Williams Eli S, Cornforth Michael N, Goodwin Edwin H, Bailey Susan M
Department of Environmental & Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA.
Methods Mol Biol. 2011;735:113-24. doi: 10.1007/978-1-61779-092-8_11.
Fluorescence in situ hybridization (FISH) has become a powerful tool for exploring genomes at the level of chromosomes. The procedure can be used to identify individual chromosomes, rearrangements between chromosomes, and the location within a chromosome of specific DNA sequences such as centromeres, telomeres, and even individual genes. Chromosome orientation FISH (CO-FISH) extends the information obtainable from standard FISH to include the relative orientation of two or more DNA sequences within a chromosome (Goodwin and Meyne, Cytogenet Cell Genet 63:126-127, 1993). In combination with a suitable reference probe, CO-FISH can also determine the absolute 5'-3' direction of a DNA sequence relative to the short arm (pter) to long arm (qter) axis of the chromosome. This variation of CO-FISH was originally termed "COD-FISH" (Chromosome orientation and direction FISH) to reflect this fact (Meyne and Goodwin, Chromosome Research 3:375-378, 1995). Telomeric DNA serves as a convenient and absolute reference probe for this purpose, since all G-rich 5'-(TTAGGG)( n )-3' telomeric sequences are terminally located and oriented away from the centromere.In the beginning, CO-FISH was used to detect obligate chromosomal inversions associated with isochromosome formation (Bailey et al., Mutagenesis 11:139-144, 1996), various pericentric inversions (Bailey et al., Cytogenetics and Cell Genetics 75:248-253, 1996), and to confirm the origin of centromeric lateral asymmetry (Goodwin et al., Chromosoma 104:345-347, 1996). More recent and sophisticated applications of CO-FISH include distinction between telomeres produced via leading- vs. lagging-strand DNA synthesis (Bailey et al., Science 293:2462-2465, 2001), identification of interstitial blocks of telomere sequence that result from inappropriate fusion to double-strand breaks (telomere-DSB fusion) (Bailey et al., DNA Repair (Amst) 3:349-357, 2004), discovery of elevated rates of mitotic recombination at chromosomal termini (Cornforth and Eberle, Mutagenesis, 16:85-89, 2001) and sister chromatid exchange within telomeric DNA (T-SCE) (Bailey et al., Nucleic Acids Res 32:3743-3751, 2004), establishing replication timing of mammalian telomeres throughout S-phase (ReD-FISH) (Cornforth et al., In: Cold Spring Harbor Symposium: Telomeres and Telomerase, Cold Spring Harbor, NY, 2003; Zou et al., Proc Natl Acad Sci USA 101:12928-12933, 2004) and in combination with -spectral karyotyping (SKY-CO-FISH) (Williams et al., Cancer Res 69:2100-2107, 2009). For more information, the reader is referred to several reviews (Bailey et al., Cytogenet Genome Res 107, 14-17, 2004; Bailey and Cornforth, Cell Mol Life Sci 64:2956-2964, 2007; Bailey, Telomeres and Double-Strand Breaks - All's Well that "Ends" Well, Radiat Res 169:1-7, 2008).
荧光原位杂交（FISH）已成为在染色体水平探索基因组的强大工具。该方法可用于识别单个染色体、染色体之间的重排，以及特定DNA序列（如着丝粒、端粒甚至单个基因）在染色体上的位置。染色体定向FISH（CO-FISH）将从标准FISH可获得的信息扩展到包括染色体中两个或多个DNA序列的相对方向（Goodwin和Meyne，《细胞遗传学与细胞基因组学》63:126 - 127，1993）。与合适的参考探针结合，CO-FISH还可以确定DNA序列相对于染色体短臂（pter）到长臂（qter）轴的绝对5'-3'方向。CO-FISH的这种变体最初被称为“COD-FISH”（染色体定向和方向FISH）以反映这一事实（Meyne和Goodwin，《染色体研究》3:375 - 378，1995）。端粒DNA为此提供了方便且绝对的参考探针，因为所有富含G的5'-(TTAGGG)(n)-3'端粒序列都位于末端且远离着丝粒定向。起初，CO-FISH用于检测与等臂染色体形成相关的专性染色体倒位（Bailey等人，《诱变》11:139 - 144，1996）、各种臂间倒位（Bailey等人，《细胞遗传学与细胞遗传学》75:248 - 253，1996），并确认着丝粒侧向不对称的起源（Goodwin等人，《染色体》104:345 - 347，1996）。CO-FISH更新的复杂应用包括区分通过前导链与滞后链DNA合成产生的端粒（Bailey等人，《科学》293:2462 - 2465，2001）、鉴定因与双链断裂不适当融合（端粒 - DSB融合）导致的端粒序列间插片段（Bailey等人，《DNA修复（阿姆斯特丹）》3:349 - 357，2004）、发现染色体末端有丝分裂重组率升高（Cornforth和Eberle，《诱变》16:85 - 89，2001）以及端粒DNA内姐妹染色单体交换（T-SCE）（Bailey等人，《核酸研究》32:3743 - 3751，2004）、确定整个S期哺乳动物端粒的复制时间（ReD-FISH）（Cornforth等人，见：《冷泉港研讨会：端粒与端粒酶》，纽约冷泉港，2003；邹等人，《美国国家科学院院刊》101:12928 - 12933，2004）以及与光谱核型分析相结合（SKY-CO-FISH）（Williams等人，《癌症研究》69:2100 - 2107，2009）。欲了解更多信息，读者可参考几篇综述（Bailey等人，《细胞遗传学与基因组研究》107, 14 - 17, 2004；Bailey和Cornforth，《细胞与分子生命科学》64:2956 - 2964，2007；Bailey，《端粒与双链断裂——结局好一切都好》，《辐射研究》169:1 - 7，2008）。
