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EcoRI fragments of DNA from Bacillus subtilis NCIB 8565, a high producer of an endo-1,3-1,4-β-d-glucanase, were ‘shot-gun’ cloned in the plasmid vector pBR325. A 3·5 kb insert, carrying single restriction sites for AvaI, BglII, ClaI, PvuI and PvuII, was shown to direct the synthesis of β-glucanase in Escherichia coli K12. Enzyme activity was demonstrated in extracellular fractions of E. coli harbouring the β-glucanase gene; however, the largest proportion (< 50%) of total enzyme activity was periplasmic in location. β-Glucanase activity and cellular location were independent of the orientation of the 3·5 kb fragment in pBR325.
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