2 resultados para Northern blotting

em BORIS: Bern Open Repository and Information System - Berna - Suiça


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The human gene deleted in malignant brain tumors 1 (DMBT1) is considered to play a role in tumorigenesis and pathogen defense. It encodes a protein with multiple scavenger receptor cysteine-rich (SRCR) domains, which are involved in recognition and binding of a broad spectrum of bacterial pathogens. The SRCR domains are encoded by highly homologous repetitive exons, whose number in humans may vary from 8 to 13 due to genetic polymorphism. Here, we characterized the porcine DMBT1 gene on the mRNA and genomic level. We assembled a 4.5 kb porcine DMBT1 cDNA sequence from RT-PCR amplified seminal vesicle RNA. The porcine DMBT1 cDNA contains an open reading frame of 4050 nt. The transcript gives rise to a putative polypeptide of 1349 amino acids with a calculated mass of 147.9 kDa. Compared to human DMBT1, it contains only four N-terminal SRCR domains. Northern blotting revealed transcripts of approximately 4.7 kb in size in the tissues analyzed. Analysis of ESTs suggested the existence of secreted and transmembrane variants. The porcine DMBT1 gene spans about 54 kb on chromosome 14q28-q29. In contrast to the characterized cDNA, the genomic BAC clone only contained 3 exons coding for N-terminal SRCR domains. In different mammalian DMBT1 orthologs large interspecific differences in the number of SRCR exons and utilization of the transmembrane exon exist. Our data suggest that the porcine DMBT1 gene may share with the human DMBT1 gene additional intraspecific variations in the number of SRCR-coding exons.

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The epidemiology, phylogeny, and biology of nonencapsulated Streptococcus pneumoniae are largely unknown. Increased colonization capacity and transformability are, however, intriguing features of these pneumococci and play an important role. Twenty-seven nonencapsulated pneumococci were identified in a nationwide collection of 1,980 nasopharyngeal samples and 215 blood samples obtained between 1998 and 2002. On the basis of multilocus sequence typing and capsule region analysis we divided the nonencapsulated pneumococci into two groups. Group I was closely related to encapsulated strains. Group II had a clonal population structure, including two geographically widespread clones able to cause epidemic conjunctivitis and invasive diseases. Group II strains also carried a 1,959-bp homologue of aliB (aliB-like ORF 2) in the capsule region, which was highly homologous to a sequence in the capsule region of Streptococcus mitis. In addition, strains of the two major clones in group II had an additional sequence, aliB-like ORF 1 (1,968 to 2,004 bp), upstream of aliB-like ORF 2. Expression of aliB-like ORF 1 was detected by reverse transcription-PCR, and the corresponding RNA was visualized by Northern blotting. A gene fragment homologous to capN of serotypes 33 and 37 suggests that group II strains were derived from encapsulated pneumococci some time ago. Therefore, loss of capsule expression in vivo was found to be associated with the importation of one or two aliB homologues in some nonencapsulated pneumococci.