Hozier J, Graham R, Westfall T, Siebert P, Davis L
Applied Genetics Laboratories, Inc., Melbourne, Florida 32901.
Genomics. 1994 Feb;19(3):441-7. doi: 10.1006/geno.1994.1092.
We have developed preparative in situ hybridization (Prep-ISH) of complex DNA populations to mitotic chromosomes as a means of generating chromosome region-specific DNA subpopulations. Prep-ISH is a combination of two cytogenetic techniques: in situ hybridization of DNA molecules to mitotic chromosomes and chromosome microdissection. Here, we present test cases demonstrating the feasibility of this approach on mouse and human genomes, using single nuclei, single chromosomes, or single chromosomal subregions to assess sensitivity, specificity, and representation of the Prep-ISH technique. Prep-ISH has a number of applications in studies of gene expression and genome organization, including efficient cytogenetic sorting of tissue-specific cDNAs and genomic DNA libraries. In addition, Prep-ISH is likely to dramatically reduce the number of candidate genes to aid in gene discovery efforts and to improve efficiency of developing transcription maps and YAC and cosmid contigs through defined cytogenetic regions.
我们已开发出针对有丝分裂染色体的复杂DNA群体的制备性原位杂交（Prep-ISH）技术，以此来生成染色体区域特异性的DNA亚群体。Prep-ISH是两种细胞遗传学技术的结合：DNA分子与有丝分裂染色体的原位杂交以及染色体显微切割。在此，我们展示了测试案例，利用单细胞核、单条染色体或单个染色体亚区域来评估Prep-ISH技术的敏感性、特异性和代表性，从而证明该方法在小鼠和人类基因组上的可行性。Prep-ISH在基因表达和基因组组织研究中有诸多应用，包括对组织特异性cDNA和基因组DNA文库进行高效的细胞遗传学分选。此外，Prep-ISH很可能会大幅减少候选基因的数量，有助于基因发现工作，并提高通过特定细胞遗传学区域绘制转录图谱以及构建酵母人工染色体（YAC）和黏粒重叠群的效率。
