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首页> 外文期刊>Biomolecular engineering >Nucleotide replacement at two sites can be directed by modified single-stranded oligonucleotides in vitro and in vivo
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Nucleotide replacement at two sites can be directed by modified single-stranded oligonucleotides in vitro and in vivo

机译：修饰的单链寡核苷酸可在体内和体外指导两个部位的核苷酸替换

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Studies involving the alteration of DNA sequences by modified single-stranded oligonucleotides in vitro and in vivo have revealed potential applications for functional genomics. Repair of a replacement, deletion, or insertion mutation has already been achieved with molecules having lengths between 25 and 74 bases. But, other vector parameters still remain to be explored. Here, the position of the single base in the vector directing the alteration was examined and the optimal site was found to be at or near the center of the vector. If that position is staggered 3' or 5', the frequencies of gene repair in vitro decreases. The potential of a single vector to direct two nucleotide changes at a specific site in a target sequence was also examined. Both targeted bases are corrected together at the same frequency if the sites are separated by three bases, but conversion linkage decreases precipitously when the distance is expanded to 15 and 27 nucleotides, respectively. These results suggest that single oligonucleotides can be used to direct nucleotide exchange at two independent sites, a reaction characteristic that may be useful for many genomics applications.
机译：涉及在体外和体内通过修饰的单链寡核苷酸改变DNA序列的研究已经揭示了功能基因组学的潜在应用。已经用长度在25至74个碱基之间的分子实现了替换，缺失或插入突变的修复。但是，其他向量参数仍有待探索。在此，检查了指导改变的载体中单个碱基的位置，发现最佳位点在载体中心或附近。如果该位置错开3'或5'，则体外基因修复的频率会降低。还检查了单个载体在靶序列中特定位点引导两个核苷酸变化的潜力。如果位点被三个碱基隔开，则两个目标碱基将以相同的频率一起校正，但是当距离分别扩展到15个和27个核苷酸时，转换键急剧下降。这些结果表明，单个寡核苷酸可用于指导两个独立位点的核苷酸交换，这一反应特性可能对许多基因组学应用有用。

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