Iron is one of the essential nutrient elements and exists in many forms in aquatic environments, and only free ferric ion can be utilized by phytoplankton. Physicochemical analysis can provide information on total iron content in natural water samples, but bioassay approach which uses living organisms as bioreporters can tell us the bioavailable fraction of dissolved iron. Anabaena sp. PCC 7120 is a filamentous and nitrogen-fixing cyanobacterium. Alr2581 is an outer membrane transporter associated with Fe uptake in Anabaena sp. PCC 7120 and its expression is enhanced under iron-limiting conditions. In this study, an Anabaena sp. PCC 7120 Fe-responsive bioreporter strain Palr2581-luxAB was constructed by fusing the promoter of gene o/r2581 to luxAB reporter gene. Our results showed that growths were no difference in the bioreporter Palr2581-luxAB grown in Fe-replete and Fe-deplete BG11 medium. However, contents of chlorophyll a of cells cultured in Fe-replete medium were much higher than those in Fe-deplete medium. LuxAB activities were significantly induced in Fe-depleted growth medium in each growth phase, indicating continuous transcription from the alr258l promoter. A quantitative assessment of bioluminescent was made by incubating cells for 0-48h in trace metal-buffered Fraquil medium containing various additions of Fe. An ideal dose-response curve was generated relating bioreporter luminescent to the pFe (pFe=lg [Fe3+ free ferric]) between 19.6 and 21.7 after 12h incubation time and was best represented by sigmoidal curve (y=41852.4-31891.8/{l+exp[(x-20.69)/0.16]}, R2=0.9875) having a linear component extending between pFe 20.7 (Fe3+=10-20-7 mol/L) and pFe 21.2 (Fe3+=10-21.2 mol/L) (y=35472x-707426, R2=0.9988). In addition, the applicability of using this Fe bioreporter to assess Fe availability in the natural environment has been tested by using water sample from Lake Donghu (contained pFe 20.56, Fe3+=10-20.56 mol/L). It indicated that the bioreporter could serve as a quantitative tool to assess Fe-nutrient status in natural freshwater environments.%鱼腥藻PCC 7120中的alr2581基因编码的蛋白质在缺铁胁迫时显著上调.将该基因的启动子Palr2581和费氏弧菌的luxAB基因融合,通过同源单交换,整合到鱼腥藻PCC 7120的基因组上,构建了可以感知环境中铁的生物报告体Palr2581-luxAB.该藻株在不含铁的BG11中培养时,启动子Palr2581的转录活性增强,LuxAB 酶活显著升高.通过测定Palr2581-luxAB藻株在不同铁浓度Fraquil培养基中的LuxAB酶活,得到了铁浓度pFe(-lg[Fe3+])与LuxAB酶活的剂量反应曲线.结果显示,12h时,LuxAB酶活随培养基中Fe3+浓度增加呈S形递减关系,其中在pFe-20.7-21.2范围内有很好的线性关系.根据这一特性,我们利用Palr2581-luxAB作为铁的生物报告体,测定了武汉市东湖水体中可利用的铁浓度为10-20.56 mol/L.研究显示,通过这一方法可以较方便地监测各种淡水中可利用的铁浓度.
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