Constructing A Versatile And Ultrasensitive Micro Rna Detection Sensor Based On Hierarchical Gold Nanostructures Synergetic Base-Stacking Effect

Single-stranded non-coding endogenous small micro RNA(mi RNA)plays an important role in a variety of biological processes.It affects cell-to-cell communication,post-transcriptional gene regulation,cell fate and other biological processes,and its abnormal expression can cause some diseases,such as: tumors.mi RNA plays a regulatory role in the development,metastasis and invasion of tumors.mi RNA plays the role of proto-oncogene or tumor suppressor gene.Its expression affects the effect of patient treatment and survival,and is a potential cancer diagnosis and prognosis biomarkers.Recently,researchers have discovered that mi RNAs help tumor immune escape through a variety of ways such as exosomes,inhibit tumor cell apoptosis,and inhibit normal immune responses.Studies have shown that certain mi RNAs can increase the sensitivity of radiotherapy and chemotherapy,and there is a certain relationship between the abnormal expression of mi RNAs and cancer resistance.mi RNA affects the occurrence and treatment of diseases,and its detection is of great significance.However,the mi RNA sequence is short(18-24 nt),high homogeneity,and low abundance,which requires sensitive and specific detection methods.Biosensors have the advantages of high sensitivity,specificity,and fast detection speed,which play an increasingly important role in the medical field.One can fix the probe with fully complementary mi RNAs on the sensing interface to capture the target,and then detect mi RNAs by adding signal molecules or using ultra-sensitive transducers.Due to the special structural characteristics of PNA,it has high sensitivity and specificity when used for mi RNA detection.Therefore,PNA probes are often introduced in the above design,but it is difficult to design completely complementary capture probes because most mi RNAs have secondary structures.This limitation limits the wide use of such methods.To avoid the influence of secondary structure,we divide the nucleic acid into fragments.Two or more juxtaposed terminal nucleotides hybridized to a longer complementary signal-stranded nucleic acid is called base stacking hybridization.Affected by dispersion attraction,short-range exchange repulsion and electrostatic interaction,this hybridization method can enhanced stability and efficiency for nucleic acid hybridization.After selecting the appropriate length of nucleic acid,we found that the efficiency of base stacking hybridization can be nearly complete complementary hybridization.The performance of a biosensor to detect molecules is closely related to its interface.The structure of the interface has an important impact on the thermodynamics and kinetics of the assembly,binding and signal transduction of biomolecules.The design and structural regulation of the biorecognition interface is of significance in the development of high-performance biosensors.The nanostructured interface is favored by scientific researchers due to its large specific surface area and high electron transfer efficiency.The hierarchical gold nanostructures interface obtained by electrochemical deposition has a large specific surface area and molecular probe deflection angle,which provides favorable dynamic hybridization conditions for nucleic acid molecular hybridizationHere,we present a base-stacking effect-mediated ultrasensitive electrochemical mi RNA sensor(BSee-mi R)with a universal sandwich configuration.In the BSee-mi R,a short capture probe(10 nt HS-DNA)selfassembled on a gold electrode surface could effectively capture the target mi RNA synergizing with another sequence based on coaxial base-stacking,which rivals the fully complementary strength.Importantly,such a sandwich structure is flexible to incorporate signal amplification strategies(e.g.,HRPTMB)that are usually difficult to achieve in short sequence detection.Using this design,the BSee-mi R achieves a broad dynamic range with a detection limit down to 7.5 f M.Furthermore,we found a hierarchical gold nanostructures synergetic base-stacking effect that could improve the sensitivity of the BSeemi R by two orders of magnitude(79.3 a M).Our BSee-mi R also has a singlebase resolution to discriminate the highly homologous mi RNAs.More importantly,this approach is universal and has been used to probe target mi RNAs varying in sequences and secondary structures.Our ultrasensitive sensor could detect mi RNA in cell lysates and human blood and distinguish cancer patients from normal people.It is expected to become a universal tool to measure relevant mi RNA for tumor diagnosis.