A sorghum (<i>Sorghum bicolor</i>) mutant with altered carbon isotope ratio

Recent efforts to engineer C4 photosynthetic traits into C3 plants such as rice demand an understanding of the genetic elements that enable C4 plants to outperform C3 plants. As a part of the C4 Rice Consortium’s efforts to identify genes needed to support C4 photosynthesis, EMS mutagenized sorghum populations were generated and screened to identify genes that cause a loss of C4 function. Stable carbon isotope ratio (δ13C) of leaf dry matter has been used to distinguishspecies with C3 and C4 photosynthetic pathways. Here, we report the identification of a sorghum (Sorghum bicolor) mutant with a low δ13C characteristic. A mutant (named Mut33) with a pale phenotype and stunted growth was identified from an EMS treated sorghum M2 population. The stable carbon isotope analysis of the mutants showed a decrease of 13C uptake capacity. The noise of random mutation was reduced by crossing the mutant and its wildtype (WT). The back-cross (BC1F1) progenies were like the WT parent in terms of 13C values and plant phenotypes. All the BC1F2 plants with low δ13C died before they produced their 6th leaf. Gas exchange measurements of the low δ13C sorghum mutants showed a higher CO2 compensation point (25.24 μmol CO2.mol-1air) and the maximum rate of photosynthesis was less than 5μmol.m-2.s-1. To identify the genetic determinant of this trait, four DNA pools were isolated; two each from normal and low δ13C BC1F2 mutant plants. These were sequenced using an Illumina platform. Comparison of allele frequency of the single nucleotide polymorphisms (SNPs) between the pools with contrasting phenotype showed that a locus in Chromosome 10 between 57,941,104 and 59,985,708 bps had an allele frequency of 1. There were 211 mutations and 37 genes in the locus, out of which mutations in 9 genes showed non-synonymous changes. This finding is expected to contribute to future research on the identification of the causal factor differentiating C4 from C3 species that can be used in the transformation of C3 to C4 plants.当前将C4光合性状改造至水稻等C3植物中的研究，亟需阐明赋予C4植物优于C3植物性能的遗传基础。作为C4水稻联盟（C4 Rice Consortium）旨在鉴定支持C4光合所需基因的研究内容之一，本研究构建并筛选了EMS（乙基甲磺酸酯）诱变的高粱种群，以鉴定导致C4功能丧失的基因。植物叶片干物质的稳定碳同位素比值（δ¹³C）常被用于区分具备C3与C4光合途径的物种。本研究报道了一株低δ¹³C特性的高粱（Sorghum bicolor）突变体：研究人员从EMS诱变处理的高粱M₂群体中筛选得到一株命名为Mut33的突变体，其表现出白化表型与生长迟滞特征。对该突变体的稳定碳同位素分析显示，其¹³C吸收能力有所下降。通过将该突变体与其野生型（wildtype, WT）亲本杂交，降低了随机诱变带来的背景噪声。回交（back-cross, BC₁F₁）后代的碳同位素比值与植株表型均与野生型亲本一致。所有携带低δ¹³C特性的BC₁F₂代植株均在长出第6片叶前死亡。对低δ¹³C高粱突变体的气体交换测定结果显示，其CO₂补偿点更高（25.24 μmol CO₂·mol⁻¹空气），且最大光合速率低于5 μmol·m⁻²·s⁻¹。为鉴定该性状的遗传决定因子，研究人员分离得到4个DNA混合池：其中2个来自表型正常的BC₁F₂植株，另外2个来自低δ¹³C的突变体BC₁F₂植株。采用Illumina测序平台对上述混合池进行测序。通过对比表型存在差异的两个混合池的单核苷酸多态性（single nucleotide polymorphisms, SNPs）等位基因频率，发现10号染色体上57,941,104至59,985,708 bp区间内的一个位点的等位基因频率为1。该区间内共存在211处突变与37个基因，其中9个基因的突变呈现非同义替换特征。本研究结果有望为后续鉴定区分C3与C4物种的因果因子提供支撑，进而服务于C3植物向C4植物的性状转化研究。