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Restriction Landmark Genomic Analysis of Cancer

癌症的限制性标志基因组分析

基本信息

批准号：
7081297
负责人：
WILLIAM A HELD
金额：
$ 35.14万
依托单位：
ROSWELL PARK CANCER INSTITUTE CORP
依托单位国家：
美国
项目类别：
财政年份：
1995
资助国家：
美国
起止时间：
1995-04-01 至 2008-06-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The overall aim of this research is to identify targets of CpG island alterations associated with human colon tumorigenesis using Restriction Landmark Genomic Scanning (RLGS). Using Notl or other restriction enzymes that have GC rich recognition sequences as the restriction landmark, RLGS targets gene rich CpG island regions and primarily detects alterations in DNA methylation status although DNA deletions and amplifications can also be detected. RLGS loci exhibiting frequent alterations associated with human colon tumorigenesis will be cloned using in silico based, virtual RLGS computational methods. The confirmation of identity between the "virtual" RLGS sequence and the "real" RLGS sequence will be accomplished by PCR amplification of the RLGS spot DNA using primers derived from the virtual sequence. Array Comparative Genomic Hybridization (CGH) will be performed on the same tumors analyzed by RLGS to integrate epigenetic and genetic aberrations associated with human colon cancer. This will allow us to establish whether there is a class of tumors with high epigenetic instability that can be distinguished from those with high genetic instability and whether there is cooperative interaction between epigenetic and genetic aberrations. The methylation status of selected RLGS loci will be evaluated by methylation sensitive PCR and bisulfite sequencing to determine the boundaries and density of DNA methylation. Quantitative expression assays will be used to assess the expression of putative target genes in primary tumors to determine whether the CpG island alteration was associated with a change in gene expression. The identification of novel targets of CpG island alteration may uncover new pathways of tumorigenesis and further our understanding of how epigenetic alterations impact on the malignant phenotype.

描述（由申请人提供）：本研究的总体目标是利用限制性地标基因组扫描 (RLGS) 来确定与人类结肠肿瘤发生相关的 CpG 岛改变的靶标。使用NotI或其他具有富含GC的识别序列作为限制性标志的限制性内切酶，RLGS靶向富含基因的CpG岛区域，并且主要检测DNA甲基化状态的改变，尽管也可以检测DNA缺失和扩增。将使用基于计算机的虚拟 RLGS 计算方法来克隆表现出与人类结肠肿瘤发生相关的频繁改变的 RLGS 基因座。 “虚拟”RLGS序列和“真实”RLGS序列之间的同一性确认将通过使用源自虚拟序列的引物对RLGS点DNA进行PCR扩增来完成。将在 RLGS 分析的相同肿瘤上进行阵列比较基因组杂交 (CGH)，以整合与人类结肠癌相关的表观遗传和遗传畸变。这将使我们能够确定是否有一类具有高度表观遗传不稳定性的肿瘤可以与具有高度遗传不稳定性的肿瘤区分开来，以及表观遗传和遗传畸变之间是否存在协同相互作用。选定的 RLGS 位点的甲基化状态将通过甲基化敏感 PCR 和亚硫酸氢盐测序进行评估，以确定 DNA 甲基化的边界和密度。定量表达测定将用于评估原发性肿瘤中推定靶基因的表达，以确定 CpG 岛改变是否与基因表达的变化相关。 CpG 岛改变的新靶标的鉴定可能会揭示肿瘤发生的新途径，并进一步加深我们对表观遗传改变如何影响恶性表型的理解。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Identification of a novel member of the snail/Gfi-1 repressor family, mlt 1, which is methylated and silenced in liver tumors of SV40 T antigen transgenic mice.

鉴定出 snail/Gfi-1 阻遏蛋白家族的新成员 mlt 1，该蛋白在 SV40 T 抗原转基因小鼠的肝肿瘤中被甲基化和沉默。

DOI：
发表时间：
2001
期刊：
Cancer research
影响因子：
11.2
作者：
Tateno,M;Fukunishi,Y;Komatsu,S;Okazaki,Y;Kawai,J;Shibata,K;Itoh,M;Muramatsu,M;Held,WA;Hayashizaki,Y
通讯作者：
Hayashizaki,Y

An arrayed human not I-EcoRV boundary library as a tool for RLGS spot analysis.

阵列人类非 I-EcoRV 边界库作为 RLGS 点分析的工具。

DOI：
10.1093/dnares/4.3.253
发表时间：
1997
期刊：
DNA research : an international journal for rapid publication of reports on genes and genomes
影响因子：
0
作者：
Plass,C;Weichenhan,D;Catanese,J;Costello,JF;Yu,F;Yu,L;Smiraglia,D;Cavenee,WK;Caligiuri,MA;deJong,P;Held,WA
通讯作者：
Held,WA

Restriction landmark genomic scanning of mouse liver tumors for gene amplification: overexpression of cyclin A2.

小鼠肝脏肿瘤的限制性标志基因组扫描以进行基因扩增：细胞周期蛋白 A2 的过度表达。