• 文献检索
  • 文档翻译
  • 深度研究
  • 学术资讯
  • Suppr Zotero 插件Zotero 插件
  • 邀请有礼
  • 套餐&价格
  • 历史记录
应用&插件
Suppr Zotero 插件Zotero 插件浏览器插件Mac 客户端Windows 客户端微信小程序
定价
高级版会员购买积分包购买API积分包
服务
文献检索文档翻译深度研究API 文档MCP 服务
关于我们
关于 Suppr公司介绍联系我们用户协议隐私条款
关注我们

Suppr 超能文献

核心技术专利：CN118964589B侵权必究
粤ICP备2023148730 号-1Suppr @ 2026

文献检索

告别复杂PubMed语法，用中文像聊天一样搜索，搜遍4000万医学文献。AI智能推荐，让科研检索更轻松。

立即免费搜索

文件翻译

保留排版，准确专业，支持PDF/Word/PPT等文件格式，支持 12+语言互译。

免费翻译文档

深度研究

AI帮你快速写综述，25分钟生成高质量综述，智能提取关键信息，辅助科研写作。

立即免费体验

由Taq聚合酶合成的胞嘧啶甲基化DNA用于检测限制性内切酶HinfI的甲基化敏感性。

Cytosine methylated DNA synthesized by Taq polymerase used to assay methylation sensitivity of restriction endonuclease HinfI.

作者信息

Colasanti J, Sundaresan V

机构信息

Cold Spring Harbor Laboratory, NY 11724.

出版信息

Nucleic Acids Res. 1991 Jan 25;19(2):391-4. doi: 10.1093/nar/19.2.391.

DOI:10.1093/nar/19.2.391
PMID:2014176
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC333607/
Abstract

We have studied the resistance of cytosine methylated DNA to digestion by the restriction endonuclease HinfI, using a simple PCR procedure to synthesize DNA of known sequence in which every cytosine is methylated at the 5 position. We find that HinfI cannot digest cytosine methylated DNA at the concentrations normally used in restriction digests. Complete digestion is possible using a vast excess of enzyme; under these conditions, the rate of HinfI digestion for cytosine methylated DNA is at least 1440-fold slower than for unmethylated DNA. The presence of an additional methylated cytosine at the degenerate position internal to the recognition sequence does not appear to increase the resistance to HinfI digestion. We also tested HhaII, an isoschizomer of HinfI, and found that it is completely inactive on cytosine methylated DNA. The procedure we have used should be of general applicability in determination of the methylation sensitivities of other restiction enzymes, as well as studies of the effects of methylation on gene expression in direct DNA transfer experiments.

摘要

我们使用一种简单的聚合酶链反应（PCR）程序来合成已知序列的DNA，其中每个胞嘧啶在5位被甲基化，以此研究了胞嘧啶甲基化DNA对限制性内切酶HinfI消化的抗性。我们发现，在限制性消化通常使用的浓度下，HinfI不能消化胞嘧啶甲基化DNA。使用大量过量的酶可以实现完全消化；在这些条件下，胞嘧啶甲基化DNA的HinfI消化速率比未甲基化DNA至少慢1440倍。识别序列内部简并位置处额外的甲基化胞嘧啶的存在似乎不会增加对HinfI消化的抗性。我们还测试了HinfI的同裂酶HhaII，发现它对胞嘧啶甲基化DNA完全无活性。我们所使用的程序在确定其他限制性酶的甲基化敏感性以及直接DNA转移实验中甲基化对基因表达的影响的研究中应具有普遍适用性。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/480036d79cea/nar00238-0184-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/3099cc3d9604/nar00238-0183-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/a925e5e691b2/nar00238-0184-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/480036d79cea/nar00238-0184-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/3099cc3d9604/nar00238-0183-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/a925e5e691b2/nar00238-0184-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/51bd/333607/480036d79cea/nar00238-0184-b.jpg

