DNA polymerase κ (Polκ) is a specialized polymerase that has multiple cellular roles such as translesion DNA synthesis, replication of repetitive sequences, and nucleotide excision repair. We have developed a method for capturing DNA synthesized by Polκ utilizing a Polκ–specific substrate, N 2 -(4-ethynylbenzyl)-2′-deoxyguanosine (EBndG). After shearing of the DNA into 200 to 500 bp lengths, the EBndG-containing DNA was covalently bound to biotin using the Cu(I)-catalyzed alkyne–azide cycloaddition reaction and isolated with streptavidin beads. Isolated DNA was then ligated to adaptors, followed by PCR amplification and next-generation sequencing to generate genome-wide repair maps. We have termed this method polymerase κ sequencing. Here, we present the human genome maps for Polκ activity in an undamaged cell line. We found that Polκ activity was enhanced in GC-rich regions, euchromatin regions, the promoter of genes, and in DNA that is replicated early in the S phase.

中文翻译：

---

人类成纤维细胞基因组中 DNA 聚合酶 κ 的活性


DNA 聚合酶 κ (Polκ) 是一种特殊的聚合酶，具有多种细胞作用，例如跨损伤 DNA 合成、重复序列复制和核苷酸切除修复。我们开发了一种利用 Polκ 特异性底物 N 2 -(4-乙炔基苄基)-2'-脱氧鸟苷 (EBndG) 捕获 Polκ 合成的 DNA 的方法。将 DNA 剪切成 200 至 500 bp 长度后，使用 Cu(I) 催化的炔烃-叠氮化物环加成反应将含有 EBndG 的 DNA 与生物素共价结合，并用链霉亲和素珠分离。然后将分离的 DNA 连接到接头上，然后进行 PCR 扩增和下一代测序，以生成全基因组修复图谱。我们将这种方法称为聚合酶κ测序。在这里，我们展示了未受损细胞系中 Polκ 活性的人类基因组图谱。我们发现 Polκ 活性在富含 GC 的区域、常染色质区域、基因启动子以及在 S 期早期复制的 DNA 中增强。