The genomic library of alkalophilic thermophilic Bacillus was constructed in E.coli. A single recombinant [pATB 507 I] out of 10,000 colonies showed growth at 50 degrees C, which produced large, opaque and mucilaginous colonies on LB. The IMViC biochemical tests suggested that the strain was E.coli, and scanning electron microscopy revealed increased cell length. The plasmid DNA from the thermophilic recombinant showed multiple bands. The cured strain showed ampicillin resistance (10 micrograms/ml) and presence of insert DNA in the genome, as revealed by southern blot analysis. The thermophilic trait was transformable with 10(-4) frequency of expression amongst the pATB 507I transformants. The hybridization of E.coli genomic DNA with labeled genomic DNA from alkalophilic thermophilic Bacillus suggested that the genetic recombination(s) of the insert DNA was an essential prerequisite for expression of thermophilic trait. The conjugative plasmid (RP4) mediated mobilization of thermophilic trait resulted in thermophilic conjugants after 2 h incubation. Southern blot analysis revealed that a single locus [5.6 kb Bam HI fragment] was involved in the transfer of thermophilic trait.
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