Beier D R, Dushkin H, Sussman D J
Division of Genetics, Brigham and Women's Hospital, Boston, MA 02115.
Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):9102-6. doi: 10.1073/pnas.89.19.9102.
We have utilized a PCR-based analysis of single-strand conformation polymorphisms to identify polymorphisms that can be used for mapping cloned DNA sequences in the mouse. We have found that single-strand conformation polymorphism analysis of sequences that are potentially less subject to conservation (i.e., intron and 3' untranslated regions) is a relatively efficient means of detecting polymorphisms between inbred strains. Fifty percent of the tested primer pairs were polymorphic between inbred strains and 90% were polymorphic between mouse species, which is a frequency comparable to that found for microsatellite repeat sequences. We have found that this technique can be readily used to determine the strain distribution pattern in a recombinant inbred series and is a simple and rapid means to obtain a map position for cloned sequences. When this strategy was tested on a number of previously mapped cloned genes, the strain distribution patterns obtained were consistent with that to be expected on the basis of the known map position. We also tested the utility of this approach for characterizing genes that have not been previously mapped. Dvl, the mouse homolog of the putative Drosophila dishevelled gene, and Adfp, encoding an adipocyte differentiation-related protein, were found to map to chromosome 4. These results were confirmed using single-strand conformation polymorphism analysis of an interspecific backcross.
我们利用基于聚合酶链反应（PCR）的单链构象多态性分析来鉴定可用于绘制小鼠克隆DNA序列图谱的多态性。我们发现，对潜在保守性较低的序列（即内含子和3'非翻译区）进行单链构象多态性分析是检测近交系之间多态性的一种相对有效的方法。50%的测试引物对在近交系之间存在多态性，90%在小鼠物种之间存在多态性，这一频率与微卫星重复序列的频率相当。我们发现，该技术可轻松用于确定重组近交系中的品系分布模式，是获得克隆序列图谱位置的一种简单快速的方法。当对一些先前已定位的克隆基因进行该策略测试时，获得的品系分布模式与基于已知图谱位置预期的模式一致。我们还测试了该方法用于表征先前未定位基因的效用。发现小鼠中假定的果蝇盘状基因的同源物Dvl以及编码脂肪细胞分化相关蛋白的Adfp定位于4号染色体。使用种间回交的单链构象多态性分析证实了这些结果。
