Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 2018, 4178045
eCollection

DNA Nanotweezers With Hydrolytic Activity for Enzyme-Free and Sensitive Detection of Fusion Gene via Logic Operation

Affiliations

DNA Nanotweezers With Hydrolytic Activity for Enzyme-Free and Sensitive Detection of Fusion Gene via Logic Operation

Yongjie Xu et al. J Anal Methods Chem.

Abstract

Gene fusion is a molecular event occurring in cellular proliferation and differentiation, and the occurrence of irregular fusion gene results in various malignant diseases. So, sensing fusion gene with high performance is an important task for integrating individual disease information. Here, we proposed a nonenzymatic and high-throughput fluorescent assay system for the detection of fusion gene by employing DNA nanotweezers with hydrolytic activity. This tweezer was assembled by three single-stranded DNAs and engineered with sensing elements and reporting subunits. In the absence of the fusion gene, the engineered tweezer remained opened and inactive which led to no signal output. However, the addition of fusion genes would cause structure alterations of the tweezer from open to close and further DNAzyme activation with the assembly of two reporting subunits. Then, the activated DNAzyme catalyzed fluorescence substrates for signal conversion. Taking BCR/ABL fusion gene as an example, the tweezer-based assay system showed not only excellent distinguishing capability towards different input targets but also high sensitivity with a detection limit of 5.29 pM. In addition to good detection performance, this system was simple and enzyme-free, offering a powerful nanometer tool as a smart nanodevice for sensing fusion detection.

Figures

Scheme 1
Scheme 1
Schematic illustration of the fluorescence imaging method based on functional DNA nanotweezer regulated by fusion gene.
Figure 1
Figure 1
Characterization of the assembly process of DNA nanotweezer using agarose gel electrophoresis. Lanes 1, 2, 3, and 4 represents strand A, strand B, strand C, and BCR/ABL fusion gene, respectively, and lanes 5 and 6 are strand A + C and strand B + C, lane 7 represents opened nanotweezer assembled from strand A + B + C, and lane 8 represented closed nanotweezer after adding BCR/ABL fusion gene to the system in lane 7. Inset: the images a and b in lanes 7 and 8 represents the nanotweezer-based assay system without and with BCR/ABL fusion gene, respectively.
Figure 2
Figure 2
Optimization of experimental conditions. (a) Time-dependent fluorescence changes from 20, 30, 40, 50, 60, to 70 min. (b) DNAzyme cleavage activity on varied reaction temperature at 25, 30 35, 40, 45, and 50°C. (c) The effect of bases number on hinge region on signal to noise by setting at 3, 4, 5, 6, and 7 nucleotides.
Figure 3
Figure 3
(a) Fluorescence images of BCR/ABL fusion gene at 0, 1 × 10−11, 5 × 10−11, 1 × 10−10, 5 × 10−10, 1 × 10−9, 5 × 10−9, and 1 × 10−8 M (image and bar a to h). (b) Dose-response curves for BCR/ABL fusion gene from 1 × 10−11 to 1 × 10−8 M. Inset: the corresponding calibration curve.
Figure 4
Figure 4
Fluorescence responses to different target inputs: random sequence (image and bar a), blank control (image and bar b), ABL gene (image and bar c), BCR gene (image and bar d), BCR/ABL fusion gene (image and bar e), and the mixture of ABL gene, BCR gene, and BCR/ABL fusion gene (image and bar f).

Similar articles

See all similar articles

Cited by 1 article

References

    1. Rabbitts T. H. Chromosomal translocations in human cancer. Nature. 1994;372(6502):143–149. doi: 10.1038/372143a0. - DOI - PubMed
    1. Rowley J. D. A new consistent chromosomal abnormality in chronic myelogenous leukaemia identified by quinacrine fluorescence and Giemsa staining. Nature. 1973;243(5405):290–293. doi: 10.1038/243290a0. - DOI - PubMed
    1. Heim S., Mitelman F. Molecular screening for new fusion genes in cancer. Nature Genetics. 2008;40(6):685–686. doi: 10.1038/ng0608-685. - DOI - PubMed
    1. D’Alessio F., Mirabelli P., Mariotti E., et al. Miniaturized flow cytometry-based BCR-ABL immunoassay in detecting leptomeningeal disease. Leukemia Research. 2011;35(10):1290–1293. doi: 10.1016/j.leukres.2011.05.016. - DOI - PubMed
    1. Soverini S., Hochhaus A., Nicolini F. E., et al. BCR-ABL kinase domain mutation analysis in chronic myeloid leukemia patients treated with tyrosine kinase inhibitors: recommendations from an expert panel on behalf of European LeukemiaNet. Blood. 2011;118(5):1208–1215. doi: 10.1182/blood-2010-12-326405. - DOI - PubMed

LinkOut - more resources

Feedback