lima kali. Spermatozoa hasil elektroporasi digunakan untuk membuahi telur. Pengujian keberhasilan insersi gen lisozim dilakukan pada tahap embrio larva, dan pada benih. Hasil pengujian, baik pada tingkat DNA maupun pada tingkat RNA dari sampel spermatozoa dan larva (whole cell), memperlihatkan hasil yang positif. Individu ikan patin yang membawa gen lisozim dan telah terintegrasi ke dalam ge-nomnya akan digunakan sebagai kandidat dalam pembentukan galur ikan patin tahan penyakit.

Keywords:
electroporation , lisozyme gene , gene expression , RNA , DNA , disease resistance

Downloads article

Download data is not yet available.

Article Details

Copyright
   

Copyright (c) 2017 Jurnal Iktiologi Indonesia

  Authors who publish with this journal agree to the following terms:  
  a.)
Authors retain copyright and grant the journal right of first publication with the work simultaneously licensed under a Creative Commons Attribution License that allows others to share the work with an acknowledgement of the work's authorship and initial publication in this journal.  
  b.) Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgement of its initial publication in this journal.  
  c.) Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).  
       
  Creative Commons License
Jurnal Iktiologi Indonesia by Masyarakat Iktiologi Indonesia (MII) is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Permissions beyond the scope of this license may be available at https://jurnal-iktiologi.org
 

 

References

Balfry SK, Iwama GK. 2004. Observation on the inherent variability of measuring lysozyme activity in Coho salmon Onchorhyn-chus kissutch. Journal Comparative Biochemistry and Physiology. 138B: 207-211.

Burge EJ, Madigan DJ, Burnett LE, Burnett KG. 2007. Lysozyme gene expression as a molecular marker of hemocyte location in Pacific white shrimp, Litopenaeus vannamei, after injection with Vibrio. Fish and Shellfish Immunology, 22(4): 327-339.

Dewi RRSPS, Alimuddin, Sudrajat AO, Sumantadinata K. 2012. Efektivitas transfer gen PhGH pada ikan patin siam (Pangasianodon hypophthalmus). Jurnal Riset Akua-kultur, 7(2): 171-180.

Dewi RRSPS, Alimuddin, Sudrajat AO, Sumantadinata K, Hayuningtyas EP. 2013. Pola ekspresi gen enhaced green fluorescent protein pada embrio dan larva ikan patin siam (Pangasianodon hypophthalmus). Jurnal Riset Akuakultur, 8(3): 339-346.

Fernandes JMO, Kemp CD, Smith VJ. 2004. Two novel muramidases from skin mucosa of rainbow trout (Onchorhynchus my-kiss). Comparative Biochemistry and Physiology, 138B: 53-64.

Fletcher GL, Hobb RS, Evans RP, Shears MA, Hahn AL, Hew CL. 2011. Lysozyme transgenic Atlantic salmon (Salmo salar L). Aquaculture Research, 42(3): 427-440.

Grinde B. 1989. A lysozyme isolated from rainbow trout acts on mastitis pathogens. Microbiology Letters, 60(2): 179-182

Hadie W, Lusiastuti AM, Sularto, Tahapari E. 2010a. Imunitas maternal terhadap Aeromonas hydrophila: pengaruhnya terhadap fekunditas dan daya tetas ikan patin siam

(Pangasianodon hypophthalmus). Jurnal Riset Akuakultur, 5 (2): 229-235

Hadie W, Kusrini E, Priyadi A, Alimuddin. 2010b. Penyisipan gen warna pada ikan Carasius auratus menggunakan metode elektroforasi dalam upaya meningkatkan kualitas ikan hias. Jurnal Riset Akuakul-tur, 5(3): 335-343.

Hadie W, Tahapari E, Hadie LE, Sularto. 2010c. Efektivitas persilangan dalam peningkatan produktivitas ikan patin melalui hibridisasi antar spesies. Jurnal Iktiologi Indonesia, 10(2): 179-184.

Hikima J, Hirono I, Aoki T. 2003. The lysozyme gene in fish. Aquatic Genome. Tokyo University of Fisheries. Tokyo. 301-309.

Hew CL, Fletcher GL. 2001. The rule of aquatic biotechnology in aquaculture. Aquaculture, 197(1-4): 191-204.

Knight DE, Scrutton MC. 1986. Gaining access to the cytosol: the technique and some application of electropermeabilization. Biochemical Journal. 234(3): 497-506.

Magnadttir B, Lange S, Gudmundsduttir S, Bog-wald J, Dalmo RA. 2005. Ontogeny of humoral immune parameters in fish. Fish and Shelfish Immunology, 19(5): 429-439.

Patrzykat A, Douglas SE. 2005. Antimicrobial peptides: cooperative approaches to protection. Protein and Peptide Letters, 12(1): 19-25.

Saurabh S, Sahoo PK. 2008. Lysozyme: an important defence molecule for fish innate immune system. Aquaculture Research, 39(3): 223-239.

Tort L, Balasch JC, Mackenzie S. 2003. Fish immune system. A crossroads between innate and adaptive responses. Immunolo-gia, 22(3): 277-286.