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目的:研究γ射线对水稻基因组的诱变效应,明确其诱发突变的类型、分布和频率。创新点:首次针对种子繁殖植物在全基因组范围及单核苷酸水平上揭示了γ射线诱发可遗传变异的频率与特征。方法:利用Illumina Hiseq2000对三种γ射线剂量辐照培育的6株水稻(日本晴)M2植株进行基因组重测序,生物信息学分析确定单碱基替换(SBS)和插入缺失(Indel)突变,以及结构变异和拷贝数等变异的频率和基因组分布。利用Sanger测序、目标片段扩增或定量多聚酶链反应(qP CR)对各类突变进行验证。综合重测序和验证结果估算诱发突变频率。结论:结果表明,γ射线既可以诱发单碱基替换,也可以诱发插入缺失突变和结构变异;水稻M2代植株中的平均突变频率达到7.5×10~(-6)~9.8×10~(-6);Indel突变频率约为SBS变异的1/4,而结构变异频率更低;SBS和Indel突变随机分布在12条染色体上,无明显的突变热点。
OBJECTIVE: To study the mutagenic effects of γ-rays on rice genome and to determine the type, distribution and frequency of induced mutations. Innovation: For the first time, the frequency and characteristics of γ-ray-induced heritable mutations in seed-reproducing plants at the genome-wide and at the single-nucleotide level are revealed. METHODS: Six rice (Nipponbare) M2 plants were irradiated with three γ-ray doses by Illumina Hiseq 2000 for genome re-sequencing. Bioinformatic analysis identified single base substitution (SBS) and indel mutations, as well as structure Variation and copy number variation and genomic distribution. Various mutations were validated using Sanger sequencing, target fragment amplification or quantitative polymerase chain reaction (qP CR). Comprehensive re-sequencing and validation results to estimate the frequency of mutation. The results showed that γ ray can induce single base substitution and induce deletion mutation and structural variation. The average frequency of mutation in M2 plants reached 7.5 × 10 -6 ~ 9.8 × 10 ~ (-1) 6). The frequency of Indel mutation was about 1/4 of that of SBS mutation, but the frequency of structural variation was lower. The SBS and Indel mutations were randomly distributed on 12 chromosomes with no obvious mutation hot spots.