| Endogenous antimicrobial peptides are an important component in innate immunity of organism and defensins are a kind of positive endogenous antimicrobial peptides, which contain cysteine-rich and distribute abroad in the kingdoms of animal and plant. Defensins are a big family among endogenous antimicrobial peptides. According to the differences of the position and connect manner of cysteines, the property of precursors, the position of expression, defensins are devided into five subfamilies: α-defensin,β-defensin,θ-defensin, insect defensin and plant defensin. β-defensins are proved to be a component of antimicrobial barrier because they distribute prominently in muscosal epithelial cells of mammals and aves. It has been reported that β-defensins express extensively in the epithelial tissues of many organs in mammals (cow, sheep, goat, pig, mouse, rat, human, monkey) and aves (chicken, turkey). In this study, we have identified in camels a novel β-defensin, named camel β-defensin-1 (caBD-1). Total RNA was extracted from the tougue epithelial tissue of a camel and the cDNA encoding caBD-1 was amplified by the reverse transcription-PCR ( RT-PCR ) with the pair of primers which were designed according to the cDNA conserve sequences of reported ruminents' (cow, sheep and goat )β-defensins. The purified RT-PCR product was cloned in pBlueselect T vector. After the restriction endonuclease pattern analysis of reombinant plasmid, the cDNA was sequenced and the result of cDNA sequencing demonstrated that the caBD-1 is belong to the family of β-defensins because the cDNA contain a open reading frame (ORF) of 192 bases which encoded a 64 amino acid prepro-peptide and the prepro-peptide contained the β-defensin consensus sequence of six invariantly spaced cysteine residues. Obstaining full-length cDNA is the essential foundation of the research on gene structures, gene expressions and gene functions. Based on the known cDNA sequence of caBD-1, we successfully cloned the 3ˊcDNA end of caBD-1 using anchored PCR RACE with a sequence specific primer as up primer and 3ˊsites adaptor primer as down primer. In addition, inverse nestd PCR can rapidly amplify 5ˊcDNA end by using a specific RT primer whose 5ˊend is phosphated and two pairs specific inverse nested PCR primers, which are designed basing on the known cDNA sequence of caBD-1. The caBD-1 cDNA which come from caBD-1 mRNA by RT is encircled, then inverse nestd PCR is performed. Contrasting with the anchored PCR, inverse nestd PCR has better specificity and higher amplification effects. It is a very effective method of amplifying 5ˊcDNA ends. The full-length caBD-1 cDNA is 322bp and includes a ORF of 192bp which encode the prepro-caBD-1 of 64 amino acid residues. With computer software we predict that the prepro-caBD-1 contain a signal peptide of 20 amino acid residues, a propiece of 6 amino acid residues, a muture peptide of 38 amino acid residues. In the muture peptide there are six invariantly spaced cysteine residues and 9 positive amino acid residues ( 2 Arginine-R, 7 Lysine-K ) , there is no negative amino acid residue in the muture peptide. Therefore the caBD-1 is a positive peptide which molecular weight is 4007.94 dalton and pI is 9.71. The results of pair distances of the cDNA and prepro-peptide sequences of caBD-1 and otherβ-defensins in human, other mammals and aves by computer software indicate that the relative relationship of caBD-1 to pBD-1 is highest (78.1%, 76.6%), to the β-defensins of cow, sheep and goat is middle (67.7%～74.9%, 51.6%～60.9%), to the β-defensins of human, monkey, mouse, rat, chicken and turkey is lowest (mostly less than 30%). Therefore, we can conclude that the relationship of caBD-1 to pBD-1 is closer than caBD-1 to other ruminant's (cow, sheep and goat) β-defensins with the sequences of cDNA and prepro-peptide. The result demonstrates the diversity of defensins evolution In order to examine the organs that express possibly caBD-1 mRNA in camels, we designed again another pair of primers, which expecte...
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