Synthesis Of Nano-bioprobe And Its Applications In DNA Damage Detection

DNA is the most important genetic substance in the life activity,and DNA damages including DNA oxidization and base modification often happen in the body.DNA methylation,as one kind of DNA base modification,doesn't change DNA sequences,but it may alter the expression of genetic gene expression leading to abnormal gene transcription and translation and influencing the storage of genetic information.DNA methylation is the earliest,most fundamental,and also the most frequently studied epigenetic mechanism.The normal cell function depends on the maintainence of the epigenetic balance,however a lot of researches show that the aberrant DNA methylation could alter chromatin structure,DNA conformation,DNA stability and the interaction between DNA and protein.These disturbances of epigenetic stability are closely related with the pathogenesis of many other critical diseases.Especially for carcinogenesis,the typical characters are the hypomethylation of the whole genome and the hypermethylation of some parts.The abnormal DNA methylation can be founded in the early period of cancer,and it will beome more serious for the gradual accumulation of various genetic alterations and the disruption of epigenetic modifications during the cancer progress.Hypermethylation reflects the abnormal DNA methylation in the promoter region of tumor suppressor gene.However both the reduction of DNA methylation in the promoter region of tumor suppressor gene and repeated sequences,and genetic instability can lead to the hypomethylation directly.Consequently,the development of highly sensitive,specific,and low-cost methods for the DNA methylation and methyltransferase assay still has a great impact on early disease diagnose and cure,and saving patient life.On the other hand,when the toxic and harmful exogenous compounds invade into the human body,they can cause direct or indirect DNA damages.Take organophosphorous(OPs)compounds,which are widely used in agriculture field,as an example.OPs are chemically unstable and easy to decompose in nature and in living organisms,and they can be left over in foods,especially in fruits and vegetables with high residual amount and long residual time.OPs exert many effects of genetic toxicity and cytotoxicity on many other tissues and organs through food chain,so that many countries all over the world have pay much more attentions to establish more stringent standards on detecting OPs remains.Therefore,the further research and development of highly sensitive detection of organophosphorus pesticide residues can not only strictly regulate and supervise the use of pesticides,but also can promote the improvement and innovation of pesticide manufacture.For these purposes,electroanalytical chemistry and nanomaterials are introduced into constructing biosensors for detecting DNA methylation,methyltransferase activity and OPs concentration.Details are described as below:(1)The specific probe can be constructed by G-rich DNA,capture DNA and target DNA,then G-rich DNA can form HRP-mimicking DNAzyme to catalyze poly-aniline deposition on the electrode so that the strong electrochemical signal can be acquired.Nucleicacidase,including restriction endonuclease Hpa? and exonuclease ?,however,can recognize and cleave the sequence in the probe,resulting in no HRP-mimicking DNAzyme formation,and electrochemical responds disappearing.When the site of CpG being mathylated,the cleavage will be blocked and the electrochemical current appears again.This detection limit of the biosensor is 0.12 U mL-1.(2)A mimic-hybridization chain reaction(mimic-HCR)amplified strategy is proposed for sensitively detecting the DNA methylation and methyltransferase activity.In the presence of methylated DNA,DNA-gold nanoparticles are captured on the electrode by sandwich-type assembly.Then mimic-HCR of two hairpin probes will be triggered by probe DNA on the Au-NPs to produce many long double-helix chains for numerous RuHex inserting.As a result,the electrochemical signal can be amplified.If no DNA is methylated,however,the specific dsDNA on the Au electrode can be cleaved by the specific nucleicacidase,resulting in the signal decrement.The thus designed sensor owes much lower detect limit of 0.03 U mL-1 due to the synergy between Au-NPs and mimic-HCR.(3)Graphene has abundant hydrophilic groups on its surface and it has a unique 2D structure.The new bionanosheet is formed by graphene and chitosan interaction for covalent binding with acetylcholinesterase on the GCE surface.This biosensor is sensitive to detect organophosphate pesticides for OPs have inhibitory effect on the activity of acetyl-cholinesterase.The results show that this biosensor has a good linear relationship ranging from 1×10-2?g L-1 to 5×102?g L-1 and the detection limit is 8.2×10-3?g L-1 moreover,it is accuracy,sensitivity,low cost and easy to operate.