Saitoh Hiroshi, Nakamura Akiko, Kuwahara Masayasu, Ozaki Hiroaki, Sawai Hiroaki
Department of Applied Chemistry, Faculty of Engineering, Gunma University, Kiryu, Gunma 376-8515, Japan.
Nucleic Acids Res Suppl. 2002(2):215-6. doi: 10.1093/nass/2.1.215.
We obtained a modified DNA aptamer against sweetener, aspartame, by in vitro selection method. The modified DNA was prepared from dATP, dGTP, dCTP and a modified dTTP bearing a terminal amino group at C-5 position in place of thymidine by PCR using a hyper thermophilic DNA polymerase, KOD Dash DNA polymerase. The synthetic 102-mer DNA with a 60-mer random region was used as an initial template for the PCR. The PCR-amplified modified DNA library was applied to an aspartame-agarose column, and then the bound modified DNA was eluted from the column for the affinity chromatography selection. Repeating the procedure, we selected the modified DNA aptamer against aspartame.
我们通过体外筛选方法获得了一种针对甜味剂阿斯巴甜的修饰DNA适配体。修饰DNA由dATP、dGTP、dCTP和一种在C-5位带有末端氨基取代胸腺嘧啶的修饰dTTP，使用超嗜热DNA聚合酶KOD Dash DNA聚合酶通过PCR制备。具有60个碱基随机区域的合成102聚体DNA用作PCR的初始模板。将PCR扩增的修饰DNA文库应用于阿斯巴甜琼脂糖柱，然后从柱上洗脱结合的修饰DNA用于亲和色谱筛选。重复该过程，我们筛选出了针对阿斯巴甜的修饰DNA适配体。
