Biological lithography: development of a maskless microarray synthesizer for DNA chips

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Abstract

We have recently reported the development of a DNA maskless array synthesizer (MAS) [Singh-Gasson et al., Nat. Biotechnol. 17 (1999) 974]. In this paper we describe in more details the technical approach and implementation of the system, and show recent results of high-density DNA chips.

Introduction

The application of semiconductor fabrication techniques to biological problems has recently created a host of novel biological “devices” and products. Lithography, in particular, has been successfully applied to the fabrication of high-density micro arrays (HDMAs), for instance for the purpose of analyzing gene expression. The analysis is performed by observing the event of a match between a sample of m-RNA and a pre-programmed short strand of DNA (oligomer) located on a “chip”. All techniques rely on the extremely high selectivity in the bonding of DNA sequences, and vary in the techniques used for fabrication and detection.

We have developed a maskless exposure system that can quickly produce HDMAs with low cost and high efficiency. The system is based on the use of a Texas Instrument Digital Light Processor, DLP, as a virtual mask [6]. An optical system based on the Offner design forms a full-field 1X image of the pattern presented on the DLP chip on the glass window of a reaction chamber connected to a DNA synthesizer, Fig. 1. The synthesizer flows the reagents in the correct sequence in synchrony with the masks presented on the DLP chip, thus creating the microarray by a sequential operation. We note that in this approach there are no moving parts and hence no overlay issues. No masks are need to be fabricated, and the whole process is error-free and low cost. The fabrication time has been reduced to about 3 h. Hence, this new tool, the micro array synthesizer (MAS) will allow rapid turnaround experiments with DNA chips. Several units of the exposure systems have been built, and are currently in use at NimbleGen Systems for the production of custom HDMAs.

Section snippets

DNA chip fabrication

DNA—deoxyribonucleic acid—is the organic molecule that encodes all of the genetic information of an organism. Formed by a sequence of four bases (A, C, T, G) arranged in a linear chain, the DNA can store prodigious amount of information. The human genome contains 2×109 basis, and thus may assume in principle 42 billion different sequences. Because of its sequential nature, DNA can be “assembled” using a base-by-base (or layer-by-layer) approach. In one of the simplest processes, a so-called DNA

Instrument description

The exposure system includes an illumination system, the micromirror array (Texas Instrument DLP), the imaging optics and the reaction cell. The illumination system is based on a high-pressure mercury short arc lamp, followed by a filtering and uniformizing optical system. The source fills uniformly a disc about 25 mm in diameter. The light is directed at an angle of 20° from the axis of the DLP, to be deflected along the DLP normal when the mirrors are tilted in the +10° deflection state. This

Results

In Fig. 2 we present an image recorded using a photosensitive film. The individual pixels are well resolved, and the separation lines are, as expected, of the order of 2.5 μm. Exposure times are of the order of 70–100 s for a 100 mW/cm2 illumination level. One of the major differences between DNA photosynthesis and photoresist processing is the lack of a high-contrast chemistry. The deprotection reaction scales proportionally to the exposure dose, i.e., yields an exponential behavior;

Conclusions

We have presented the result of the initial development of the micro array synthesizer, a laboratory desktop unit that brings an unprecedented level of flexibility and turnaround time to DNA chips fabrication. With the MAS, a scientist can program and fabricate a DNA chip in a matter of hours, opening the way to hitherto impossible studies.

Acknowledgments

This work was initially funded by the University of Wisconsin and, in part, by a NIH SBIR grant. This work would not have been possible without the contribution of NimbleGen System staff (E. Nuywasir, T. Albert, K. Johnson, A. Pitas, J. Singh) and of CNTech (J. Wallace, Q. Leonard).

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