Zhang J, Fang Y, Hou J Y, Ren H J, Jiang R, Roos P, Dovichi N J
Department of Chemistry, University of Alberta, Edmonton, Canada.
Anal Chem. 1995 Dec 15;67(24):4589-93. doi: 10.1021/ac00120a026.
Four-color DNA cycle sequencing was performed on an M13mp18 template using dye-labeled primers. Sequencing fragments were separated by capillary electrophoresis at 60 degrees C and at an electric field of 150 V/cm. The sieving medium was 5%T, non-cross-linked polyacrylamide in 7 M urea. The use of high temperature for the separation reduces formation of secondary structures in the sequencing fragments, generating a sequence that is free of compressions without the use of strongly denaturing gels. The use of high temperatures also increases the separation rate compared with room-temperature operation. Fragments up to 640 bases are separated in less than 2 h.
使用染料标记引物对M13mp18模板进行四色DNA循环测序。测序片段在60℃、150V/cm电场下通过毛细管电泳分离。筛分介质为7M尿素中的5%T非交联聚丙烯酰胺。在分离过程中使用高温可减少测序片段中二级结构的形成，无需使用强变性凝胶即可生成无压缩的序列。与室温操作相比，高温的使用还提高了分离速率。长达640个碱基的片段在不到2小时内即可分离。
