Regulation of the htpX gene of Xylella fastidiosa and its expression in E. coli.
Xylella fastidiosa was the first phytopathogen to be completely sequenced, and its genome revealed several interesting features to be used in functional studies. In the present work, the htpX gene, which encodes a protein involved in the heat shock response in other bacteria, was analyzed by RT-PCR by using cells derived from different cultural conditions. This gene was induced after a temperature upshift to 37 degrees C after growth in minimal medium, XDM, but showed constitutive expression in rich medium or in XDM plus plant extracts. Sequences upstream to the htpX gene, containing a putative regulatory region, were also transferred to E. coli, and the thermoregulation was maintained in the new host, since it was constitutively transcribed at 37 degrees C or 45 degrees C in all culture media tested, but not at 28 degrees C in minimal culture medium. The gene was also cloned into the expression vector pET32Xa/LIC, and the expression of the corresponding protein was confirmed by Western blotting.
Coltri
Patricia P
PP
Centro de Biologia Molecular e Engenharia Genética (CBMEG), Universidade Estadual de Campinas, P.O. Box 6010, CEP 13083-875, Barão Geraldo, Campinas, SP, Brazil.
Rosato
Yoko B
YB
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D018869:Heat-Shock Response / Q000235:genetics; D012329:RNA, Bacterial / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D011994:Recombinant Proteins / Q000032:analysis; D012045:Regulatory Sequences, Nucleic Acid; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044167:Xylella / Q000235:genetics* / Q000254:growth & development
D001426:Bacterial Proteins; D003470:Culture Media; D006360:Heat-Shock Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D011994:Recombinant Proteins