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ChIA-PET法（、ペアエンドタグの配列解析によるクロマチン相互作用解析）は、クロマチン免疫沈降 (ChIP)を元にした濃縮、 Chromosome conformation capture(訳語未定), Paired-End Tags(ペアエンドタグ), および ultra-high-throughput sequencing(DNAシークエンシングおよび遺伝子参照) の組み合わせによって、染色体に含まれるDNA塩基配列の相互作用を全染色体にわたって決定するために開発された手法(Fullwood & Yijun, 2009)である。 遺伝子は、プロモーターなどの制御領域、インシュレーターなどの境界領域や転写因子結合領域(TFBS)など離れた領域からの制御も受けるばあいもある。制御領域と遺伝子本体の領域の動的相互作用を明らかにすることには、医学などの立場からみると遺伝子の働きの制御の本質的に重要な部分を理解する事になるという重要性がある(Maston et al., 2006)(訳注: 原文は、"Uncovering the interplay between regulatory regions and gene coding regions is essential for understanding the mechanisms governing gene regulation in health and disease")。 ChIA-PETは、染色体上での近接遠隔を問わない、TFBSやプロモーターと遺伝子本体の、他の手法では出来ない機能的な相互作用の同定が可能である。 ChIA-PETはまた、細胞のcell differentiation(分化), proliferation(細胞分裂), and 胚発生などの過程で働いている機構を明らかにする場合にも用いる事ができる。DNA結合性転写因子タンパクやプロモーター領域に対してChIA-PET interactomeマップを作成する事で、治療介入(therapeutic intervention)においてより良い標的を見つける事が可能である(Fullwood & Yijun, 2009)。 ==手法== ChIA-PET法は、クロマチン免疫沈降法(Kuo & Allis, 1999)および3C法を組み合わせた手法である(The ChIA-PET method combines ChIP-based methods (Kuo & Allis, 1999), and Chromosome_conformation_capture (3C), to extend the capabilities of both approaches.)。 ChIP-Seq法が転写因子結合領域(TFBS)を決定するのに広く用いられる手法である一方、染色体間の大域的相互作用の測定には3C法が用いられていた(Dekker et al., 2002)(ChIP-Sequencing (ChIP-Seq) is a popular method used to identify TFBS while 3C has been used to identify long-range chromatin interactions (Dekker et al., 2002).)。 However, both suffer from limitations when used independently to identify de-novo long-range interactions genome wide. While ChIP-Seq is typically used for genome-wide identification of TFBS (Barski et al., 2007; Wei et al., 2006), it provides only linear information of protein binding sites along the chromosomes (but not interactions between them), and suffers from high genomic background noise (false positives). Additionally, only a small amount of sequences generated by ChIP-Seq uniquely map to the genome, and an even smaller amount are functional TFBS (Johnson et al., 2007). While 3C is capable of analyzing long-range chromatin interactions, it cannot be used genome wide and, like ChIP-Seq, also suffers from high levels of background noise. Since the noise increases in relation to the distance between interacting regions (max 100kb), laborious and tedious controls are required for accurate characterization of chromatin interactions (Dekker et al., 2006). The ChIA-PET method successfully resolves the issues of non-specific interaction noise found in ChIP-Seq by sonicating the chip fragments in order to separate random attachments from specific interaction complexes. The next step, which is referred to as enrichment, reduces complexity for genome-wide analysis and adds specificity to chromatin interactions bound by pre-determined TFs (transcription factors). The ability of 3C approaches to identify long-range interactions is based on the theory of proximity ligation. In regards to DNA inter-ligation, fragments that are tethered by common protein complexes have greater kinetic advantages under dilute conditions, than those freely diffusing in solution or anchored in different complexes. ChIA-PET takes advantage of this concept by incorporating linker sequences onto the free ends of the DNA fragments tethered to the protein complexes. In order to build connectivity of the fragments tethered by regulatory complexes, the linker sequences are ligated during nuclear proximity ligation. Therefore, the products of linker-connected ligation can be analyzed by ultra-high-throughput PET sequencing and mapped to the reference genome. Since ChIA-PET is not dependent on specific sites for detection as 3C and 4C are, it allows unbiased, genome-wide de-novo detection of chromatin interactions (Fullwood et al., 2009). ==Workflow== 抄文引用元・出典: フリー百科事典『 ウィキペディア（Wikipedia）』 ■ウィキペディアで「ChIA-PET」の詳細全文を読む スポンサード リンク
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