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:: Volume 15, Issue 4 (12-2021) ::
JAD 2021, 15(4): 47-55 Back to browse issues page
Gene transfer biotechnology in zebrafish as a gene transfer research model in farmed aquatic animals
ُShirin Jamshidi *, Ashkan Banan
International Sturgeon Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Rasht, Iran
Abstract:   (570 Views)
The polymerase chain reaction (PCR) was applied to create the colored transgenic ornamental zebrafish with the specific primers designed for muscle promoter of two genes and genes fragments cloned in Tol2 vector containing GFP gene in the ORF region. The 2000 bp and 2300 bp fragments of MYLZ2 and CKMB promoters' genes respectively were amplified by PCR, cloned into Tol2 plasmid expression vector containing SalI restriction enzyme site and transformed into E. coli Dh5α. Positive clones were tested for having fragments by colony PCR with anchor primers and subcultured for extracting recombinant plasmid miniprep. The extracted recombinant plasmid was tested with SalI enzyme digestion and sequenced with universal primers. Positive recombinant plasmids were applied for injection with transcript produced synthetically in the lab into zebrafish one-cell embryo. In this study, 2000 bp and 2300 bp fragments of MYLZ2 and CKMB promoters, respectively, were cloned in an expression vector Tol2 and confirmed by PCR, sequencing and enzymatic digestion. Also, recombinant plasmid was subcultured and purified for injection with a synthetic transcript. One-cell embryos injected by coinjection materials (recombinant plasmid and transcript) were tested by fluorescence microscope for positive GFP expression, where a few offspring of F0 generation were positive for GFP expression in their muscles. Recombinant plasmids with muscle-specific promoters (MYLZ2 and CKMB) were able to induce expression of GFP in some F0 embryos in a transient form and could be applied for consolidation in the next fish generation. 
Keywords: cloning, transgenic colorful zebra fish, recombinant Tol2 vector, transcript
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Type of Study: Research | Subject: Special
Received: 2019/12/14 | Accepted: 2020/01/24 | Published: 2021/12/31
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Jamshidi ُ, Banan A. Gene transfer biotechnology in zebrafish as a gene transfer research model in farmed aquatic animals. JAD. 2021; 15 (4) :47-55
URL: http://aqudev.liau.ac.ir/article-1-677-en.html

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Volume 15, Issue 4 (12-2021) Back to browse issues page
نشریه توسعه آبزی پروری Journal of Aquaculture Development
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