{"title":"真核细胞瞬时测定DNA复制的改进方法","authors":"Maggie Sullivan, William R. Folk","doi":"10.1016/0735-0651(88)90016-7","DOIUrl":null,"url":null,"abstract":"<div><p>We describe an improved method for quantifying covalently closed circular DNAs replicated in eukaryotic cells. Normally, for such assays, plasmid DNAs transfected into animal cells must be extensively digested with restriction enzymes such as Dpnl and the products of digestion of the input DNA separated from the Dpnl-resistant (replicated) DNAs by electrophoresis. In the procedure described, the input, unreplicated DNA is nicked by light digestion with Dpnl and subsequently denatured and eliminated by S1 nuclease digestion. The replicated Dpnl-resistant DNA is detected by slot blot hybridization. Multiple samples can be rapidly assayed, with a considerable reduction in expense and labor.</p></div>","PeriodicalId":77714,"journal":{"name":"Gene analysis techniques","volume":"5 3","pages":"Pages 54-56"},"PeriodicalIF":0.0000,"publicationDate":"1988-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/0735-0651(88)90016-7","citationCount":"1","resultStr":"{\"title\":\"An improved procedure for measuring DNA replication with transient assays in eukaryotic cells\",\"authors\":\"Maggie Sullivan, William R. Folk\",\"doi\":\"10.1016/0735-0651(88)90016-7\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>We describe an improved method for quantifying covalently closed circular DNAs replicated in eukaryotic cells. Normally, for such assays, plasmid DNAs transfected into animal cells must be extensively digested with restriction enzymes such as Dpnl and the products of digestion of the input DNA separated from the Dpnl-resistant (replicated) DNAs by electrophoresis. In the procedure described, the input, unreplicated DNA is nicked by light digestion with Dpnl and subsequently denatured and eliminated by S1 nuclease digestion. The replicated Dpnl-resistant DNA is detected by slot blot hybridization. Multiple samples can be rapidly assayed, with a considerable reduction in expense and labor.</p></div>\",\"PeriodicalId\":77714,\"journal\":{\"name\":\"Gene analysis techniques\",\"volume\":\"5 3\",\"pages\":\"Pages 54-56\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"1988-05-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1016/0735-0651(88)90016-7\",\"citationCount\":\"1\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Gene analysis techniques\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/0735065188900167\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Gene analysis techniques","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/0735065188900167","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
An improved procedure for measuring DNA replication with transient assays in eukaryotic cells
We describe an improved method for quantifying covalently closed circular DNAs replicated in eukaryotic cells. Normally, for such assays, plasmid DNAs transfected into animal cells must be extensively digested with restriction enzymes such as Dpnl and the products of digestion of the input DNA separated from the Dpnl-resistant (replicated) DNAs by electrophoresis. In the procedure described, the input, unreplicated DNA is nicked by light digestion with Dpnl and subsequently denatured and eliminated by S1 nuclease digestion. The replicated Dpnl-resistant DNA is detected by slot blot hybridization. Multiple samples can be rapidly assayed, with a considerable reduction in expense and labor.
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