An electrochemical biosensor for double-stranded Wnt7B gene detection based on enzymatic isothermal amplification.
Wnt7B gene plays an important role in the development and progression of breast cancer, gastric cancer, esophageal cancer and pancreatic cancer. While, the natural state of DNA is double stranded, which makes it difficult to be directly detected. Here, we develop an electrochemical biosensor method for Wnt7B gene detection without the need to denature the target. This method firstly used nicking enzyme for exploiting in the double-stranded DNA (dsDNA). Then, long single-stranded DNA (ssDNA) was generated from the cutting site through polymerase extension reaction. Whereafter, the long ssDNA triggered a hairpin self-assembly recycling reaction, which gave rise to another isothermal amplification reaction. Last, short ssDNA was formed after the this amplification process, which could hybridize with the capture probe immobilized on Au electrode and result in signal variation. This method showed excellent analytical performance for dsDNA, of which the linear range was 2fM to 500pM and the detection limit was 1.6fM (S/N=3). It also showed an good results when applied to the real sample of Wnt7B gene detection.
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Junlong
J
Li
Department of Clinical Laboratory, Chongqing Municipal Jiulongpo District People's Hospital, Chongqing 400050, PR China.
Zhongping
Z
Chen
Department of Chemistry, Tsinghua University, Beijing 100084, PR China.
Yu
Y
Xiang
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Lili
L
Zhou
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Ting
T
Wang
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Zhang
Z
Zhang
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Kexin
K
Sun
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Dan
D
Yin
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China.
Yi
Y
Li
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, PR China. Electronic address: guomingxie@cqmu.edu.cn.
Guoming
G
Xie
D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D015374:Biosensing Techniques / Q000295:instrumentation; D003217:Conductometry / Q000295:instrumentation*; D015342:DNA Probes / Q000235:genetics; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006801:Humans; D009374:Neoplasms, Experimental / Q000235:genetics / Q000378:metabolism*; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000295:instrumentation*; D051153:Wnt Proteins / Q000032:analysis* / Q000235:genetics*
D014408:Biomarkers, Tumor; D015342:DNA Probes; C490675:WNT7B protein, human; D051153:Wnt Proteins
Double-stranded DNA detection; Electrochemical biosensor; Isothermal amplification; Wnt7B gene