| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 1999: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
Various environmental stresses are present in the nature. The common stresses are physicochemical stresses such as temperature, osmotic pressure, radiation, pressure, gas composition, contamination of toxic chemicals/metals, etc. Microorganisms quickly respond to those environmental stresses at genetic level for their survival or homeostasis. The present study attempted to screen gene elements resulting gene expression caused by response to various stress conditions.
At first, we attempted to construct a genome library of marine cyanobacterium, Synechococcus sp., using a marine cyanobacterium as a host organism. However, it was unexpectedly unsuitable to construct the library, since the transformation efficiency cyanobacteria was low. E.coli cells were, then, employed as the host organism for the construction of total genome library of the Synechococcus strain. Before the transformation of E.coli, a plasmid vector baring replication region for cyanobacteria was constructed. The lauxAB as a reporter gene was also inserted in the vector at the down stream of cloning site. Gene fragments between 0.5-2.0 kbp in length were used for cloning. Genome library with 2000 clones was constructed using E.coli as a host. Vectors were purified from 84 clones selected randomly. Size of inserted fragments was analyzed by the agarose gel eloctrophoresis. Only 25% of clones retained gene fragments between 0.5-2.0 kbp in length and the others retained smaller fragments. Luminescences due to the expression of LauxAB were observed from 20% of the clones, indicating that cloned fragments in those clones contain promoter region(s). These results indicated that the constructed vector system must be efficient for screening of stress response elements.
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