Construction And Preliminary Application Of Recombinant Bacterias Of Luciferase Genes From Vibrio Qinghaiensis Sp.-Q67

This paper compare the expression and luminescnce of luxAB and luxCDABE genes from Vibrio qinghaiensis sp.—Q67 in Escherichia coli to make preparations for the construction of recombinant bacteria which could especially detect heavy metals.In order to obtain the homologous fragments of luxAB and luxCDABE,primers were designed.Then get the product through polymerase chain reaction(PCR)?As the next process,the product got before were cloned into vector pET-22 b and then transfered into Escherichia coli DH5? for genetic sequencing.The positive clones were propagated and the plasmids extracted were transfered into Escherichia coli BL21(DE3)? Performance of bioluminescence of recombinant Escherichia coli encoding thermostable luciferase from Vibrio qinghaiensis were detected.After about one hour culture with 1?L/mL decanal and 1?L/mL IPTG the luxAB and luxCDABE recombinant bacteria got the highest RLU: 750618.5and 140901.5 respectively.The luxAB recombinant bacteria got higher RLU which is different from the result of other researcher got ? Both of the recombinant bacteria did not emit light without decanal which shows that luxCDE genes did not expressed in the luxCDABE recombinant bacteria.This phenomenon could be ascribed to mismatching of luxCDE and the expression system or metabolism products of themselves.To verify the points the toxicity of heavy metals to recombinant bacterias was detected.EC50 of luxAB and luxCDABE recombinant bacteria of Cd2+?Hg2+?Cr6+are 5.160?5.129?7.097 mg/L and 6.137?7.643?7.736 mg/L respectively.The expression of luxAB is more effective than luxCDABE in E.Coli.Lux AB recombinant bacteria had lower EC50 values showing that the luxAB recombinant bacterias are more sensitive towards heavy metals.Finally this study successfully constructed recombinant bacterias of lux genes from Vibrio qinghaiensis sp.—Q67 and studied the application of the recombinant bacterias which would make preparations for the further study of lux genes.