Bovine NRAMPI gene promoter sequence and polymorphism
Bovine tuberculosis is still present in different areas of Italy. Identification of resistant genotypes could be useful both to integrate the eradication programme and to improve a healthy-animal oriented selection. Moreover Mycobacterial infections are interesting models to study mechanisms of resistance to intracellular microbial antigens. As a candidate gene for M. bovis infection resistance in cattle we studied NRAMP1 gene (Natural Resistance Associated Macrophage Protein 1), formerly known as Bcg/Lsh/Ity gene. NRAMP1 gene owns to a family of proteins, the metal-ion carrier protein family, which is extremely conserved among different organisms from bacteria to superior vertebrates, suggesting a key role in all living organisms. Particularly NRAMP1 is involved in intracellular parasite immune control, including M. bovis. It could be involved in the phagosoma maturation process as it modulates the intraphagosoma content of ions, generating toxic radical and allowing the bacteriostatic and bactericidal action of macrophages. NRAMP1 gene is mapped on chromosome 2 in cattle (position q43-q44). It consists of 15 exons, codifying a 60kd membrane protein. NRAMP1 promoter region presents anchor sites for immune-active molecules, which represent NRAMP1 transcriptional factors. To amplify bovine NRAMP1 promoter we designed 8 primer pair set on ovine, murine and human sequences of conserved homologous regions. Using this primer panel we amplified genomic DNA from 10 cows out of 7 autochthonous Italian breeds. Particularly the animals were 4 Italian Friesian, 1 Aosta Red Pied, 1 Aosta Black Pied, 1 Aosta Chestnut, 1 Oropa Red Pied, 1 Rendena, 1 Burlina cattle. Out of the 8 primer pair panel, 3 primer pairs specifically amplified by PCR 3 different amplicons (995, 458 and 737pb), partially overlapping. Direct sequence of amplicons, carried on by Big Dye Terminators chemical (Applied Biosystems) and automatically analysed by ABI-Prism 377 (Applied Biosystems), allowed identifying a consensus sequence of 1491pb (Genbank# AY078403), which included part of bovine NRAMP1 exon1 (80bp) and 1411 bp at 5'end of exon1. Bovine consensus sequence of codifying region showed 100% homology with sheep when analysed by BLASTS. In addition, the promoter region showed a high homology with sheep promoter. Nevertheless, all the bovine sequences showed an insert of 51 pb, which lacks in sheep. No significant differences have been recorded among the different breeds, apart a g-a SNP (single nucleotide polymorphism) at position 731 pb in two Friesian cattle in heterozygous status order. Nevertheless, this SNP it is not located within regions, which represent transcription factor anchor loci. Further analysis on a larger sample could show a higher polymorphism of transcription factor anchor loci that could be related to different intracellular antigens response. Research supported by Italian MURST 40%-Cofin. 1999. (Texte intégral)
| Main Authors: | , , , |
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| Format: | conference_item biblioteca |
| Language: | eng |
| Published: |
CIRAD
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| Subjects: | L10 - Génétique et amélioration des animaux, tuberculose, séquence nucléotidique, génotype, http://aims.fao.org/aos/agrovoc/c_7997, http://aims.fao.org/aos/agrovoc/c_27583, http://aims.fao.org/aos/agrovoc/c_3225, |
| Online Access: | http://agritrop.cirad.fr/512047/ |
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