相似文献

1
Cytosine methylated DNA synthesized by Taq polymerase used to assay methylation sensitivity of restriction endonuclease HinfI.由Taq聚合酶合成的胞嘧啶甲基化DNA用于检测限制性内切酶HinfI的甲基化敏感性。
Nucleic Acids Res. 1991 Jan 25;19(2):391-4. doi: 10.1093/nar/19.2.391.
2
Restriction endonuclease cleavage of 5-methyl-deoxycytosine hemimethylated DNA at high enzyme-to-substrate ratios.在高酶与底物比例下，5-甲基脱氧胞嘧啶半甲基化DNA的限制性内切酶切割
Nucleic Acids Res. 1993 Feb 11;21(3):681-6. doi: 10.1093/nar/21.3.681.
3
Cytosine methylation: quantitation by automated genomic sequencing and GENESCAN analysis.胞嘧啶甲基化：通过自动基因组测序和基因扫描分析进行定量
Biotechniques. 1996 Jul;21(1):126-33. doi: 10.2144/96211rr04.
4
Restriction endonuclease isoschizomers ItaI, BsoFI and Fsp4HI are characterised by differences in their sensitivities to CpG methylation.限制性内切核酸酶同裂酶ItaI、BsoFI和Fsp4HI的特征在于它们对CpG甲基化的敏感性存在差异。
Nucleic Acids Res. 1997 Aug 15;25(16):3196-8. doi: 10.1093/nar/25.16.3196.
5
In Z-DNA the sequence G-C-G-C is neither methylated by Hha I methyltransferase nor cleaved by Hha I restriction endonuclease.在Z-DNA中，G-C-G-C序列既不会被Hha I甲基转移酶甲基化，也不会被Hha I限制性内切酶切割。
Proc Natl Acad Sci U S A. 1984 Jun;81(11):3268-72. doi: 10.1073/pnas.81.11.3268.
6
Cloning, expression, and purification of a thermostable nonhomodimeric restriction enzyme, BslI.一种耐热非同源二聚体限制酶BslI的克隆、表达及纯化
J Bacteriol. 2000 Feb;182(4):949-55. doi: 10.1128/JB.182.4.949-955.2000.
7
The effect of C(5) cytosine methylation at CpG sequences on mitomycin-DNA bonding profiles.CpG序列中C(5)胞嘧啶甲基化对丝裂霉素-DNA结合谱的影响。
Bioorg Med Chem. 2001 Apr;9(4):863-73. doi: 10.1016/s0968-0896(00)00301-1.
8
Measurement of DNA-protein equilibria using gel chromatography: application to the HinfI restriction endonuclease.使用凝胶色谱法测量DNA-蛋白质平衡：应用于HinfI限制性内切酶
Biochemistry. 1985 Jun 4;24(12):3049-54. doi: 10.1021/bi00333a037.
9
Site-dependent cleavage of pBR322 DNA by restriction endonuclease HinfI.限制性内切酶HinfI对pBR322 DNA的位点依赖性切割。
Nucleic Acids Res. 1983 Jun 25;11(12):4109-26. doi: 10.1093/nar/11.12.4109.
10
Recognition of native DNA methylation by the PvuII restriction endonuclease.PvuII限制性内切酶对天然DNA甲基化的识别。
Nucleic Acids Res. 2000 Aug 15;28(16):3143-50. doi: 10.1093/nar/28.16.3143.

引用本文的文献

1
Restriction endonuclease cleavage of 5-methyl-deoxycytosine hemimethylated DNA at high enzyme-to-substrate ratios.在高酶与底物比例下，5-甲基脱氧胞嘧啶半甲基化DNA的限制性内切酶切割
Nucleic Acids Res. 1993 Feb 11;21(3):681-6. doi: 10.1093/nar/21.3.681.
2
PCR with 5-methyl-dCTP replacing dCTP.使用5-甲基-dCTP替代dCTP进行聚合酶链式反应（PCR）。
Nucleic Acids Res. 1991 Mar 11;19(5):1081-5. doi: 10.1093/nar/19.5.1081.
3
Binding sites for maize nuclear proteins in the terminal inverted repeats of the Mu1 transposable element.玉米核蛋白在Mu1转座元件末端反向重复序列中的结合位点。

本文引用的文献

1
Digestion of highly modified bacteriophage DNA by restriction endonucleases.用限制性内切核酸酶消化高度修饰的噬菌体DNA。
Nucleic Acids Res. 1982 Mar 11;10(5):1579-91. doi: 10.1093/nar/10.5.1579.
2
Restriction enzyme digestion of hemimethylated DNA.半甲基化DNA的限制性内切酶消化
Nucleic Acids Res. 1981 Jun 11;9(11):2509-15. doi: 10.1093/nar/9.11.2509.
3
The somatic replication of DNA methylation.DNA甲基化的体细胞复制。
Mol Gen Genet. 1991 Sep;229(1):17-26. doi: 10.1007/BF00264208.
Cell. 1981 Apr;24(1):33-40. doi: 10.1016/0092-8674(81)90498-0.
4
Inheritance of mutator activity in Zea mays as assayed by somatic instability of the bz2-mu1 allele.通过bz2-mu1等位基因的体细胞不稳定性测定玉米中突变活性的遗传。
Genetics. 1986 Dec;114(4):1293-312. doi: 10.1093/genetics/114.4.1293.
5
The inheritance of epigenetic defects.表观遗传缺陷的遗传
Science. 1987 Oct 9;238(4824):163-70. doi: 10.1126/science.3310230.
6
Effect of in vitro DNA methylation on beta-globin gene expression.体外DNA甲基化对β-珠蛋白基因表达的影响。
Proc Natl Acad Sci U S A. 1988 Jul;85(13):4638-42. doi: 10.1073/pnas.85.13.4638.
7
Molecular analysis of the maize anthocyanin regulatory locus C1.玉米花青素调控位点C1的分子分析
Proc Natl Acad Sci U S A. 1986 Dec;83(24):9631-5. doi: 10.1073/pnas.83.24.9631.
8
Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors.改良的M13噬菌体克隆载体和宿主菌株：M13mp18和pUC19载体的核苷酸序列。
Gene. 1985;33(1):103-19. doi: 10.1016/0378-1119(85)90120-9.
9
Effect of site-specific methylation on DNA modification methyltransferases and restriction endonucleases.位点特异性甲基化对DNA修饰甲基转移酶和限制性内切核酸酶的影响。
Nucleic Acids Res. 1989;17 Suppl(Suppl):r389-415. doi: 10.1093/nar/17.suppl.r389.
10
Restriction enzymes and their isoschizomers.限制性内切酶及其同裂酶。
Nucleic Acids Res. 1989;17 Suppl(Suppl):r347-87. doi: 10.1093/nar/17.suppl.r347.