Cloning and sequencing of European sea bass (Dicentrarchus labrax L.) growth hormone cDNA using polymerase chain reaction and degenerate oligonucleotides.
To clone and sequence the cDNA of the growth hormone of European sea bass (Dicentrarchus labrax L.) (sbGH), total pituitary RNA was reverse transcribed and amplified using polymerase chain reaction (PCR). Degenerate oligonucleotides, designed by comparing available GH cDNA sequences from related teleost species, were used as primers to amplify the 5' end and the core region of sbGH cDNA, while the 3' end was amplified according to the Rapid Amplification of cDNA ends (RACE) method. SbGH cDNA contains an open reading frame encoding a preprotein of 204 amino acids. The deduced amino acid sequence shows a putative signal peptide of 17 amino acids, suggesting that the mature hormone consists of 187 amino acids. Sequence comparison indicates a high degree of conservation of GH cDNAs within the Percoidei infraorder. Our procedure based on degenerate oligonucleotides and PCR provides a straightforward approach to clone GH cDNAs from other related bony fishes.
Doliana
R
R
Department of Biology, University of Padova, Italy.
Bortolussi
M
M
Colombo
L
L
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001492:Bass / Q000235:genetics*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D013006:Growth Hormone / Q000235:genetics*; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction
D015345:Oligonucleotide Probes; D013006:Growth Hormone; D004247:DNA