Structure and antigenicity analysis of the IgG gene for Nyctereutes procyonoides.

OBJECTIVE: Nyctereutes procyonoides immunoglobulin G (IgG) gene is partially cloned. MATERIAL AND METHODS: In order to obtain a certain length (966bp) of Nyctereutes procyonoides immunoglobulin G (IgG), two pairs of primers are designed according to the conserved nucleotide sequence of canine (GenBank:AF354265, AF354265, AF354266, AF354267) and mink (GenBank: L07789). Using Bioinformatics technology and Western-blot to analyze antigenicity of Nyctereutes procyonoides IgG-B gene. RESULTS: The homology for nucleotide sequence of IgG between Nyctereutes procyonoides and canine (IgG A, IgG B, IgG C, IgG D), mink, Homo sapiens, Oryctolagus cuniculus, Mus musculus, Anas platyrhynchos and gallus were respectively (88.1%, 93.6%, 85.4%, 87.2%), 83.7%, 74.8%, 71.8%, 69.2%, 51.6%, 48.4%. It can be seen that there was high homology of aminoacid sequence between IgG of Nyctereutes procyonoides and IgG (A, B, C, D) of canine. And the serum antibody of Nyctereutes procyonoides had obviously cross-reaction with HRP conjugated rabbit anti-dog IgG, compared with those of canine, oryctolagus cuniculus, mus musculus, mink, gallus. CONCLUSIONS: We successfully got Nyctereutes procyonoides immuneglobulin G (IgG) gene (Gen- Bank: KM010191). There is the closest ties of consanguinity of IgG exist between Nyctereutes procyonoides and canine among the mammal through the genetic evolution. The detection and treament of canine distemper can be used on Nyctereutes procyonoides.

[Expression of Edwardsiella tarda luxS gene at different growth stage].

OBJECTIVE: Purpose of this work was to explore the distribution of LuxS/AI-2 quorum sensing system in Edwardsiella, and analyze expression characteristics and biological function of the key gene luxS accompanying the growth of Edwardsiella. METHODS: The full-length of AI-2/LuxS of Edwardsiella tarda was cloned by PCR based on the sequence on NCBI, then characteristics and conservative structure of this protein-coding gene were analyzed using web database and bioinformatics tools. The anti-rabbits serum was prepared after this protein was purified through prokaryotic expression. The expression level of luxS gene was analyzed during different growth stages using Western blot and further the distribution of luxS gene in Edwardsiella tarda was studied by this technique. To explore whether the specific LuxS is AI-2 dependent we used the method of antibody neutralization to analyze the effect of the anti-rabbits serum on the growth of Edwardsiella tarda. RESULTS] The luxS gene was obtained by PCR, its length was 516 bp, and the sequence was highly conserved in Edwardsiella tarda. Results of Western blot analysis showed that LuxS expression level was the lowest in the lag phase and began increasing when entered index phase. It reached the peak in the late index phase and decreased in decline phase. Moreover, Antibody neutralization results showed that, it can elongate the growth plateau phase, but it has no significant effect on bacterial growth. CONCLUSION: The key gene of luxS was highly conserved, and LuxS/AI-2 was widely distributed among Edwardsiella tarda. The expression level of luxS gene was different during every growth period, expression of LuxS protein reached the highest level in the late index phase.

Genetic and antigenic analysis of mink's immunoglobulin G Fc region.

Mink's immunoglobulin (Ig) G Fc gene was cloned, the gene was analyzed by phylogenetic analysis, and western blot was done to prove that the detection of distemper and canine parvovirus in dogs and minks can be universal. In order to get the certain length of Fc segment gene, a pair of primers is designed, which according to the Fc segment gene sequences of mink's IgG (L07789) published by GenBank, extracted total RNA from the spleen of minks and amplified it by RT-PCR. The results showed that the Fc segment gene contained 606 bp. Then it was sequenced after the amplified fragments were cloned into the vector PEasy-T1. Then the genetic evolution was analyzed. An antibody hybridization test was done through western blot. The results showed that nucleotide sequence homologies between minks and canines were 85%, and amino acid sequence homologies between minks and canines were 80.5%. Mink IgG heavy chain can effectively combine to anti-dog IgG by western blot. It was concluded that mink's and dog's IgG Fc had the closest relationship in mammalian through the analysis of the genetic evolution. Based on the above analysis and related literature, we concluded that we could detect mink diseases with a dog diagnosis reagent, or treat mink diseases with dog antiserum.