Development Of Aptamer-based Rapid Detection Methods For Ochratoxin And Estrogen

Aptamer is a single-stranded DNA or RNA that can bind to a target with high affinity and high specificity.It has the advantages of small size,high stability,easy chemical synthesis and modification.And aptamer was combined with a variety of analytical techniques,widely used in food safety and environmental monitoring and analysis technology research.In this study,a lateral flow fluorescent strip based on aptamer for the specific detection of ochratoxin A was constructed.Based on the principle of fluorescence resonance energy transfer,a rapid and sensitive detection method for estradiol was established;A visual and rapid nanogold colorimetric detection method for progesterone was established based analysis and study of effect of the length of the aptamer sequence.The main research content is as follows:1.In this study,simple,specific,sensitive,and easily applied Cy5-labeled aptamer based lateral flow strip biosensors in the competitive format were developed for one-step detection of OTA in corn samples.The test strip showed an excellent linear relationship in the range from 1 ng mL^-1 to 1000 ng mL^-1 with the LOD of 0.40 ng mL^-11 and recoveries from 96.4%to 104.7%in spiked corn samples.Thus,the strip sensor developed in this study is an acceptable alternative for rapid detection of the OTA level in grain samples.2.A competitive fluorescent aptasensor was constructed based on fluorescence resonance energy transfer?FRET?for determination of 17?-estradiol?E2?.With a competitive"turn-on"aptasensor,E2could be quantitatively detected by monitoring fluorescence enhancement.Under optimized conditions,this assay exhibited a linear response to E2 within the range from 0.1 ng mL^-1 to 10?g mL^-1,with the detection limit of 0.1 ng mL^-1?0.35 nM?.This assay has been successfully applied for determination of E2 in environmental,biological and food samples with a low cost and simple pretreatment process,requiring no complicated operations.The results showed high selectivity and good recovery.The present strategy holds great potential in detection of E2 in complex matrixes and is also important in environmental monitoring,food safety and human health.3.A rapid colorimetric assay for progesterone based on different lengths of aptamers was established.The affinity of two aptamers and the sensitivity to nanogold colorimetry were studyed and analyzed.The results showed that the 25mer aptamer method was 5 times more sensitive than the 60mer aptamer.The60mer aptamer chain was too long,when the aptamer was combined with progesterone,and the redundant bases were still adsorbed on the surface of the nanogold particles.The two aptamer nanogold colorimetry methods showed a good linear relationship in each detection concentration range?R²>0.9?.The results of spiked recovery in river water samples were good?RSD<6.5%?.Although the sensitivity of this method needs to be improved,there is less research on the detection method of progesterone based on aptamers,and this study provides new ideas for further aptamer modification and the establishment of a more sensitive progesterone detection method.