由于缺少在全基因組范圍內(nèi),，對(duì)5-羥甲基胞嘧啶（5hmC）進(jìn)行單個(gè)堿基水平分辨率的作圖手段，對(duì)5hmC的研究一直進(jìn)展緩慢,。5月17日Cell雜志在線發(fā)表了Chuan He研究組的研究論文“Base-Resolution Analysis of 5-Hydroxymethylcytosine in the Mammalian Genome”將這一研究方向推進(jìn)了一大步,。
傳統(tǒng)的親和純化手段不能準(zhǔn)確定位5hmC，也不能確定它在每一修飾位點(diǎn)的相對(duì)豐度,。Chuan He等人運(yùn)用Tet輔助的重亞硫酸鹽測(cè)序（TAB-Seq）技術(shù),，與傳統(tǒng)的重亞硫酸鹽測(cè)序技術(shù)結(jié)合,在基因組范圍內(nèi)以單個(gè)堿基的分辨率對(duì)5hmC進(jìn)行作圖，并確定5hmC與5mC的相對(duì)豐度,。
將這一技術(shù)應(yīng)用于胚胎干細(xì)胞,，他們不僅證實(shí)了5hmC在哺乳動(dòng)物基因組中廣泛分布，而且顯示5hmC位點(diǎn)更易于出現(xiàn)在某些特殊序列當(dāng)中,，并具有在兩條DNA鏈分布的不對(duì)稱性,。
他們還發(fā)現(xiàn)鄰近轉(zhuǎn)錄因子結(jié)合的DNA位點(diǎn)存在高水平的5hmC和低水平的5mC。此外,，5hmC分布水平在不同功能序列元件之間差異也很大,。這提示5hmC的修飾和保持在這些元件之間的機(jī)制是不同的。（生物谷Bioon.com）
doi:10.1016/j.cell.2012.04.027
Base-Resolution Analysis of 5-Hydroxymethylcytosine in the Mammalian Genome
Miao Yu, Gary C. Hon, Keith E. Szulwach et al.
Summary
The study of 5-hydroxylmethylcytosines (5hmC) has been hampered by the lack of a method to map it at single-base resolution on a genome-wide scale. Affinity purification-based methods cannot precisely locate 5hmC nor accurately determine its relative abundance at each modified site. We here present a genome-wide approach, Tet-assisted bisulfite sequencing (TAB-Seq), that when combined with traditional bisulfite sequencing can be used for mapping 5hmC at base resolution and quantifying the relative abundance of 5hmC as well as 5mC. Application of this method to embryonic stem cells not only confirms widespread distribution of 5hmC in the mammalian genome but also reveals sequence bias and strand asymmetry at 5hmC sites. We observe high levels of 5hmC and reciprocally low levels of 5mC near but not on transcription factor-binding sites. Additionally, the relative abundance of 5hmC varies significantly among distinct functional sequence elements, suggesting different mechanisms for 5hmC deposition and maintenance.
