Abstract
We have constructed two broad-host-range Gram+/Gram- probe vectors designed for the cloning of bacterial genetic expression and secretion signals. These vectors make use of a silent reporter gene encoding the mature alpha-amylase from Bacillus licheniformis whose reactivation can easily be monitored on iodine-stained starch plates. Shotgun cloning of Enterococcus faecalis DNA fragments allowed recovery of several cassettes directing transcription, translation of the reporter gene and secretion of alpha-amylase. Sequence analysis revealed, in each case, the presence of a putative promoter, ribosome-binding site and signal peptide similar to those described in other Gram+ bacteria.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Bacillus / enzymology
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Bacillus / genetics
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Base Sequence
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Chromosomes, Bacterial
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Cloning, Molecular / methods*
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Enterococcus faecalis / genetics*
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Escherichia coli / genetics
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Gene Expression Regulation, Bacterial*
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Genetic Vectors / genetics*
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Molecular Sequence Data
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Mutagenesis, Insertional / genetics
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Protein Biosynthesis
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Protein Conformation
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Protein Sorting Signals / genetics
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Recombinant Fusion Proteins / genetics
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Restriction Mapping
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Sequence Homology, Nucleic Acid
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Transcription, Genetic
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alpha-Amylases / genetics*
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alpha-Amylases / metabolism
Substances
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Protein Sorting Signals
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Recombinant Fusion Proteins
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alpha-Amylases
Associated data
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GENBANK/M74891
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GENBANK/M74892
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GENBANK/M77275
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GENBANK/M77276
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GENBANK/M77277
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GENBANK/M77278
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GENBANK/M77279
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GENBANK/M79309
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GENBANK/M79310
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GENBANK/M90357
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GENBANK/S43285
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GENBANK/S43286
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GENBANK/S43293
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GENBANK/S43295
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GENBANK/S43297