Experimental Studies Of The Sandwich-type Electrochemical Enzymes-based Lna-modified DNA Biosensor For Detection Of Relative Fusion Genes In Acute Promyelocytic Leukemia

As a novel biosensor, electrochemical DNA biosensor has been widely notified for fast detection, sensitivity, minitype and easy operation. Electrochemical DNA biosensor considered as an interdisciplinary study about electrochemistry and molecular biology has far-reaching significance and applicable value in the study of genetic project and clinical medicine. Recently, locked nucleic acid technology evolves with the development of genetic engineering technology to become a kind of new biological high technology. As it has advantages of high selectivity and affinity to DNA and RNA, good thermostability, nucleic acid technology has been applied in gene research, clinical diagnosis and treatment.In the paper, a novel DNA electrochemical probe (locked nucleic acid, LNA) was first designed and involved in building a sandwich-type electrochemical DNA biosensor for detection of PML/RARαfusion gene in acute promyelocytic leukemia (APL). The stability and the specificity of the DNA biosensor were also been investigated.Firstly, the sandwich-type electrochemical enzymes-based LNA-modified DNA biosensor, which had combinated locked nucleic acid (LNA) probe and electrochemical enzyme immunoassay, was developed for PML/RARαfusion gene in APL in this research. Under the optimal condition of the hybridization, while detecting the complementary DNA standard concentration range of synthetic PML/RARαfusion gene from 1.0×10 -13 to 1.0×10–11 mol/L, there was a good linear relationship between the current signal and the concentration of complementary DNA, the lower limit was 7.4×10 -14 mol/L. Otherwise, the biosensor shows a good specificity to distinguish the complementary sequence and different mismatch sequences. The obvious difference of results after hybridization demonstrated this biosensor could detect synthetic PML/RARαfusion gene qualitatively and quantitatively in simple system.Secondly, the specificity and the sensitivity of the DNA biosensor were investigated further under the mixed hybridization solution contained different kinds of mismatch sequences as interference background. Under the optimal condition of the hybridization, while detecting the complementary DNA in mixed system standard concentration range of synthetic PML/RARαfusion gene from 5.0×10 -9 to 5.0×10 -11 mol/L, there was a good linear relationship between the current signal and the concentration of complementary DNA, the lower limit was 9.6×10 -12 mol/L. It can be seen that under a certain interference background, the electrochemical DNA biosensor still possesses good molecular recognizability and can hybrid with completely complementary sequences well. It provided a good basis of further detection research in practical samples.