Lü X C, Montelius-Alatalo K, Helou K, Klinga-Levan K, Islam Q, Levan G, Röhme D
Department of Genetics, Lundberg Laboratory, Göteborg University, Sweden.
Somat Cell Mol Genet. 1997 Jan;23(1):63-74. doi: 10.1007/BF02679956.
We have applied the representational difference analysis (RDA) to isolate genetic markers for a deletion on the rat chromosome RNO5q22-33. This deletion occurred in anchorage independent sublines of a normal rat fibroblast x mouse hepatoma cell hybrid (BS181) (Islam 1989). Normal rat tissue DNA provided the "tester" and the BS181 hybrid DNA the "driver" in the RDA hybridization/selection reactions. Out of twelve RDA derived DNA sequences that were analyzed in detail using a rat X mouse cell hybrid panel for chromosome mapping, nine (75%) were found to represent RNO5 deletions, whereas the other three were new RFLPs mapping to other chromosomes. In two cases, the RDA sequences were also analyzed by fluorescence in situ hybridization (FISH) and found to give distinct signals in the RNOq22-33 region. This result emphasizes teh significance of the previous cytogenetic analysis of this hybrid, which indicated the presence of a gene for the suppression of anchorage independence, Sai 1, in this deletion region. The RDA derived sequences isolated by this work will provide a valuable source of new genetic markers for the further detailed analysis of the Sai 1 deletion region.
我们应用代表性差异分析（RDA）来分离大鼠染色体RNO5q22 - 33上缺失区域的遗传标记。这种缺失发生在正常大鼠成纤维细胞x小鼠肝癌细胞杂交体（BS181）的非锚定依赖性亚系中（Islam，1989）。在RDA杂交/选择反应中，正常大鼠组织DNA作为“检测者”，BS181杂交体DNA作为“驱动者”。在使用大鼠X小鼠细胞杂交面板进行染色体定位的详细分析的12个RDA衍生DNA序列中，有9个（75%）被发现代表RNO5缺失，而另外3个是定位到其他染色体的新的限制性片段长度多态性（RFLP）。在两个案例中，RDA序列也通过荧光原位杂交（FISH）进行了分析，发现在RNOq22 - 33区域产生了明显的信号。这一结果强调了对该杂交体先前细胞遗传学分析的重要性，该分析表明在这个缺失区域存在一个抑制非锚定依赖性的基因Sai 1。这项工作分离出的RDA衍生序列将为进一步详细分析Sai 1缺失区域提供新的遗传标记的宝贵来源。
