线粒体基因orf352的破坏部分恢复了细胞质雄性不育水稻的花粉发育
植物线粒体基因组有时携带细胞质雄性不育(CMS)相关基因。这些基因已经在农业上被利用，在各种作物上产生高产的F1杂交种子。Orf352基因被报道为水稻(Oryza Sativa)中的RT102型CMS基因，但其在CMS中的作用缺乏因果证据。
在这里，我们利用线粒体靶向转录激活因子样效应子核酸酶(MitoTALENs)，从细胞质雄性不育水稻RT102A的线粒体基因组中敲除orf352。我们在RT102A中分离出18个独立的转化事件，这些事件导致orf352的基因组编辑，包括它在几种植物中完全从线粒体基因组中移除。对mitoTALEN靶位点周围的序列分析表明，它们诱导的双链断裂可以通过同源重组修复。
在5‘靶位点附近，修复涉及与orf284相同的序列，而修复3’靶位点产生了各种新的序列，这些新序列产生了由orf352片段组成的新的嵌合基因。
含有编码ORF352的179~352氨基酸的新线粒体基因的植株表现出与RT102A相同的缩小花粉粒表型，而缺失orf352或含有编码ORF352的211~352氨基酸的新基因的植株虽然不能结实，但部分挽救了花粉的活力和萌发。
这些结果表明，orf352基因的缺失部分恢复了花粉的发育，表明ORF352的179~210位氨基酸可能参与了花粉败育的建立。
Plant mitochondrial genomes sometimes carry cytoplasmic male sterility (CMS)-associated genes. These genes have been harnessed in agriculture to produce high-yielding F1 hybrid seeds in various crops. The gene orf352 was reported to be an RT102-type CMS gene in rice (Oryza sativa), although a causal demonstration of its role in CMS is lacking. Here, we employed mitochondrion-targeted transcription activator-like effector nucleases (mitoTALENs), to knock out orf352 from the mitochondrial genome in the cytoplasmic male sterile rice RT102A. We isolated 18 independent transformation events in RT102A that resulted in genome editing of orf352, including its complete removal from the mitochondrial genome in several plants. Sequence analysis around the mitoTALEN target sites revealed the repair of their induced double-strand breaks via homologous recombination. Near the 5' target site, repair involved sequences identical to orf284, while repair of the 3' target site yielded various new sequences that generated new chimeric genes consisting orf352 fragments. Plants with a new mitochondrial gene encoding amino acids 179 to 352 of ORF352 exhibited the same shrunken pollen grain phenotype as RT102A, whereas plants either lacking orf352 or harboring a new gene encoding amino acids 211 to 352 of ORF352 showed partial rescue of pollen viability and germination, although they failed to set seed. These results demonstrated that disruption of orf352 partially restored pollen development, indicating that amino acids 179 to 210 from ORF352 may contribute to the establishment of pollen abortion.
doi: https://doi.org/10.1101/2021.02.24.432723
文章来源原创，转载请注明来源。
温馨提示：
为方便PaperRSS粉丝们科研、就业等话题交流。我们根据10多个专业方向（植物、医学、药学、人工智能、化学、物理、财经管理、体育等），特建立了30个国内外博士交流群。群成员来源欧美、日韩、新加坡、清华北大、中科院等全球名校。