REAL-TIME DETECTION OF DNA HYBRIDIZATION AND MELTING ON OLIGONUCLEOTIDE ARRAYS BY USING OPTICAL-WAVE GUIDES

被引:117
作者
STIMPSON, DI
HOIJER, JV
HSIEH, WT
JOU, C
GORDON, J
THERIAULT, T
GAMBLE, R
BALDESCHWIELER, JD
机构
[1] Abbott Laboratories, North Chicago
[2] California Institute of Technology, Pasadena
[3] Haviland Meteorite Crater, Haviland
[4] Oncor, Gaithersburg, MD 20877
关键词
D O I
10.1073/pnas.92.14.6379
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
The challenge of the Human Genome Project is to increase the rate of DNA sequence acquisition by two orders of magnitude to complete sequencing of the human genome by the year 2000. The present work describes a rapid detection method using a two-dimensional optical wave guide that allows measurement of real-time binding or melting of a light-scattering label on a DNA array. A particulate label on the target DNA acts as a light-scattering source when illuminated by the evanescent wave of the wave guide and only the label bound to the surface generates a signal. Imaging/visual examination of the scattered light permits interrogation of the entire array simultaneously. Hybridization specificity is equivalent to that obtained with a conventional system using autoradiography. Wave guide melting curves are consistent with those obtained in the liquid phase and single-base discrimination is facile. Dilution experiments showed an apparent lower limit of detection at 0.4 nM oligonucleotide. This performance is comparable to the best currently known fluorescence-based systems. In addition, wave guide detection allows manipulation of hybridization stringency during detection and thereby reduces DNA chip complexity. It is anticipated that this methodology will provide a powerful tool for diagnostic applications that require rapid cost-effective detection of variations from known sequences.
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页码:6379 / 6383
页数:5
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