Screening hypermethylated regions by methylation-sensitive single-strand conformational polymorphism.
Screening hypermethylated regions by methylation-sensitive single-strand conformational polymorphism.
复制标题
通过甲基化敏感的单链构象多态性筛选高甲基化区域。
DOI：
10.1006/abio.1999.4445
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发表时间：
2000
期刊：
影响因子：
--
通讯作者：
Jarrard,DF
中科院分区：
文献类型：
--
作者：
Kinoshita,H;Shi,Y;Sandefur,C;Jarrard,DF
DNA甲基化是一种重要的表观遗传修饰，可以改变含有CpG岛的基因的转录。在本报告中，我们证明了熟悉的技术单链构象多态性（SSCP）可以很容易地应用于亚硫酸处理的DNA，以检测大（~ 250 bp）富含cg的区域的甲基化差异。在甲基化敏感的SSCP （Ms-SSCP）技术中，亚硫酸氢钠修饰DNA在5-甲基胞嘧啶保持不变的条件下将未甲基化的胞嘧啶转化为尿嘧啶。然后用亚硫酸处理过的DNA特异性引物扩增待测区域。所得产物然后进行非变性凝胶分析。基于差异带迁移率，Ms-SSCP能够灵敏地检测甲基化密度的变化，并确定样品中是否存在未甲基化的等位基因。Ms-SSCP是一种快速而简单的技术，用于筛选多个样品的甲基化变化。与现有技术相比，它有几个优势，包括利用纳克量的DNA，避免测序cg富集区域的困难，以及筛选DNA大区域的多个甲基化位点。
DNA methylation is an important epigenetic modification that alters transcription in those genes containing CpG islands. In this report we demonstrate that the familiar technique single-strand conformation polymorphism (SSCP) can be easily applied to bisulfite-treated DNA to detect methylation differences over large (∼250 bp) CG-enriched regions. In the methylation-sensitive SSCP (Ms-SSCP) technique, sodium bisulfite modification of DNA converts unmethylated cytosine to uracil under conditions in which 5-methylcytosine remains unaltered. Amplification of these regions to be tested is then performed using primers specific for bisulfite-treated DNA. The resulting products are then subjected to nondenaturing gel analysis. Based on differential band mobility, Ms-SSCP is able to sensitively detect alterations in methylation density and determine if unmethylated alleles are present in a sample. Ms-SSCP is a rapid and simple technique for screening multiple samples for methylation changes. It has several advantages over existing techniques, including the utilization of nanogram amounts of DNA, the avoidance of difficulties in sequencing CG-enriched regions, and the screening of multiple sites for methylation across large regions of DNA.
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DOI：
10.1073/pnas.93.18.9821
发表时间：
1996-09-03
影响因子：
11.1
作者：
Herman, JG;Graff, JR;Baylin, SB
通讯作者：
Baylin, SB
影响因子：
14.9
作者：
Gonzalgo, ML;Jones, PA
通讯作者：
Jones, PA
影响因子：
--
作者：
S. Baylin;J. Herman;J. Graff;P. Vertino;J. Issa
通讯作者：
S. Baylin;J. Herman;J. Graff;P. Vertino;J. Issa
用于检测点突变的单链构象多态性（SSCP）技术的优化
影响因子：
3.9
作者：
D. Glavač;M. Dean
通讯作者：
M. Dean
影响因子：
11.2
作者：
D. Jarrard;H. Kinoshita;Yan Shi;C. Sandefur;Douglas G Hoff;L. Meisner;Chawnshang Chang;J. Herman;W. Isaacs;N. Nassif
通讯作者：
D. Jarrard;H. Kinoshita;Yan Shi;C. Sandefur;Douglas G Hoff;L. Meisner;Chawnshang Chang;J. Herman;W. Isaacs;N. Nassif