Cloning and Characterization of Pangasianodon hypophthalmus Growth Hormone Gene and its Heterologous Expression

Sekar, M and Singh, Shiva Dhar and Gupta, Subhodh (2014) Cloning and Characterization of Pangasianodon hypophthalmus Growth Hormone Gene and its Heterologous Expression. Applied Biochemistry and Biotechnology, 173 (6). pp. 1446-1468. ISSN 1470-8744

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Abstract

Pangasianodon hypophthalmus is one of the fast-growing catfish of freshwater origin, and its growth is attributed by the action of growth hormone (GH). In this study, the growth hormone gene (PhGH) of 3.0 kb was characterized, and it is composed of five exons and four introns and having characteristics of an upstream region that contains TATA, CAAT boxes, and binding sites of important transcription factors like Pit-1a, CRE, CREB, CREBP, Ap-1, SP1, and TBP. The full-length cDNA sequence of 1,069 bp was isolated using RACE technique, and it is composed of untranslated regions of 60 and 403 bp at 5′ and 3′, respectively, with an open reading frame of 603 bp that encodes a putative polypeptide of 200 amino acids with an estimated molecular mass of 22.57 kDa. The precursor of PhGH is composed of 22 amino acid signal peptides and 178 amino acid mature peptides. Five conserved Cys residues (Cys71, Cys135, Cys173, Cys190, and Cys198) and two possible sites of N-glycosylation (145th and 197th) were detected on GH polypeptide. The PhGH gene showed more than 90 % sequence similarity with other catfishes, and the phylogeny constructed revealed the close proximity of Siluriformes fishes with Cypriniformes fishes. The PhGH gene was observed to be expressed predominantly in pituitary tissues while weekly expressed in extrapituitary tissues. Further, the recombinant PhGH was expressed in Escherichia coli using His-tag expression vector pET 32(a), and the recombinant protein of ~23 kDa was confirmed by western blotting. Our findings suggest that the identified functional GH gene would provide basic information in transgenic studies aiming for faster growth rate. This recombinant growth hormone (GH) may be produced in large scale to exploit its growth-promoting function in other cultured fishes.

Item Type: Article
Uncontrolled Keywords: Growth enhancement, Aquaculture, Transgenic fish, Recombinant growth hormone,Tissue-specific expression
Subjects: Fish Genetics
Demersal Fishes > Cat fishes
Divisions: CMFRI-Kochi > Marine Capture > Demersal Fisheries Division
Subject Area > CMFRI > CMFRI-Kochi > Marine Capture > Demersal Fisheries Division
CMFRI-Kochi > Marine Capture > Demersal Fisheries Division
Subject Area > CMFRI-Kochi > Marine Capture > Demersal Fisheries Division
Depositing User: Arun Surendran
Date Deposited: 18 Nov 2021 05:25
Last Modified: 18 Nov 2021 05:25
URI: http://eprints.cmfri.org.in/id/eprint/15502

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