Stevenson C L, Johnson R W, Vo-Dinh T
Health and Safety Research Division, Oak Ridge National Laboratory, TN 37831-6101.
Biotechniques. 1994 Jun;16(6):1104-11.
A method fo nonradioactive DNA sequencing, which has been marketed commercially, uses four different fluorescent tags to label DNA fragments and fixed-wavelength excitation/fluorescence detection of the labels. This study presents an alternative method of detection based on synchronous luminescence (SL), in which both excitation and emission wavelengths are scanned simultaneously. This approach has proven in the past to have significant advantages over fixed-wavelength luminescence in the analysis and identification of fluorescent analytes. In this paper, the utility of synchronous excitation was investigated as a method for DNA sequencing with fluorescent tags. A laser-based SL instrument, recently developed in this laboratory, was used to resolve the spectra of a mixture of the dyes used in the fluorescence-based sequencing scheme. A preliminary limit of detection of 720 zeptomoles (10(-21) M) of fluorescein isothiocyanate dye in solution was achieved with this instrument. The results presented here suggest that the SL technique could result in an increase in sensitivity and a decrease in error rate of identification during DNA sequencing.
一种已在市场上商业化销售的非放射性DNA测序方法，使用四种不同的荧光标签标记DNA片段，并对标签进行固定波长激发/荧光检测。本研究提出了一种基于同步发光（SL）的替代检测方法，其中激发波长和发射波长同时进行扫描。过去已证明，在荧光分析物的分析和鉴定方面，这种方法比固定波长发光具有显著优势。本文研究了同步激发作为一种使用荧光标签进行DNA测序的方法的实用性。本实验室最近开发的基于激光的SL仪器用于解析荧光测序方案中所用染料混合物的光谱。用该仪器实现了溶液中异硫氰酸荧光素染料720仄摩尔（10^(-21) M）的初步检测限。此处给出的结果表明，SL技术可能会提高DNA测序过程中的灵敏度并降低鉴定错误率。
