Molecular Cloning And Expression Analysis Of Polyamine Metabolism-related Genes In Piglet

In order to study the function of some Polyamine metabolism-related genes of pig, according to mouse and human similar gene sequence searched in their ESTs and UniGene, we designed the primers of the seven genes, successfully cloned by RT-PCR. They were Arginine decarboxylase (ADC) , Ornithine decarboxylase (ODC), Ornithine aminotransferase (OAT), Ornithine decarboxylase antizyme 1 ( OAZ1), Ornithine decarboxylase antizyme 2 (OAZ2), Ornithine decarboxylase antizyme 3 (OAZ3) and Polyamine-modulated factor 1 (PMF1). Finally, we analyzed the tissue expression of porcine the genes'mRNA by the semi-quantitative RT-PCR before and after weaning, with 18S rRNA as the internal standard. The results were as follows:ADC: The sequence of pig ADC consisted of 1602 bp, with an open reading frame of 1380 bp. The predicted protein sequence was 460 amino acid residues. Identity analysis showed that the nucleotide sequence of pig ADC shared 91.1% and 87.4% homology with that of human and mouse, respectively. The homology of the deduced protein sequence was 89.7% and 87%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a low expression of porcine ADC mRNA in body and no finding in some tissues. After weaning, the expression increased significantly in appendix and rectal, while decreased slightly in other tissuesODC: The sequence of pig ODC consisted of 1422 bp, with an open reading frame of 1383 bp. The predicted protein sequence was 461 amino acid residues. Identity analysis showed that the nucleotide sequence of pig ODC shared 86.1%, 84.8% and 84.4% homology with that of human, mouse and rat respectively. The homology of the deduced protein sequence was 93.1%, 89.8% and 90.5%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a wide spread expression of porcine ODC mRNA. After weaning, the expression increased significantly in appendix and rectal, while decreased slightly in jejunum, ileum and colon.OAT: The sequence of pig OAT consisted of 1465 bp, with an open reading frame of 1317 bp. The predicted protein sequence was 439 amino acid residues. Identity analysis showed that the nucleotide sequence of pig OAT shared 87.1%, 82.2% and 82.2% homology with that of human, mouse and rat respectively. The homology of the deduced protein sequence was 91.8%, 90.5% and 89.8%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a wide spread expression of porcine OAT mRNA. After weaning, the expression increased significantly in renal, while it decreased slightly in duodenum, jejunum and ileum.OAZ1: The sequence of pig OAZ1 consisted of 874 bp, with an open reading frame of 681 bp. The predicted protein sequence was 227 amino acid residues. Identity analysis showed that the nucleotide sequence of pig OAZ1 shared 92.5%, 85.1% and 84.3% homology with that of human, mouse and rat respectively. The homology of the deduced protein sequence was 93.4%, 83.8% and 82.5%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a wide and rich spread expression of porcine OAZ1 mRNA. After weaning, the expression of porcine OAZ1 mRNA increased in different magnitude, except in stomach.OAZ2: The sequence of pig OAZ2 consisted of 678 bp, with an open reading frame of 568 bp. The predicted protein sequence was 189 amino acid residues. Identity analysis showed that the nucleotide sequence of pig OAZ2 shared 98.1%, 97.9% and 97.2% homology with that of human, mouse and rat respectively. The homology of the deduced protein sequence was 99.5%, 98.9% and 98.9%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a wide and rich spread expression of porcine OAZ2 mRNA. After weaning, all the expression increased in different magnitude, especially in the appendix.OAZ3: predicted protein sequence was 461 amino acid residues. Identity analysis showed that the nucleotide sequence of pig OAZ3 shared 89.6%, 85.5% and 84.8% homology with that of human, mouse and rat respectively. The homology of the deduced protein sequence was 90.4%, 87.5% and 84.2%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a wide spread expression of porcine OAZ3 mRNA and low expression in some tissues. After weaning, the expression of porcine PMF1 mRNA, in the intestinal tissues, increased in different magnitude. PMF1: The sequence of pig PMF1 consisted of 703 bp, with an open reading frame of 615 bp. The predicted protein sequence was 460 amino acid residues. Identity analysis showed that the nucleotide sequence of pig PMF1 shared 86.9% and 76.6% homology with that of human and mouse, respectively. The homology of the deduced protein sequence was 84% and 70.9%, respectively. The analysis of the semi-quantitative RT-PCR showed that, there was a wide spread expression of porcine PMF1 mRNA and low expression in cerebellum and stomach. The expression of porcine PMF1 mRNA was similar to the AZ family in jejunum, ileum and colonic, but decreased slightly after weaning.