POSITIONAL CLONING OF MEN1 GENE

这个项目的目的是鉴定基因和突变。 对常染色体显性遗传病MEN1的易感性负责 与11q13上的标记连锁。我们使用间期的努力-FISH，躯体 和减少辐射的细胞杂交以及基因组的构建 克隆重叠群使我们能够生成2.5 Mb的物理地图 由标记D11S480和D11S913定义的MEN1间隔。我们有 构建了YAC重叠群、细菌克隆(PAC/BAC/P1)重叠群和 该基因组区域的粘粒重叠群。开发了近100个STSS 主要来自这些克隆的末端序列使我们能够 生成高密度的STS内容地图。该重叠群提供模板 用于测序、转录鉴定和转录精细定位。 从这些克隆中分离出的多态标记在我们的 继续努力缩小杂合性缺失(LOH)的间隔 MEN1肿瘤的研究及MEN1连锁不平衡研究 血统。我们目前的MEN1间隔是1.3 Mb(PYGM-D11S913)。一个 成绩单地图(目前接近20份成绩单，其中11份已知 基因)正在为1.3MB的MEN1区域建造中。目前正在努力 以获得每个转录本的全长cdna序列并 分析它们的表达和外显子-内含子的组织。到目前为止，有6个 对20个MEN1患者的转录本进行了突变分析 Southern印迹、Northern印迹和SSCP/DDF方法，剔除AS MEN1的候选基因。

The purpose of this project is to identify the gene and the mutations responsible for the predisposition to MEN1, an autosomal dominant disease linked to markers at 11q13. Our efforts using interphase-FISH, somatic and radiation-reduced cell hybrids as well as construction of genomic clone contigs have allowed us to generate a physical map for the 2.5 Mb MEN1 interval defined by the markers, D11S480 and D11S913. We have constructed a YAC contig, a bacterial clone (PAC/BAC/P1) contig and a cosmid contig for this genomic region. Nearly 100 STSs developed primarily from the end sequences of these clones have allowed us to generate a high-density STS-content map. This contig provides templates for sequencing, transcript identification, and transcript fine mapping. Polymorphic markers isolated from these clones have been helpful in our continued efforts to narrow the interval by LOH (loss of heterozygo-sity) studies in MEN1 tumors and linkage disequilibrium studies in MEN1 pedigree. Our current MEN1 interval is 1.3 Mb (PYGM-D11S913). A transcript map (currently nearing 20 transcripts including 11 known genes) is under construction for the 1.3MB MEN1 region. Efforts are being made to obtain full length cDNA sequence of each transcript and to analyze their expression and exon-intron organization. So far, 6 of the 20 transcripts have been analyzed for mutations in MEN1 patients by Southern blot, Northern blot and by SSCP/DDF methods, and eliminated as candidate genes responsible for MEN1.