Li C, Tucker P W
Department of Microbiology, University of Texas Southwestern Medical Center, Dallas 75235.
Nucleic Acids Res. 1993 Mar 11;21(5):1239-44. doi: 10.1093/nar/21.5.1239.
We have developed a strategy for DNA sequencing based on exonuclease III digestion followed by double strand specific endonuclease digestion and direct dideoxynucleotide sequencing reaction. This strategy eliminates the need for subcloning, oligonucleotide primers, and prior knowledge of the DNA to be sequenced. All template and primer duplexes needed for sequencing a complete insert can be prepared in one day from uncharacterized starting DNA. Sequence information can be obtained from different regions of the DNA simultaneously. The method uses double-stranded DNA to generate single-stranded template and primer, and thus produces high quality sequence results. Commercially available dideoxy-sequencing kits are well suited for this method. The strategy should be applicable for both automatic and routine laboratory DNA sequencing.
我们已经开发出一种基于核酸外切酶III消化，随后进行双链特异性核酸内切酶消化和直接双脱氧核苷酸测序反应的DNA测序策略。该策略无需亚克隆、寡核苷酸引物以及对要测序的DNA的先验知识。从未表征的起始DNA中，一天内就能制备出测序完整插入片段所需的所有模板和引物双链体。可同时从DNA的不同区域获得序列信息。该方法利用双链DNA生成单链模板和引物，从而产生高质量的序列结果。市售的双脱氧测序试剂盒非常适合这种方法。该策略应适用于自动和常规实验室DNA测序。
