Molecular Cloning and Expression Analysis of HGITM cDNA in Trionyx sinensis

Abstract:

In this study, we obtained the full-length cDNA of the GHITM gene from Trionyx sinensis for the first time using the RACE (rapid-amplification of cDNA ends) method. The full-length cDNA sequence was 2650 bp, including a 123 bp 5-UTR, 1477 bp 3-UTR, and 1050 bp open reading frame (ORF) that encoded 349 amino acid residues. The isoelectric point (pI) of this peptide was 10.01, and the molecular mass was 37.12 kDa. The amino acid sequence was composed of the extracellular region, the transmembrane region, and the intracellular region. The transmembrane region was composed of 7 transmembrane domains. The phylogenetic analysis of the amino acid sequences showed that T. sinensis, Chrysemys picta bellii, and Chelonia mydas belonged to the same branch, the three species of crocodiles formed a branch, and birds formed another branch. The expression of GHITM in different tissues was also analyzed with quantitative real-time PCR. The results showed that GHITM was expressed in all the tested tissues, including the liver, pituitary, muscle, spleen, kidney, heart, intestine, and gonad, with the high expression observed in the liver, muscle, and pituitary. At the specifications of 50 g and 500 g, the expression of GHITM was significantly higher in the liver of males than females (P<0.05). Low temperature incubation can inhibit the expression of GHITM in fetal embryos (P<0.05). The results indicated that GHITM was related to the growth and embryo development of T. sinensis.