Caracterização molecular do gene da β-tubulina em Phakopsora pachyrhizi / Molecular characterization of the β-tubulin gene in Phakopsora pachyrhizi

AUTOR(ES)
DATA DE PUBLICAÇÃO

2008

RESUMO

This study was carried out to characterize either the β- tubulin gene from the Phakopsora pachyrhizi and the protein codified by this gene. The urediniospores were collected in seven Brazilian counties. The genomic DNA was extracted and the β-tubulin gene was amplified by PCR technique through five combinations of specific primers on such a way to covering the whole region under study. Then, the obtained fragments were purified and sequenced. The sequences were aligned and compared among each others and with sequences obtained in the GenBank. The β-tubulin gene from P. pachyrhizi showed 2815 pairs of bases and was constituted by nine exons and eight introns. Its opened reading phase (ORF) showed 1341 base pairs. It was verified that 452 base pairs correspond to the promoter region, whereas 234 pairs belong to the untranslated extremity 3 . The promoter region showed the typical signs involved in the beginning of the transcription, as being three possible sequencies TATA box and five possible sequencies CAAT box. A domain of 30 nucleotides constituted by cytosine and thymine is inside the promotor region between the basis pairs - 205 and -175 upstream the codon beginning the translation. Some sites involved in the splicing mechanism are found, as flanking those eight introns, whereas the site GT(A/G)(N)GT was found at the extremity 5 of the intron and the site (C/T)AG at extremity 3 . The consensus sequence AATAAA that plays an important role in the polyadenilation process of the precursory mRNA was found after 44 pairs of bases downstream the TAG termination codon. The ORF found for this gene codifies a protein constituted by 446 amino acids, that shows high identity (above 89%) with β-tubulin in other fungus species related. When comparing the amino acid sequence predicted for β-tubulin, this protein was identified to have four conserved domains as follows: domain of linkage to nucleotides, domain of linkage to taxol, domain of alpha/beta interface and domain of beta/ alpha interface. The phylogenetic analyses based on alignment of the β-tubulin gene provided the allocation of P. pachyrhizi together with the other fungus causing the rusts. The comparison of the introns of the β-tubulin gene of P. pachyrhizi with introns of Melampsora lini and Uromyces fabae, as the two last ones being the only rustcausing fungus species with the β-tubulin gene completely sequenced until the moment, showed that both number and position of the introns were conserved. However, the sequences corresponding to each intron showed low identity (around 30%).

ASSUNTO(S)

phakopsora pachyrhizi β caracterização molecular β -tubulin phakopsora pachyrhizi molecular characterization -tubulina biologia molecular

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