行政院國家科學委員會專題研究計畫期中進度報告:以標籤式突變法分析Salmonella choleraesuis活體內毒力因子(2/3)
Date Issued
2004
Date
2004
Author(s)
張照夫
DOI
922313B002136
Abstract
A signature-tagged mutagenesis (STM) system was adapted to identify the genes
required for in vivo survival of Salmonella enterica serovar Choleraesuis (Sal.
Choleraesuis) in pigs. An STM library was constructed in Sal Choleraesuis (kanamycin
sensitive) by conjugation with a suicide plasmid vector (pUTminiTn5Km2) which
contains unique DNA sequence tags and a kanamycin resistance marker. Sal
Choleraesuis (kanamycin-resistant) mutants were obtained by transposon mutation,
resulting in gene inactivation by insertion of the suicide vector carrying a unique tag.
960 mutants of Sal Choleraesuis were pooled together (48 mutants/pool) and screened
in a pig model. Out of 960 mutants, 34 of them were not recovered from the
mesenteric lymph node. These 34 mutants were considered as virulence-attenuated
mutants. In order to identify the virulence genes, the regions flanking the kanamycin
gene of the attenuated mutants were amplified by inverse PCR and sequenced. The
sequence analysis showed that the 34 mutants had insertions in 34 different genes that
could be grouped in eight classes: (i) genes encoding cell envelope; (ii) type III
secretion system; (iii) gene encoded in a virulence plasmid; (iv) genes involved in
transport; (v) genes coding for Gifsy-1 prophage protein; (vi) genes encoding proteins
involved in regulatory; (vii) genes involved in metabolic pathways; and (viii) DNA
sequences that showed similarity to hypothetical genes with unknown function.
Subjects
Samonella Choleraesuis
signature-tagged mutagenesis
virulence factor
in
vivo expression
vivo expression
mutant
SDGs
Publisher
臺北市：國立臺灣大學獸醫學系暨研究所
Type
report
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