Navin A, Prekeris R, Lisitsyn N A, Sonti M M, Grieco D A, Narayanswami S, Lander E S, Simpson E M
The Jackson Laboratory, 600 Main Street, Bar Harbor, Maine, 04609, USA.
Genomics. 1996 Sep 1;36(2):349-53. doi: 10.1006/geno.1996.0473.
Representational difference analysis (RDA) was used to generate Y-specific probes by enriching for and cloning the differences between the male (XY) and the female (XX) C57BL/6J mouse genomes. Characterization of 35 clones revealed 12 families related by sequence similarity. One clone from each family was chosen for detailed analysis by Southern blot hybridization, polymerase chain reaction (PCR) on normal and aberrant genomes (Sxr), and fluorescence in situ hybridization. From one difference product we have characterized 12 Y-specific probes for hybridization, created seven male-specific PCR assays, mapped all repeat families, and identified one repeat with a distinct XY homology. We report the first cloning of a Y-specific long interspersed repeat element (LINE) fragment. In total, RDA has identified six novel Y Chromosome repeat families and allowed us to extend the characterization of six known Y repeats. We conclude that this novel use of RDA for whole chromosome subtraction successfully enriches chromosome-specific sequences and is suitable for the rapid generation of new Y Chromosome-specific probes.
代表性差异分析（RDA）用于通过富集并克隆雄性（XY）和雌性（XX）C57BL/6J小鼠基因组之间的差异来生成Y特异性探针。对35个克隆的特性分析揭示了12个因序列相似性而相关的家族。从每个家族中选择一个克隆，通过Southern印迹杂交、对正常和异常基因组（Sxr）进行聚合酶链反应（PCR）以及荧光原位杂交进行详细分析。从一个差异产物中，我们鉴定了12个用于杂交的Y特异性探针，创建了7种雄性特异性PCR检测方法，绘制了所有重复家族的图谱，并鉴定了一个具有独特XY同源性的重复序列。我们报道了首个Y特异性长散在重复元件（LINE）片段的克隆。总体而言，RDA已鉴定出6个新的Y染色体重复家族，并使我们能够扩展对6个已知Y重复序列的特性分析。我们得出结论，RDA这种用于全染色体消减的新用途成功富集了染色体特异性序列，适用于快速生成新的Y染色体特异性探针。
