Data/hora: 19/06/2024 23:29:00

Biblioteca(s): | Embrapa Gado de Corte.
Data corrente: | 22/03/2011
Data da última atualização: | 09/09/2014
Autoria: | ANDREOTTI, R.; SOARES, M.; CUNHA, R. C.; LEITE, F. P. L.
Afiliação: | RENATO ANDREOTTI E SILVA, CNPGC; UFPEL; UFPEL; UFPEL.
Título: | Heterologous expression in Pichia pastoris of gene BMTI optimized.
Ano de publicação: | 2010
Fonte/Imprenta: | In: INTERNATIONAL CONGRESS OF PARASITOLOGY, 12., 2010, Melbourne. Conference abstracts. Melbourne: The Australian Society for Parasitology:World Federation of Parasitologists, 2010.
Páginas: | p.149-152

Idioma:

Inglês
Português

Notas: | ICOPA XII

Conteúdo:

The cattle tick, Rhipicephalus microplus, is the main tick species that undermines productivity losses in livestock resulting in the production of milk and meat throughout Brazil. Its control is primarily chemical and faces problems due to the resistance of ticks to several active principles used. The immune control is a viable alternative to tick control, and the protein BMTI is a strong candidate as a vaccine antigen. The aim of this work was to synthesize the sequencing of BMTI (BmTIS), optimizing it to "codon usage" of Pichia pastoris in an attempt to improve its expression. This sequence was cloned in eukaryotic expression vector pPICZaA, respecting the make reading the AOX1 promoter, present in the plasmid and that is strongly induced by the presence of methanol, following the sign "a-factor for protein secretion, and the tail of poly-histidine. The resulting plasmid, pPICZaBmTIS, X33 was transformed into strains of P. pastoris. Clones were characterized by PCR using primers of the vector and also by dot blot analysis using anti-histidine. A clone (X33BmTIS17) was sequenced and selected to be induced. A single colony clone X33BmTIS17 was inoculated with 25mL of minimal medium with glycerol and incubated at 200rpm, 300C, overnight. The culture was centrifuged at 1500 g, 5min and resuspended in minimal medium with methanol. The supernatant was subjected to SDS-PAGE 0.7%, which showed a major band with the approximate size of 49 kDa. In Western bloting, a band at the same time was recognized by the anti-histidine and by bovine serum immunized with a synthetic based pepitídeo (BmTI). Thus, a recombinant protein (rBmTI) was
expressed, and was recognized by the anti-histidine and characterized as potential antigenic by bovine serum immunized with the synthetic pepitídeo.

Thesagro: | Carrapato; Sanidade Animal.
Thesaurus Nal: | Rhipicephalus microplus.

Categoria do assunto:

H Saúde e Patologia
L Ciência Animal e Produtos de Origem Animal

Registro original: | Embrapa Gado de Corte (CNPGC)

Exemplares:

CNPGC - (13798-1) - UPC - SP - PP -  -  - 004/001 - DIS
CNPGC - (13823-1) - UPC - SP - PP -  -  - 003/015 - DIS