Anti-gene padlocks eliminate Escherichia coli based on their genotype.
Anti-gene padlocks eliminate Escherichia coli based on their genotype.
复制标题
反基因挂锁根据基因型消除大肠杆菌。
DOI：
10.1093/jac/dkm482
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发表时间：
2008
期刊：
影响因子：
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通讯作者：
Eshleman,JamesR
中科院分区：
文献类型：
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作者：
Shi,Chanjuan;Parker,AntonyR;Hua,Li;Morrell,CraigN;Lee,SooChin;Bandaru,Viswanath;Dumler,JStephen;Wu,TC;Eshleman,JamesR
目前存在几种靶向核酸的治疗策略;然而，大多数方法靶向信使RNA，而不是基因组DNA。我们描述了一种新的基于阿糖胞苷的策略，称为抗基因挂锁（AGPs），消除土方ichiacolibased对他们的genotype.MethodsThe战略采用了一种寡核苷酸与双发夹结构的AGP的两条链是互补的两条链的靶基因。我们测试了AGP的体外结合和DNA聚合的抑制。将AGPs电穿孔到细菌细胞中，与氨苄青霉素抗性质粒一起沿着有和没有基因靶点，并测量细胞存活率。ResultsIn vitro，AGPs以序列依赖的方式结合DNA靶点并抑制DNA合成。当转化到基因组中含有10、20或30个bplacZ或20个bpproADNA靶标的细菌细胞中时，AGP选择性地杀死或以其他方式抑制这些细胞的生长，而那些缺乏靶标的细胞表现出很小的毒性（如果有的话）。单次转化导致携带靶细胞损失约30%至40%。结构-功能实验进行定义必需的AGP requires.ConclusionsThese结果表明，AGP可能是一个有用的工具，以消除特定的细胞群体。
ObjectivesSeveral therapeutic strategies that target nucleic acids exist; however, most approaches target messenger RNA, rather than genomic DNA. We describe a novel oligonucleotide-based strategy, called anti-gene padlocks (AGPs), which eliminateEscherichiacolibased on their genotype.MethodsThe strategy employs an oligonucleotide with a double hairpin structure where both strands of the AGP are complementary to both strands of a target gene. We tested AGPs forin vitrobinding and inhibition of DNA polymerization. AGPs were electroporated into bacterial cells with and without gene targets along with an ampicillin resistance plasmid, and cell survival was measured.ResultsIn vitro, AGPs bound the DNA target in a sequence-dependent fashion and inhibited DNA synthesis. When transformed into bacterial cells containing 10, 20 or 30 bplacZor 20 bpproADNA targets in their genomes, AGPs selectively killed or otherwise inhibited growth of these cells, while those lacking the target demonstrated little, if any, toxicity. A single transformation resulted in ∼30% to 40% loss of target-bearing cells. Structure–function experiments were performed to define essential AGP requirements.ConclusionsThese results suggest that AGPs may be a useful tool to eliminate specific cell populations.