Schoenmakers E F, Geurts J M, Kools P F, Mols R, Huysmans C, Bullerdiek J, Van den Berghe H, Van de Ven W J
Laboratory for Molecular Oncology, Center for Human Genetics, University of Leuven, Belgium.
Genomics. 1995 Oct 10;29(3):665-78. doi: 10.1006/geno.1995.9952.
Cytogenetic analysis of a variety of benign solid tumors, among which uterine leiomyoma, lipoma, pleomorphic salivary gland adenoma, and pulmonary chondroid hamartoma, has indicated that these tumors often display chromosome breakpoints in region q13-q15 of chromosome 12. In previous studies, we have reported that these breakpoints map between locus D12S8 and the CHOP gene, the latter of which has been shown to be consistently rearranged in myxoid liposarcomas with t(12;16)(q13;p11). Here, we report directional chromosome walking studies starting from D12S8 and resulting in the construction of a YAC contig of about 6 Mb. This YAC contig, whose orientation on chromosome 12 was determined by double-color fluorescence in situ hybridization (FISH) analysis, has at least double coverage and consists of 75 overlapping YAC clones, all isolated from CEPH YAC libraries. Their insert sizes were estimated by contour-clamped homogeneous electric field (CHEF) gel electrophoresis. Chromosomal localization and chimerism of the YACs were investigated by FISH analysis. Chimerism of YAC clones was independently determined by restriction mapping. On the basis of YAC end-derived DNA markers and sequence-tagged sites (STSs), with an average spacing of approximately 70 kb, as well as restriction enzyme analysis, a long-range physical map was established for the 6-Mb DNA region of chromosome 12 covered by the YAC contig. Within the YAC contig, the relative positions of various known genes, an expressed sequence-tagged site, and a number of CEPH/Généthon polymorphic markers were determined. The latter data allow full integration of our mapping data with those obtained by CEPH/Généthon as well as those reported at the Second International Workshop on Human Chromosome 12 Mapping. Finally, this YAC contig constitutes the basis for the contstruction of a transcriptional map of this region and is likely to facilitate identification of genes involved in the formation of various benign solid tumor types.
对多种良性实体瘤进行的细胞遗传学分析表明，这些肿瘤（包括子宫平滑肌瘤、脂肪瘤、多形性涎腺腺瘤和肺软骨样错构瘤）常常在12号染色体的q13 - q15区域显示染色体断点。在先前的研究中，我们报道这些断点位于基因座D12S8和CHOP基因之间，后者已被证明在伴有t(12;16)(q13;p11)的黏液样脂肪肉瘤中持续发生重排。在此，我们报道了从D12S8开始的定向染色体步移研究，并构建了一个约6 Mb的酵母人工染色体（YAC）重叠群。通过双色荧光原位杂交（FISH）分析确定了该YAC重叠群在12号染色体上的方向，它至少有两倍覆盖率，由75个重叠的YAC克隆组成，所有克隆均从CEPH YAC文库中分离得到。通过轮廓夹钳均匀电场（CHEF）凝胶电泳估计了它们的插入片段大小。通过FISH分析研究了YAC的染色体定位和嵌合现象。通过限制性酶切图谱独立确定了YAC克隆的嵌合现象。基于平均间距约为70 kb的YAC末端衍生DNA标记和序列标签位点（STS）以及限制性酶切分析，为YAC重叠群覆盖的12号染色体6 Mb DNA区域建立了一个长距离物理图谱。在该YAC重叠群内，确定了各种已知基因、一个表达序列标签位点以及一些CEPH/Genethon多态性标记的相对位置。后一组数据使我们的图谱数据能够与CEPH/Genethon获得的数据以及在第二届人类12号染色体图谱国际研讨会上报道的数据完全整合。最后，这个YAC重叠群构成了构建该区域转录图谱的基础，并且可能有助于鉴定参与各种良性实体瘤类型形成过程的基因。
