Con Sectruction Of Genetically Engineered Bacteria For Detection And Degradation Of Formaldehyde

As a native toxin, formaldehyde has strong toxic effect on the organism and is one of the main pollutants of indoor air pollution. How to control formaldehyde pollution in the air has caused great concern, and degradation of formaldehyde by microorganism will be an effective way for the removal of formaldehyde.This paper synthesized a of microcystin degrading enzyme gene which optimizated by Lactococcus lactis bias codon, constructed a FADH gene engineering bacteria Lactococcus lactis L6 to expression. In order to solve the increasingly severe pollution of the lake reservoir water body and provide a solution for the safety of drinking water.The FADH gene from Sphingomonas sp. ACM-3962 was composed of 336 amino acid residues. The Lactococcus lactis encoding codons preference GC content lower,according to the 2504 code region and 696252 codons from Lactococcus lactis SK11,choose the abundant and preferred codons to optimize. Total 246 codons was replaced.After codon optimization, the FADH expression in Lactococcus lactis increased37.14%.Sphingomonas ACM-3962 HOCO degradation gene cluster, between the FADHC and FADH gene, there is a positive promoter and a reverse promoter, respectively, to control the expression of a FADHnd FADHC. The fragment was cloned and the green fluorescent protein and the red fluorescent protein were added to the E.coli to detect and verify its function. The results show that the promoter of promoter FADH can start the promoter of A FADH gene expression, and it can be expressed in the downstream proteins of LR microcystin..Insert the promoter-GFP FADH gene and the FADH A gene into the pMG36 e vector,and construct the recombinant vector p MG-FADH. After amplification and verification, the recombinant vector p MG- FADH was transformed into E.coli BL21 and L.lactis ML23 through electric shock method, and the gene engineering bacteria L.lactis L6 was constructed to express FADH.The culture medium was placed in the darkroom of UV light irradiation, to observe the color of bacteria liquid, visible bacteria liquid issued a green fluorescence, show that in the presence of HOCO and FADH promoter can start GFP-FADH fusion expression, the bacterium liquid in L.lactis ML23 emits green fluorescence under UVlight. The results showed that the positive promoter of promoter GFP-FADHnd the expression of A FADH fusion protein could also be initiated in Lactobacillus..Adding the freeze-dried FADH powder to the HOCO solution, HOCO can be hydrolyzed to linear molecule in 3h by more than 97%, and the toxicity of HOCO is greatly reduced.Adding the lyophilized Lactococcus lactis powder to the HOCO solution, HOCO can be hydrolyzed to linear molecule in 3h by more than 99%. Acoording to the unlyophilized Lactococcus lactis, the enzyme activity of FADH was lossed 30%, The death number of the cell is about 48%, so the directly use of freeze-dried Lactococcus lactis has good application prospects.Using the freeze-dried Lactococcus lactis to degradate HOCO in natural water,compared with laboratory buffer conditions, the enzyme activity was greatly reduced.This is because of the lacking of nutrients in natural water and p H is demanding. It also shows that, HOCO degradation in natural water by Lactococcus lactis lyophilized,also need to be studied.In the air containing 100 ppm formaldehyde is induced by Lactococcus lactis in the green fluorescent protein expression. The fluorescence intensity and the amount of bacteria showed a linear relationship. When the air contains 20 ppm formaldehyde,green fluorescent protein began to express, 60 ppm above, the green fluorescence intensity can be quantitatively detected. The filter paper is used to detect the biological sensor, which can respond to the formaldehyde in the environment, and the obvious green fluorescence is observed under the ultraviolet lamp. Construction of Lactococcus lactis gene engineering bacteria can response to the formaldehyde in air environment, in order to formaldehyde in air testing, can be used as the detection of formaldehyde in air of biosensors.