Investigation of Hybridization Efficiency of a Sequence-Orientated Coaxial DNA Probe Microarryed on Biochips Using Atomic Force Microscope

Abstract:

Two sequence-inversed probes were microarrayed on glass slides to study the hybridization efficiency with their DNA targets. A fluorescence laser scanner and an atomic force microscope (AFM) were utilized to investigate the efficiency in different hybridization cases and their corresponding depth changes on the chips. The sequences of two targets were designed to be fully complementary to their shared DNA probe in a coaxial stacking configuration. In other words, after the first DNA target is hybridized (pre-hybridizing) onto the probe, the second one is stacked onto the non-hybridized region of the same probe. The pre-hybridizing and the second DNA targets were distinguished by two distinct fluorescent dyes. The enhancement of the hybridization efficiency was investigated through the comparison between the stacking and individual hybridization configurations. AFM was used to measure the depths of two probes at different steps of hybridization. The results indicated that the depths increased as the hybridization proceeded. Probe#1, pre-hybridizing close to the chip surface, obtained a thicker depth than the other probe pre-hybridizing away from the chip surface, Probe#2. A hypothesis was proposed to explain how the depth variation was associated with the observed hybridization efficiency.

• Periodical: Applied Mechanics and Materials (Volumes 284-287)
• Pages: 315-319
• DOI: https://doi.org/10.4028/www.scientific.net/AMM.284-287.315
• Online since: January 2013
• Authors: Jui Chuang Wu, Dan Kai Yang, Yane Shu Lin, Jun Yi Chen
• Keywords: Atomic Force Microscopy (AFM), DNA Biochips, DNA Hybridization Efficiency, Microarray, Stacking Hybridization

References

[1] R. Koehler and N. Peyret, Effects of DNA secondary structure on oligonucleotide probe binding efficiency, Comput. Biol. Chem., 29 (2005) 393-397.

DOI: 10.1016/j.compbiolchem.2005.09.002

[2] P. Wu, P. Hogrebe, and D. Grainger, DNA and protein microarray printing on silicon nitride waveguide surfaces, Biosens. Bioelectron., 21 (2006) 1252-1263.

DOI: 10.1016/j.bios.2005.05.010

[3] Y. Luo, A. Chen, and K. Li, Oligonucleotide probes with improved hybridization efficiency and specificity in detection of genetic polymorphisms in arrays , European Patent, WO2002095057 (2002).

[4] S. Tao, H. Gao, F. Cao, X. Ma, and J. Cheng, Blocking oligo-a novel approach for improving chip-based DNA hybridization efficiency, Mol. Cell. Probes, 17 (2003) 197-202.

DOI: 10.1016/s0890-8508(03)00053-7

[5] K.M. Lang, D. A. Hite, R. W. Simmonds, R. McDermott, D. P. Pappas, and John M. Martinis, Conducting atomic force microscopy for nanoscale tunnel barrier characterization, Review of Scientific Instruments 75 (2004) 2726-2731.

DOI: 10.1063/1.1777388

[6] A. Ebner, P. Hinterdorfer, and H. J. Gruber, Comparison of different aminofunctionalization strategies for attachment of single antibodies to AFM cantilevers , Ultramicroscopy, 107 (2007) 922-927.

DOI: 10.1016/j.ultramic.2007.02.035

[7] O. Medalia, J. Englander, R. Guckenberger, and J. Sperling, AFM imaging in solution of protein–DNA complexes formed on DNA anchored to a gold surface,Ultramicroscopy 90 (2002) 103-112.

DOI: 10.1016/s0304-3991(01)00141-3

[8] J. M. Song, M. Culha, P. M. Kasili, G. D. Griffin, and T. Vo-Dinh, A compact CMOS biochip immunosensor towards the detection of a single bacteria, Biosensors and Bioelectronics, 20 (2005) 2203-2209.

DOI: 10.1016/j.bios.2004.08.033

[9] M. H. Rouillat, V. Dugas, and J. R. Martin, Characterization of DNA chips on the molecular scale beforeand after hybridization with an atomic force microscope, App. Surface Sci., 252 (2005) 1765-1771.

DOI: 10.1016/j.apsusc.2005.03.132

[10] D. Yang, J. Huang, C. Chen, H. Su, J. Wu, Effect of co-axially hybridized gene targets on hybridization efficiency of microarrayed DNA probes, J. Chi Inst. Chem. Eng. 39 (2008) 187.

DOI: 10.1016/j.jtice.2010.04.012

[11] K. Jiang, J. Huang, C. Chen, H. Su, J. Wu, Effect of co-axially hybridized gene targets on hybridization efficiency of microarrayed DNA probes, J. Taiwan Inst. Chem. Eng, 42 (2011) 5-12.

DOI: 10.1016/j.jtice.2010.04.012