Heat shock proteins are a family of molecular chaperones that are involved in many aspects of protein homeostasis. In the present study, a full-length cDNA, encoding the constitutively expressed 70-kDa heat shock cognate protein （Hsc70）, was isolated from swordtail fish （Xiphophorus helleri） and designated as XheHsc70. The Xhehsc70 cDNA was 2 104 bp long with an open reading frame of 1 941 bp, and it encoded a protein of 646 amino acids with a theoretical molecular weight of 70.77 kDa and an isoelectric point of 5.04. The deduced amino acid sequence shared 94.1%-98.6% identities with the Hsc70s from a number of other fish species. Tissue distribution results show that the Xhehsc70 mRNA was expressed in brain, heart, head kidney, kidney, spleen, liver, muscle, gill, and peripheral blood. After immunization with formalin-killed Vibrio alginolyticus cells there was a significant increase in the XhehscT0 mRNA transcriptional level in the head kidney of the vaccinated fish compared with in the control at 6, 12, 24, and 48 h as shown by quantitative real time RT-PCR. Based on an analysis of the amino acid sequence of XheHsc70, its phylogeny, and Xhehsc70 mRNA expression, XheHsc70 was identified as a member of the cytoplasmic Hsc70 （constitutive） subfamily of the Hsp70 family of heat shock proteins, suggesting that it may play a role in the immune response. The Xhehsc70 cDNA sequence reported in this study was submitted to GenBank under the accession number JF739182.
[Objective] The aim was to study the sequence of aroA gene （synthetic gene in metabolic pathway of aromatic amino acids） of a pathogenic bacterium （Flavobacterium johnsoniae） causing gill-rote disease.[Method] Genomic DNA of strain M165 of F.johnsoniae was used as a template,three specific nested primers of 5＇end and 3＇end and arbitrary primer were used to amplify the aroA gene of strain M165 through Thermal asymmetric interlaced PCR （TAIL-PCR）.And the obtained sequence was analyzed.[Result] The electrophoresis determination result showed that the size of amplified product was consist with the expected product; sequencing analysis suggested that the full-length of aroA gene was 1 230 bp,enconding 410 amino acids.The amino acid sequence of aroA protein showed the highest level of similarity to amino acids sequence of aroA protein of F.johnsoniae.[Conclusion] TAIL-PCR provided a simple and efficient new method for the cloning of the gene sequence.Obtaining of full-length of aroA gene provided a foundation for further investigation on the effects of nutrition correlation factors such as aroA on the virulence of and virulence of F.johnsoniae.
