DETECTING AND MAPPING SEQUENCE CHANGES IN MOUSE DNA

计划使用小鼠肾脏RNA的cdna克隆文库来鉴定 近交系小鼠DNA序列的遗传变异。变种 将通过杂交标记的含有cDNAs的克隆质粒DNA来识别 从小鼠肝脏中插入限制性的、电泳化的DNA 转移到硝化纤维滤纸或DMB纸上。DNA的来源将会 是5组重组自交系的7个祖先株。 每个探针将杂交到一组具有以下特征的限制性片段 通过硝化棉的放射自显影可以看到其相对分子质量 过滤器。如果不存在变种，每个菌株都会表现出相同的一套分子 对给定探测器的碎片进行加权。如果存在可识别的变量，则 一个或多个限制性片段的相对分子质量将会改变。 这些变异将使用适当的一组 重组近交系(RI)菌株。进一步的映射可能涉及使用适当的 合适的亲本品系间的同源品系和杂交后代。 使用重组自交系作为作图工具，目前有40% 有机会立即映射一个新的基因座，所有的基因座最终都会收到 地图位置。一旦限制片段长度中的变体被映射，它们 将成为定位其他基因的重要参考基因座。50%的分析 设想每年克隆一次cDNA。其中许多都会检测到变异。我们 能够检测到测试了第一个克隆的变体，从而预测了一个 使用这种方法的新变种的快速繁殖。

It is planned to use a library of cDNA clones from mouse kidney RNA to identify genetic variation in the DNA sequence of the inbred mouse strains. The variants will be recognized by hybridizing labeled cloned plasmid DNA's containing cDNA inserts to restricted, electrophoresed DNA from mouse livers which has been transferred to nitrocellulose filters or to DMB-paper. The source of DNA will be the seven progenitor strains of five sets of recombinant inbred (RI) strains. Each probe will hybridize to a set of restriction fragments with characteristic molecular weight, which can be seen by autoradiography of the nitrocellulose filters. If no variant exists each strain will show the same set of molecular weight fragments for a given probe. If a recognizable variant exists, the molecular weight of one or more of the restriction fragments will be altered. Such variants will be analyzed genetically using the appropriate set of recombinant inbred (RI) strains. Further mapping may involve use of appropriate congenic strains and progeny of crosses between appropriate parental strains. The use of recombinant inbred strains as a mapping tool currently gives a 40% chance of immediately mapping a new locus, and all loci will eventually receive map positions. Once the variants in restriction fragment length are mapped they will become important reference loci for mapping other genes. Analysis of 50 cDNA clones per year is envisioned. Many of these will detect variation. We were able to detect a variant with the first clone tested and thus project a rapid proliferation of new variants using this method.