Fluorescence Detection Of Sheep Disease Markers In Body Fluids Using Functional Microporous MOF

Animal husbandry is an important industry in the national economy.How to improve the survival rate and reproduction rate of animal husbandry is the key concern of the entire animal husbandry industry.Among them,mycoplasma ovipneumoniae（MO）is one of the major infectious diseases causing serious losses to the sheep breeding industry.This disease has a high infectious rate and can cause stunting or even death in sheep.Due to the unsatisfactory effect of vaccines,early diagnosis is still the most effective method to cut off the chain of transmission.However,the traditional method has poor economic efficiency and complicated operation,and a new detection method for the detection of mycoplasma ovipneumoniae needs to be proposed.In addition,it is difficult to improve the reproductive ability of sheep by traditional breeding methods,and the purpose of improving breeding can be achieved by detecting the sheep multiple birth gene（Fec B）.In this paper,two fluorescent biosensors were constructed,which were applied to the detection of MO and Fec B gene,respectively.1)A novel alternative nanoprobe for MO was prepared using porous metal–organic frameworks（MOF）as the scaffold and hairpin DNA with specific sequences of MO and G4 segment as the probe.The preparation was based on the strong fluorescence emission by Th T in the limitation of G-quadruplex（G4）with a cavity structure.The use of MOF effectively limits the folding behavior of G4 as the part of the probe and improves the defect of the strong background signal caused by the free-state G4probe in a buffer.The results from the selectivity experiment showed that only a trace of the target with lowerΔG could be the“key”to the highly efficient triggering of the release behavior of the G4probe from MOF and then change the fluorescence behavior of Th T.The DNA targets could be determined by observing the change in the signal.More importantly,the probe showed a low detection limit and a good linear correlation between the concentration of target DNA ranging from 1×10^-10 M to 1×10^-6 M not only in buffer but also in natural complex media.Besides,the operation involved in the whole strategy was simple,and the total cost was low.All these findings implied the value of the probe in further clinical diagnosis.This study was novel in reporting the successful construction of theΔG-sensitive sensor for MO.2)A MOF-based fluorescent biosensor was constructed to detect Fec B gene.The MOF with biological enzymatic activity is shielded by poly A and poly C.When the Fec B gene with GT single base mismatch exists,due to the pairing of the mismatched base and poly A/poly C,poly A and poly C are induced from the MOF surface.And the surface of exposed MOF catalyzed the substrate o-phenylenediamine（OPD）to2,3-diaminophenazine（DAP）to produce a strong fluorescence signal.The results show that the biosensor has excellent selectivity for GT mismatch,and can distinguish GT mismatch between TT mismatch and AT mismatch.Linearity ranges from 1×10^-9 M to 1×10^-5 M in buffer and 5%serum,with a detection limit as low as 0.247 n M in serum.Moreover,the biosensor has good stability,and the fluctuation of Fec B detection signal within one week of preparation is less than 5%,showing good application potential.