一种快速的突变体库创建方法

      突变体库是功能基因组学研究的有力武器。随着技术的发展，植物突变体库的创建方法已经非常丰富，比如：T-DNA或（反）转座子插入、EMS或辐射诱变、RNAi或基因编辑干涉。不同的方法都有着各自的优缺点。最新的《The Plant Cell》杂志报道了一个利用快中子（fast-neutron, FN）辐射和全基因组测序创建的水稻突变体库。

      快中子是核反应中未经过慢化剂慢化的一类中子，用其进行辐射诱变可以产生多样的突变类型，包括：碱基替换、片段缺失或插入、染色体倒位或异位、随机复制等，这也是它相对于其他突变方法的显著优势。研究人员将日本特早熟粳稻品种Kitaake（生育期只有9周）作为受体，利用快中子辐射创建了一个突变体库。对1504个突变株系进行45倍覆盖度的全基因组测序分析，鉴定了91513个突变位点，这些位点影响了32307个基因（占全部基因的58%）。研究人员将这些突变体信息整合成一个名为KitBase（http://kitbase.ucdavis.edu/）的生物信息数据库，可以查询突变基因和位点序列，查看突变表型。
      快中子辐射可以产生多样的突变类型（其中碱基替换的突变类型最多，其次是片段缺失），全基因组测序可以快速地鉴定突变位点， Kitaake品种极短的生育期有助于提升性状鉴定的速度。这样快速的突变体创建和鉴定方案对于植物功能基因组研究以及利用基因编辑技术进行品种改良来说意义重大。
 
The Plant Cell, 2017 June 2
The Sequences of 1,504 Mutants in the Model Rice Variety Kitaake Facilitate Rapid Functional Genomic Studies
Author
Guotian Li, Rashmi Jain, Mawsheng Chern……Kerrie W. Barry, Jeremy Schmutz and Pamela C. Ronald*
*: Departmentof Plant Pathology and the Genome Center, University of California, Davis; Lawrence Berkeley National Laboratory, USA
Abstract
The availability of a whole-genome sequenced mutant population and the cataloging of mutations of each line at a single-nucleotide resolution facilitate functional genomic analysis. To this end, we generated and sequenced a fast-neutron-induced mutant population in the model rice cultivar Kitaake (Oryza sativa L. ssp. japonica), which completes its life cycle in 9 weeks. We sequenced 1,504 mutant lines at 45-fold coverage and identified 91,513 mutations affecting 32,307 genes, i.e., 58% of all rice genes. We detected an average of 61 mutations per line. Mutation types include single base substitutions, deletions, insertions, inversions, translocations, and tandem duplications. We observed a high proportion of loss-of-function mutations. We identified an inversion affecting a single gene as the causative mutation for the short-grain phenotype in one mutant line. This result reveals the usefulness of the resource for efficient, cost-effective identification of genes conferring specific phenotypes. To facilitate public access to this genetic resource, we established an open access database called KitBase that provides access to sequence data and seed stocks. This population complements other available mutant collections and gene-editing technologies. This work demonstrates how inexpensive next-generation sequencing can be applied to generate a high-density catalog of mutations.