RESUMO
A fragment of 570 bp corresponding to exon 5 and 6 of integrin beta 2 (ITGB2) gene was amplified for screening D128G mutation in one hundred and fifty two buffaloes (Bubalus bubalis) which causes bovine leukocyte adhesion deficiency syndrome (BLAD) in cattle, as well as to ascertain polymorphism. TaqI PCR-RFLP revealed no such mutation thus indicating the absence of bubaline leukocyte adhesion deficiency (BuLAD) allele in animals under study. However, the polymorphism studies using MspI restriction enzyme revealed two genotypic patterns viz. AA pattern (bands of 293, 141, 105, and 31 bp) and BB pattern (bands of 293, 105, 77, 64, and 31 bp). The sequences of A and B alleles were submitted to the GenBank (EU853307 and AY821799).
Assuntos
Búfalos/genética , Búfalos/imunologia , Antígenos CD18/genética , Síndrome da Aderência Leucocítica Deficitária/veterinária , Alelos , Animais , Sequência de Bases , Bovinos , Análise Mutacional de DNA , Primers do DNA/genética , Testes Genéticos , Índia , Síndrome da Aderência Leucocítica Deficitária/genética , Síndrome da Aderência Leucocítica Deficitária/imunologia , Dados de Sequência Molecular , Polimorfismo de Fragmento de Restrição , Especificidade da EspécieRESUMO
Natural resistance associated macrophage protein 1 (NRAMP1), an integral transmembrane protein, is reported to influence the intraphagosomal microbial replication and thereby confer resistance to several intracellular pathogens in mice. In bovine, a significant association of (GT)(13) allelic variant of polymorphic microsatellite at 3' untranslated region (UTR) of NRAMP1 gene with natural resistance to brucellosis has been established. The present study was aimed to detect polymorphism at 3'UTR of NRAMP1 gene in Hariana breed of Bos indicus cattle and Holstein Friesian crossbred (B. indicusxBos taurus) cattle, and to determine the association of this polymorphism with resistance/susceptibility to brucellosis. The (GT)(n) polymorphism at 3'UTR in terms of variation in fragment length was determined using denaturing polyacrylamide gel analysis of radioisotope incorporated amplicon of 174 bp. Screening of a total of 100 samples (comprising 50 random samples of each breed) revealed that animals were of same genotype, i.e., homozygous (GT)(13)/(GT)(13). Sequencing of amplicons from representative animals confirmed the presence of (GT)(13) repeat. For association study, the animals that were positive in all three serological tests (viz., RBPT, STAT and ELISA) and had history of abortion were grouped as "affected"; whereas the animals that were negative in all these tests and completed third lactation without any history of abortion were grouped as "non-affected". Since, all animals belonging to either group were homozygous (GT)(13), association could not be established. However, the present study demonstrated that the presence of (GT)(13) allele even in homozygous condition could not provide enough resistance to brucellosis in a naturally infected herd.
Assuntos
Regiões 3' não Traduzidas/genética , Cruzamento , Brucelose Bovina/imunologia , Proteínas de Transporte de Cátions/genética , DNA Bacteriano/análise , Polimorfismo Genético , Alelos , Animais , Brucelose Bovina/genética , Bovinos , Cruzamentos Genéticos , Feminino , Masculino , Repetições de MicrossatélitesRESUMO
In the present study, water buffalo MHC (Bubu)-DRB cDNA was cloned and characterized. The 1022 base long-amplified cDNA product encompassed a single open reading frame of 801 bases that coded for 266 amino acids. The Bubu-DRB sequence showed maximum homology with the BoLA-DRB3*0101 allele of cattle. A total of seven amino acid residues were found to be unique for the Bubu-DRB sequence. The majority of amino acid substitutions was observed in the ß(1) domain. Residues associated with important functions were mostly conserved. Water buffalo DRB was phylogenetically closer to goat DRB*A.
RESUMO
The genetic diversity of MHC class II DQ genes was investigated in riverine buffalo (Bubalus bubalis) by PCR-RFLP and sequencing. Highly variable regions (exons 2-3) of DQ genes were amplified from 152 buffaloes and genotyped by PCR-RFLP. Alleles identified by differential restriction patterns were sequenced for the characterization. PCR-RFLP was a rapid method to discriminate between DQA1 and duplicated DQA2 genes in buffalo, however, the method appeared to be inadequate for determining the more complicated DQB genotypes. A total of 7 and 10 alleles were identified for DQA and DQB loci, respectively. Nucleotide as well as amino acid variations among DQ alleles particularly at peptide binding regions were high. Such variations were as expected higher in DQB than DQA alleles. The phylogenetic analysis for both genes revealed the grouping of alleles into two major sub-groups with higher genetic divergence. High divergence among DQ allelic families and the isolation of two diverse DQA and DQB sequences from individual samples indicated duplication of DQ loci was similar in buffalo to other ruminants.
Assuntos
Búfalos/genética , Búfalos/imunologia , Genes MHC da Classe II , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Primers do DNA/genética , Evolução Molecular , Duplicação Gênica , Variação Genética , Antígenos de Histocompatibilidade Classe II/genética , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Homologia de Sequência de AminoácidosRESUMO
In the present study, we explored structural and functional variations and possible duplication of the major histocompatibility complex (MHC)-DQA gene in water buffalo (Bubalus bubalis). Two cDNA sequences, amplified from one individual water buffalo, were designated as Bubu-DQA1 (DQA*0101) and -DQA2 (DQA*2001). The percentage of nucleotide and amino acid similarity between Bubu-DQA1 and -DQA2 revealed that these sequences display more similarity to alleles of respective DQA1 and DQA2 genes from other ruminant species than to each other. The phylogenetic analysis also revealed a considerably larger genetic distance between these two genes than between homologous genes from other species. The larger genetic distance between DQA*0101 and DQA*2001, and the presence of different bovine DQA putative locus specific amino acid motifs, suggests these sequences are non-allelic. This finding is consistent with DQA gene duplication in other ruminants.
Assuntos
Búfalos/genética , Búfalos/imunologia , Genes MHC da Classe II , Antígenos de Histocompatibilidade Classe II/genética , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Bovinos , Primers do DNA/genética , DNA Complementar/genética , DNA Complementar/isolamento & purificação , Duplicação Gênica , Dados de Sequência Molecular , Filogenia , Ruminantes/genética , Ruminantes/imunologia , Homologia de Sequência de Aminoácidos , Especificidade da EspécieRESUMO
In the present study, water buffalo MHC (Bubu)-DRB cDNA was cloned and characterized. The 1022 base long-amplified cDNA product encompassed a single open reading frame of 801 bases that coded for 266 amino acids. The Bubu-DRB sequence showed maximum homology with the BoLA-DRB3*0101 allele of cattle. A total of seven amino acid residues were found to be unique for the Bubu-DRB sequence. The majority of amino acid substitutions was observed in the β1 domain. Residues associated with important functions were mostly conserved. Water buffalo DRB was phylogenetically closer to goat DRB*A.