CLONING THE X-LINKED RETINITIS PIGMENTOSA GENES

X连锁视网膜色素变性是最严重的赫雷德色素变性之一， 人类失明的原因，但它的生物化学基础仍然是单一的， 知道的 最近的连锁数据表明存在两个不同的基因座， 两个区域化到X染色体的近端短臂：一个在 Xp（RP3）和另一个更近端连接到DXS7（RP2）。 本研究 该提案旨在克隆和表征RP3基因，并定义 进一步的RP2基因座的染色体定位。 以隔离 RP3基因，基因组克隆最近获得的区域之间的 将进一步分析CGD位点和JBB，以确定保守的 指示含外显子序列的序列，并检测 在不相关的男性的DNA面板微缺失或重排， 通过Southern印迹分析或脉冲场凝胶电泳进行XRP。 定位在RP3基因座附近或RP3基因座内的其它克隆将是异源的。 通过从最近的侧翼开始的双向染色体步移 噬菌体文库中的标记和通过分析已经获得的酵母 人工染色体（YAC）克隆含有来自该基因的序列， 地区 检测保守序列的克隆将用于分离 候选RP3 cDNA，其将进一步通过北方 分析和测序。 一旦发现了候选序列， 未来的努力将针对识别RP3突变 基因座，阐明基因的结构组织和开发 研究RP3细胞和生化特性的试剂 蛋白 为了更精确地定位RP2位点， 将连接DXS7和DXS255之间区域的限制性图谱， 确定HTF岛屿将被开发，目的是确定 包含RP2基因的限制性片段。

X-linked Retinitis Pigmentosa is one of the most sever e of the hered- itary causes of blindness in man, yet its biochemical basis remains un- known. Recent linkage data indicate the existence of two distinct loci, both regionalized to the proximal short arm of the X chromosome: one at Xp (RP3) and the other more proximal linked to DXS7 (RP2). This research proposal aims to clone and characterize the gene for RP3 and define further the chromosomal localization of the RP2 locus. To isolate the RP3 gene, genomic clones recently obtained from the region between the CGD locus and JBB will be analyzed further in order to identify conserved sequences indicative of exon containing sequences and to detect either microdeletions or rearrangements in DNA panels of unrelated males with XRP by either Southern blot analysis or pulse field gel electrophoresis. Additional clones that map close to or within the RP3 locus will be iso- lated by bidirectional chromosomal walking from the nearest flanking markers in phage libraries and by analysis of already obtained Yeast Artificial Chromosome (YAC) clones that contain sequences from this region. Clones that detect conserved sequences will be used to isolate candidate RP3 cDNAs which will be further characterized by Northern analysis and sequencing. Once a candidate sequence had been found, future efforts will be targeted towards identifying mutations at the RP3 locus, elucidating the structural organization of the gene and developing reagents to study the cellular and biochemical properties of the RP3 protein. To localize more precisely the RP2 locus a detailed long range restriction map that will link up regions between DXS7 and DXS255 and identify HTF islands will be developed with the aim of identifying the restriction fragment that contains the RP2 gene.