Use of RNA Arbitrarily Primed-PCR Fingerprinting To Identify Vibrio cholerae Genes Differentially Expressed in the Host following Infection
IR@IICB: CSIR-Indian Institute of Chemical Biology, Kolkata
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| Title |
Use of RNA Arbitrarily Primed-PCR Fingerprinting To Identify Vibrio cholerae Genes Differentially Expressed in the Host following Infection
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| Creator |
Chakrabortty, Amit
Das, Soumita Majumder, Sabita Mukhopadhyay, Kanchan Roychoudhury, Susanta Chaudhuri, Keya |
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| Subject |
Infectious Diseases and Immunology
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| Description |
Evidence suggests that a repertoire of Vibrio cholerae genes are differentially expressed in vivo, and regulation of
virulence factors in vivo may follow a different pathway. Our work was aimed at characterization of in
vivo-grown bacteria and identification of genes that are differentially expressed following infection by RNA
arbitrarily primed (RAP)-PCR fingerprinting. The ligated rabbit ileal loop model was used. The motility of in
vivo-grown bacteria increased by 350% over that of in vitro-grown bacteria. Also, the in vivo-grown cells were
more resistant to killing by human serum. By using the RAP-PCR strategy, five differentially expressed
transcripts were identified. Two in vitro-induced transcripts encoded polypeptides for the leucine tRNA
synthatase and the 50S ribosomal protein, and the three in vivo-induced transcripts encoded the SucA and
MurE proteins and a polypeptide of unknown function. MurE is a protein involved in the peptidoglycan
biosynthetic pathway. The lytic profiles of in vivo- and in vitro-grown cells suspended in distilled water were
compared; the former was found to be slightly less sensitive to lysis. Ultrathin sections of both cells observed
under the transmission electron microscope revealed that in contrast to the usual wavy discontinuous membrane
structure of the in vitro-grown cells, in vivo-grown cells had a more rigid, clearly visible double-layered
structure. The V. cholerae murE gene was cloned and sequenced. The sequence contained an open reading frame
of 1,488 nucleotides with its own ribosome-binding site. A plasmid containing the murE gene of V. cholerae was
transformed into V. cholerae 569B, and a transformed strain, 569BME, containing the plasmid was obtained.
Ultrathin sections of 569BME viewed under a transmission electron microscope revealed a slightly more rigid
cell wall than that of wild-type 569B. When V. cholerae 569B and 569BME cells were injected separately into
ligated rabbit ileal loops, the transformed cells had a preference for growth in the ileal loops versus laboratory
conditions.
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| Publisher |
American Society for Microbiology
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| Date |
2000
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| Type |
Article
PeerReviewed |
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| Format |
application/pdf
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| Identifier |
http://www.eprints.iicb.res.in/848/1/INFECTION_AND_IMMUNITY_68(_7)_3878%2D3887;2000[31].pdf
Chakrabortty, Amit and Das, Soumita and Majumder, Sabita and Mukhopadhyay, Kanchan and Roychoudhury, Susanta and Chaudhuri, Keya (2000) Use of RNA Arbitrarily Primed-PCR Fingerprinting To Identify Vibrio cholerae Genes Differentially Expressed in the Host following Infection. Infection and Immunity, 68 (7). pp. 3878-3887. |
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| Relation |
http://dx.doi.org/10.1128/IAI.68.7.3878-3887.2000
http://www.eprints.iicb.res.in/848/ |
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