Vitellogenin, a biomarker for environmental estrogenic pollution, of Reeves' pond turtles: Analysis of similarity for its amino acid sequence and cognate mRNA expression after exposure to estrogen

Abstract

Vitellogenin (VTG), a biomarker for environmental estrogenic pollution, can be detected in the bloodstream of oviparous animals before morphological and functional abnormalities appear due to exposure to environmental estrogens. Reports observing VTG in turtles have been limited. We therefore cloned and sequenced a partial cDNA of VTG in Reeves' pond turtle, Chinemys reevesii. The cloned cDNA fragment possessed the start codon and 2,229 bp, encoding 743 amino acid residues. A sequence of deduced amino acid from the cDNA did not contain a high serine content, such as that which exists in phosvitin. Two N-glycosylation sites were found in the sequence. The sequence was compared to those of two birds (chicken and herring gull), one amphibian (Xenopus), and five fishes (carp, zebrafish, eel, haddock, and red seabream). The C. reevesii VTG was similar to that of herring gull (78%, value of positives), chicken (76%), Xenopus (69%), eel (63%), red seabream (62%), haddock (62%), carp (62%), and zebrafish (61%). The phylogenetic tree showed that C. reevesii VTG existed between the amphibian and birds, and it was present far from fish VTGs. A reverse transcription-polymerase chain reaction method was employed to detect the mRNA expression of the C. reevesii VTG through the use of primers designed from our sequence. The VTG mRNA expression (292 bp) was proven in the total RNA extraction from the liver of the juvenile turtles which were treated with estradiol-17β. The information herein would be useful for ecotoxicological studies using freshwater turtles and these findings are expected to contribute positively towards wildlife conservation.