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识别序列CGCG中任何一个胞嘧啶的甲基化都会抑制ThaI对DNA的切割。

Methylation of either cytosine in the recognition sequence CGCG inhibits ThaI cleavage of DNA.

作者信息

Strobl J S, Thompson E B

出版信息

Nucleic Acids Res. 1984 Nov 12;12(21):8073-83. doi: 10.1093/nar/12.21.8073.

DOI:10.1093/nar/12.21.8073
PMID:6209609
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC320278/
Abstract

ThaI (CGCG) sites which overlap HhaI (GCGC) sites in phi X174 and pBR322 DNA were methylated in vitro with HhaI methylase and S-adenosylmethionine to yield CGmCG, mCGCG or mCGmCG (5-methylcytosine, mC). Methylation of either cytosine in the ThaI recognition sequence rendered the DNA resistant to ThaI cleavage. Rat pituitary cell genomic DNA was digested with ThaI or 2 other known methylation-sensitive enzymes, AvaI or XhoI. After electrophoresis and ethidium bromide straining of the DNA, all 3 enzymes showed the infrequent DNA cleavage characteristic of methylation-sensitive enzymes. Comparison of pituitary growth hormone (GH) genes bearing strain-specific degrees of methylation showed the less methylated gene to be more frequently cut by either AvaI or ThaI. ThaI resistant sites in GH genes were cleaved by ThaI after exposing cells to 5-azacytidine, an inhibitor of DNA methylation. We conclude that ThaI is a useful restriction enzyme for the analysis of mC at CGCG sequences in eukaryotic DNA.

摘要

在φX174和pBR322 DNA中与HhaI（GCGC）位点重叠的ThaI（CGCG）位点，在体外使用HhaI甲基化酶和S-腺苷甲硫氨酸进行甲基化，产生CGmCG、mCGCG或mCGmCG（5-甲基胞嘧啶，mC）。ThaI识别序列中任何一个胞嘧啶的甲基化都会使DNA对ThaI切割产生抗性。用ThaI或其他两种已知的甲基化敏感酶AvaI或XhoI消化大鼠垂体细胞基因组DNA。对DNA进行电泳和溴化乙锭染色后，所有这3种酶都显示出甲基化敏感酶特有的罕见DNA切割特征。比较具有菌株特异性甲基化程度的垂体生长激素（GH）基因，发现甲基化程度较低的基因更频繁地被AvaI或ThaI切割。在将细胞暴露于DNA甲基化抑制剂5-氮杂胞苷后，GH基因中的ThaI抗性位点被ThaI切割。我们得出结论，ThaI是分析真核DNA中CGCG序列处mC的有用限制酶。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/0641a26677b6/nar00339-0128-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/b54f93e90d4b/nar00339-0123-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/fc98f18709ca/nar00339-0125-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/e7138e71739e/nar00339-0126-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/0641a26677b6/nar00339-0128-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/b54f93e90d4b/nar00339-0123-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/fc98f18709ca/nar00339-0125-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/e7138e71739e/nar00339-0126-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/5271/320278/0641a26677b6/nar00339-0128-a.jpg

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