Affinity Isolation of Defined Genomic Fragments Cleaved by Nuclease S1-based Artificial Restriction DNA Cutter
Affinity Isolation of Defined Genomic Fragments Cleaved by Nuclease S1-based Artificial Restriction DNA Cutter
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基于核酸酶 S1 的人工限制性 DNA 切割器切割的确定基因组片段的亲和分离
DOI：
10.1002/cpnc.76
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发表时间：
2019
影响因子：
--
通讯作者：
Makoto Komiyama
中科院分区：
文献类型：
--
作者：
Arivazhagan Rajendran;Narumi Shigi;Jun Sumaoka;Makoto Komiyama
人类基因组极易受到各种修饰、损伤和损伤的影响。为了分析与人类基因组的特定区域结合的损伤和蛋白质，基因组应该在所需的位置被片段化，并且应该分离出感兴趣的区域。少数可用的分离人类基因组所需区域的方法存在严重缺陷，只能应用于特定的序列或需要繁琐的实验程序。我们最近开发了一种新的方法，利用一对伪互补多肽核酸(PcPNAs)和S1核酸酶来分离DNA定点切割所释放的所需基因组片段。当与生物素结合时，其中一个pcPNAs可用于亲和纯化切割产物。在这里，我们报告了一种详细的方法来分离确定的千碱基长度的DNA片段，这些片段可以应用于质粒或基因组DNA，并且不受序列的限制。©2019 John Wiley&Sons，Inc.
The human genome is highly susceptible to various modifications, lesions, and damage. To analyze lesions and proteins bound to a defined region of the human genome, the genome should be fragmented at desired sites and the region of interest should be isolated. The few available methods for isolating a desired region of the human genome have serious drawbacks and can only be applied to specific sequences or require tedious experimental procedures. We have recently developed a novel method to isolate a desired fragment of the genome released by site‐specific scission of DNA using a pair of pseudo‐complementary peptide nucleic acids (pcPNAs) and S1 nuclease. When conjugated to biotin, one of the pcPNAs can be used to affinity purify the cleavage product. Here we report a detailed protocol to isolate defined kilobase‐length DNA fragments that can be applied to plasmid or genomic DNA and is not limited by sequence. © 2019 by John Wiley & Sons, Inc.