We developed bulked segregant analysis as a method for rapidly identifying markers linked to any specific gene or genomic region. Two bulked DNA samples are generated from a segregating population from a single cross. Each pool, or bulk, contains individuals that are identical for a particular trait or genomic region but arbitrary at all unlinked regions. The two bulks are therefore genetically dissimilar in the selected region but seemingly heterozygous at all other regions. The two bulks can be made for any genomic region and from any segregating population. The bulks are screened for differences using restriction fragment length polymorphism probes or random amplified polymorphic DNA primers. We have used bulked segregant analysis to identify three random amplified polymorphic DNA markers in lettuce linked to a gene for resistance to downy mildew. We showed that markers can be reliably identified in a 25-centimorgan window on either side of the targeted locus. Bulked segregant analysis has several advantages over the use of near-isogenic lines to identify markers in specific regions of the genome. Genetic walking will be possible by multiple rounds of bulked segregation analysis; each new pair of bulks will differ at a locus identified in the previous round of analysis. This approach will have widespread application both in those species where selfing is possible and in those that are obligatorily outbreeding.

译文

我们开发了大量的分离物分析,作为一种快速识别与任何特定基因或基因组区域相关的标记的方法。从单个杂交的分离种群中产生两个大量的DNA样本。每个池或整体包含对于特定性状或基因组区域相同但在所有未连接区域任意的个体。因此,这两个大块在选定区域在遗传上是不同的,但在所有其他区域似乎是杂合的。这两个大块可以用于任何基因组区域和任何分离的种群。使用限制性片段长度多态性探针或随机扩增的多态性DNA引物筛选大块的差异。我们已经使用大量的分离物分析来鉴定生菜中的三个随机扩增的多态性DNA标记,这些标记与抗霜霉病的基因相关。我们表明,可以在目标基因座两侧的25厘摩根窗口中可靠地识别出标记。与使用近等基因系鉴定基因组特定区域中的标记相比,大量分离物分析具有多个优势。通过多轮膨胀的分离分析,遗传行走是可能的; 每对新的大块在上一轮分析中确定的基因座上会有所不同。这种方法将在可能自交的物种和强制近交的物种中得到广泛应用。

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