基于稳定同位素标记的DNA测序方法初步探索

Fluorescent dye labeling-based approaches are the standard methods for DNA sequencing, featuring of high sensitivity, high throughput, and cost-effectiveness. However, fluorescent dyes also have intrinsic limitations, such as complicated spectra interferences during multiple dye component detection. In this project, we will explorer new DNA sequencing method based on stable isotope labeling and capillary electrophoresis-inductively coupled plasma mass spectrometry detection. The stable isotope labeling-based DNA sequencing method is expected to act as an effective supplementary to the existing fluorescent dye labeling-based method. Due to the absence of limitations associating with fluorescent dye (complicated spectra correction), the proposed strategy may realize the accurate sequencing and comparison of two or even multiple DNAs in a single capillary.The problems derived from varied electrophoretic mobility of multiple capillary will be overcome during the comparison of DNA sequences. The strategy is expected to play an important role in sequence variant validation, forensic evidence identification, and other applications in need of accurate sequence comparison.

基于荧光染料标记体系的DNA测序方法具有高灵敏度、高通量和低成本的优点，是目前的普遍使用的标准测序方法。但是荧光染料作为标记物用于核酸测序时也有其局限性，比如在激光照射下多组分同时检测时需要复杂的光谱校正等。在本项目中，我们拟探索采用稳定同位素（镧系元素同位素）标记技术，与毛细管电泳-电感耦合等离子体质谱检测相结合，构建DNA测序分析新方法。稳定同位素标记测序方法有望成为现有荧光标记高通量测序方法的有效补充。由于排除了荧光染料的多色荧光光谱干扰的局限，基于稳定同位素标记的测序方法有望实现在同一根毛细管中两条甚至多条DNA的序列分析和精准比对。同一毛细管中的序列比对将克服不同毛细管泳道间迁移率的差异对测序结果重现性和准确度的影响。方法有望在序列变异确认、法医证物鉴定等需要精准比对的应用领域发挥积极作用。