Total RNA was isolated from pituitary of grouper (
Epinephelus coioides
),and SMART cDNA was synthesized from 50ng RNA.Two genes in the growth hormone/prolactin family were cloned from SMART cDNAs plasmid library.The growth hormone (GH) cDNA contains 938 bp and encodes 204 amino acids (aa).The prolactin (PRL) cDNA contains 1429 bp and encodes 212 aa.The deduced aa sequences of grouper GH,PRL and SL were compared with those of other eight species.Average genetic distances and a dendrogram showing the hierarchical structure of affinity among different nine fishes were implemented in Mega 2 and CLUSTAL W1.64 b.Branch and bound strap analysis (1000 replicates) were performed and the degree of support for particular nodes were produced.The three phylogenetic trees drawn from the whole aa sequence of GH,PRL and SL were almost identical,
Epinephelus coioides
,
Perca flavescens
,
Sparus aurata
and
Paralichthys olivaceus
were clustered into one group,
Oncorhynchus keta
and
Oncorhynchus mykiss
were clustered into one group,
Carassius auratus
and
Ictalurus punctatus
were clustered into one group,while
Anguilla japonica
alone was clustered into another group.Alignment of the deduced aa sequence and the phylogenetic tree based on GH/PRL/SL showed the same clustering as the present hierarchy of fish and a higher conservation of SL than GH and PRL among various fish species is suggested in this text.Although there is only 24%-31% identity between the three hormones overall,the C-terminal region shows a higher identity.Grouper SL appears to be more closely related to GH (30.8%) than to PRL (25.6%),which is slightly more than the identity of GH to PRL (24.1%).
