Capilla-Perez Laia, Solier Victor, Portemer Virginie, Chambon Aurelie, Hurel Aurelie, Guillebaux Alexia, Vezon Daniel, Cromer Laurence, Grelon Mathilde, Mercier Raphael
CNRS, Institut Jean-Pierre Bourgin, INRA, AgroParisTech, Université Paris-Saclay, Versailles, France.
Front Plant Sci. 2018 Sep 19;9:1339. doi: 10.3389/fpls.2018.01339. eCollection 2018.
Genetic screens have been crucial for deciphering many important biological processes, including meiosis. In , previous forward screens have likely identified almost all the meiotic genes that when mutated lead to a pronounced decrease in fertility. However, the increasing number of genes identified in reverse genetics studies that play crucial roles in meiosis, but do not exhibit strong phenotypes when mutated, suggests that there are still many genes with meiotic function waiting to be discovered. In this study, we produced 897 homozygous mutant lines using Ethyl Methyl Sulfonate (EMS) mutagenesis followed by either single seed descent or haploid doubling. Whole genome sequencing of a subset of lines showed an average of 696 homozygous mutations per line, 195 of which (28%) modify a protein sequence. To test the power of this library, we carried out a forward screen looking for meiotic defects by observing chromosomes at metaphase I of male meiosis. Among the 649 lines analyzed, we identified 43 lines with meiotic defects. Of these, 21 lines had an obvious candidate causal mutation, namely a STOP or splicing site mutation in a gene previously shown to play a role in meiosis (, and ). Interestingly, this was the first time that six of these genes were identified in a forward screen in (, , and ). These results illustrate the potential of this mutant population for screening for any qualitative or quantitative phenotype. Thus, this new mutant library is a powerful tool for functional genomics in . The HEM (Homozygote EMS Mutants) lines are available at the Versailles stock center.
遗传筛选对于破译包括减数分裂在内的许多重要生物学过程至关重要。在过去，正向筛选可能已经鉴定出了几乎所有在发生突变时会导致育性显著下降的减数分裂基因。然而，反向遗传学研究中鉴定出的在减数分裂中起关键作用但发生突变时不表现出强表型的基因数量不断增加，这表明仍有许多具有减数分裂功能的基因有待发现。在本研究中，我们使用甲基磺酸乙酯（EMS）诱变，随后通过单粒传或单倍体加倍产生了897个纯合突变系。对一部分品系进行全基因组测序显示，每个品系平均有696个纯合突变，其中195个（28%）会改变蛋白质序列。为了测试这个文库的效力，我们通过观察雄性减数分裂中期I的染色体进行了一次正向筛选，寻找减数分裂缺陷。在分析的649个品系中，我们鉴定出43个具有减数分裂缺陷的品系。其中，21个品系有一个明显的候选因果突变，即在先前已证明在减数分裂中起作用的基因中存在一个终止或剪接位点突变（以及）。有趣的是，其中六个基因是首次在正向筛选中被鉴定出来（以及）。这些结果说明了这个突变群体在筛选任何定性或定量表型方面的潜力。因此，这个新的突变文库是用于功能基因组学研究的强大工具。HEM（纯合子EMS突变体）品系可在凡尔赛种质中心获得。
