Biomonitoring Of Heavy Metal By Recombinant Luminescent Bacteria

To meet the demands of biomonitoring of heavy metal pollution in water environment,automatic on-linetoxicity test based on recombinant luminescent bacteria(RLB)is urgently needed.RLB can transform bioavailable heavy metal into easy-measurable and quantizable luminescence signal,hence,it can be used to evaluate the biotoxicity of heavy metal pollutions.Five RLBs with defferent propertieswere constructed.The heavy metal sensing components of cad,mer,pbr,cue and copwereamplified from Klebsiella pneumoniae,Pseudomonas putida SP1,Pseudomonas sp.ND6,Rahnella aquatilis HX2 and Klebsiella oxytoca M5 al by PCR method,and then subcloned into p UCD615 with luminescent gene lux CDABE,be aimed at controlling the expression of lux gene.The recombinant p UCD615 wastransferred into E.coli competent cell,thus we have constructed successfully five RLB strains namedp THE14-E.Coli,p THE30-E.Coli,p THE57-E.Coli,p THE10-E.Coli and p THE11-E.Coli respectively,which show response to Hg,Cd,Pb,Cu,or Ag.Internal standard method wasestablished to avoid false negative in RLB test.In this method,4 values are required for toxicity calculation and concentration range determination: RLUN,luminescence of background;RLUP,luminescence of positive sample;RLUS,luminescence of water sample;RLUI,luminescence of water sample that contains positive substance.An automatic on-line biosensor based on RLB method wasconstructed.In this biosensor,a mechanical arm and relevant instruments are controlled by an upper computer to automatically carry out all testing procedures of RLB test,including cell suspension preparation,water sample preparation,mixed reaction,toxicity test,and system cleaning.A two-dimension toxicity biosensor wasalso constructed to monitor acute toxicity and toxicity towardsheavy metalsimultaneously.whichuses natural luminescent bacteria and recombinant luminescent bacteria(RLB)as recognition elements towards differing comprehensive toxicity and heavy metal toxicity.thesoftware control system and hardware equipments are optimized to make it more easily control.It possesses promisingand applicative in the matter of on-line monitoring acute toxicity and heavy metals toxicity of in environmental water.