Use of lac fusion to approach the high TyrB production with a runaway-replication vector

An easy and efficient method was proposed to achieve high production of the tyrB gene product on a runaway-replication vector. The DNA containing the tyrB promoter fused transcriptionally with the lacZ gene was cloned into a runaway-replication plasmid, pRA90. With the aid of this gene fusion, the e xpression level of tyrB in response to temperature alterations, the inducti on cell density, the temperature upshift rate, and various culture medium w as investigated by measuring LacZ activities. The optimal condition thus se t up for LacZ production was adopted to yield TyrB by a similar clone harbo ring an intact tyrB gene. As a result, TyrB was over-amplified 135-fold in terms of specific activity in comparison with the wild-type level. This wor k demonstrates the potential use of lacZ fusion to probe the sound conditio n for high TryB production with uncontrolled-replication plasmids.