A polymer Lab-on-a-Chip for genetic analysis using the arrayed primer extension on microarray chips
- 377 Downloads
In this work a polymer lab-on-a-chip (LOC), fabricated through MEMS technology, was employed to execute a genetic protocol for the Single Nucleotide Polymorphisms (SNPs) detection. The LOC was made in Poly (methyl methacrylate) (PMMA) and has two levels: the lower one for the insertion and mixing of the reagents, the upper one for the interfacing with the DNA microarray chip. The hereditary hearing loss was chosen as case of study, since the demand for testing such a particular disorder is high and genetics behind the condition is quite clear. The Arrayed Primer EXtension (APEX) genetic protocol was implemented on the LOC to analyze the SNPs. A low density (for detection of up to 10 mutations) and a high density microarray chips (for detection of 245 mutations in 12 genes), containing the primers for the extension, were employed to carry out the APEX reaction on the LOC. Both the microarray chips provide a signal to noise ratio and efficiency comparable with a detection obtained in a conventional protocol in standard conditions. Moreover, significant reduction of the employed PCR volume (from 30 μL to 10 μL) was obtained using the low density chip.
KeywordsLab-on-a-Chip DNA microarray PCR (polymerase chain reaction) APEX (Arrayed Primer EXtension) Single Nucleotide Polymorphisms (SNPs)
This work was funded and accomplished in the framework of the MANUNET ERA-Net Project “AUDAX - Automation of a Device based on APEX technology” (2008 Manunet Transnational Call), in cooperation with project EU31699 funded by Enterprise Estonia (LOC (Lab-on-a-chip) based genotyping technology –AUDAX applied research/product development.
- B. S. Gaylord, M. R. Massie, S. C. Feinstein, G. C. Bazan, Proc. Natl. Acad. Sci. U S A 102, 34–39 (2005)Google Scholar
- K.-Y. Hwang, J.-H. Kim, K.-Y. Suh, J. S. Ko, N. Huh,, Sens. and Actuators B: Chem. 155, 422–429 (2011)Google Scholar
- S.L. Marasso, E. Giuri, G. Canavese, R. Castagna, M. Quaglio, I. 442 Ferrante, D. Perrone, M. Cocuzza, Biomed. Microdevices 13, 19– 443 27 (2011a)Google Scholar
- S. L. Marasso, G. Canavese, M. Cocuzza, Microelectron. Eng. 88, 8 2322--2324 (2011b)Google Scholar
- M.-I. Mohammed, G. J. Sills, M. J. Brodie, E. M. Ellis, J. M. Girkin, Sens. and Actuators B: Chem. 139, 83–90 (2009)Google Scholar
- S. Petralia, R. Verardo, E. Klaric, S. Cavallaro, E. Alessi, C. Schneider, Sens. and Actuators B: Chem. 187, 99–105 (2013)Google Scholar
- M. Podder, J. Ruan, B. W. Tripp, Z. E. Chu, S. J. Tebbutt, BMC Med. Genomics 1, 5–5 (2008)Google Scholar
- A. Polini, Polini, E. Mele, A. G. Sciancalepore, S. Girardo, A. Biasco, A. Camposeo, R. Cingolani, D. A. Weitz, D. Pisignano, Biomicrofluid. 4, 3 15–18 (2010)Google Scholar
- J. Pullat, W. Kusnezow, K. Jaakson, M. Beier, J.D. Hoheisel, A. Metspalu, Biotechnol 25, 133–141 (2008)Google Scholar
- Soper, S. A., Ford, S. M., Qi, S., McCarley, R. L., Kelly, K., & Murphy, M. C., Anal. Chem. 72, 642 A-651 A (2000)Google Scholar