High-resolution, genome-scale mapping of natural variation between reproductively isolated individuals

摘要 遗传学的主要目标之一是了解自然变异的个体如何以及为什么不同 表型。绘制这种性状变异的分子基础可能很简单，但只有在 可以进行杂交。我们发现，一种很少被研究的蠕虫物种--细小线虫--有一个中位数 寿命比相关线虫长50%，我们观察到 酿酒酵母是其分支中唯一可以充当机会性病原体的物种。在 目前的提议，我们使用大规模平行版本的倒数来剖析这些物种的分歧 半合子试验。我们通过产生有活力的、不育的F1来创造半合子突变的基因组 物种间的杂交并使它们接受转座子诱变。我们把半合子汇集在一起，测量 它们的寿命和适合性在基于测序的分析中，并测试克隆之间的频率差异 含有给定基因的双亲等位基因的池。结果是一个基因座目录，在这些基因座上 物种间的差异会影响兴趣特征。在我们使用酵母(目标1)和蠕虫(目标2)的原理证明中， 我们将发现野生物种中出现的等位基因，以提高健康寿命和抗应激能力。这些 结果将与削弱适合度的等位基因形成鲜明对比，这些等位基因通常在种内研究中被映射。 我们酵母基因的同源基因作为流行真菌毒力基因的候选者将立即引起人们的兴趣 病原体和我们的蠕虫寿命因子的同源基因将非常适合于分析 哺乳动物。这项工作将是有史以来第一次对遗传结构和分子进行全面的调查 生殖隔离个体间性状变异的遗传学。而我们首创的方法将使 任何真核系统中的遗传解剖，在该系统中，F1杂交体或其组织是 可用。

SUMMARY One of the primary goals of genetics is to understand how and why naturally varying individuals differ in phenotype. Mapping the molecular basis of this trait variation can be straightforward, but only in individuals that can be interbred. We have found that a little-studied worm species, Caenorhabditis latens, has a median lifespan 50% longer than related nematodes, and we have observed robust, conserved thermotolerance in Saccharomyces cerevisiae, the only species of its clade that can act as an opportunistic pathogen. In the current proposal, we dissect these species divergences using a massively parallel version of the reciprocal hemizygote test. We create a genomic complement of hemizygote mutants, by generating viable, sterile F1 hybrids between species and subjecting them to transposon mutagenesis. We pool the hemizygotes, measure their longevity and fitness in sequencing-based assays, and test for differences in frequency between clones of the pool bearing the two parents' alleles of a given gene. The result is a catalog of loci at which variants between species influence the trait of interest. In our proof of principle using yeast (Aim 1) and worm (Aim 2), we will uncover alleles that have arisen in wild species to boost healthspan and stress resistance. These results will stand in contrast to the alleles weakening fitness that are often mapped in intra-specific studies. Orthologs of our yeast loci will be of immediate interest as candidates for virulence genes in prevalent fungal pathogens, and orthologs of our worm lifespan factors will be well-suited to analysis for anti-aging effects in mammals. This effort will be the first-ever comprehensive survey of the genetic architecture and molecular genetics of trait variation between reproductively isolated individuals. And the methods we pioneer will enable genetic dissection in any eukaryotic system in which banks of F1 hybrid individuals or their tissues are available.