Data Sheet 1_Mitochondrial genome of Lonicera macranthoides: features, RNA editing, and insights into male sterility.docx

IntroductionMitochondria are essential organelles that provide energy for plants. They are semi-autonomous, maternally inherited, and closely linked to cytoplasmic male sterility (CMS) in plants. Lonicera macranthoides, a widely used medicinal plant from the Caprifoliaceae family, is rich in chlorogenic acid (CGA) and its analogues, which are known for their antiviral and anticancer properties. However, studies on the mitogenome of L. macranthoides still remain limited.
MethodsThe mitochondrial DNA contained in the whole genome DNA was extracted from a male sterile cultivar of L. macranthoides, named ‘Yulei 1’. Next-generation sequencing (NGS) and third-generation sequencing (TGS) technologies were combined to obtain the mitogenome. RNA editing events were identified by integrating the mitogenome data with RNA sequencing data from leaf, stem, and flower tissues. The potential causes of male sterility in ‘Yulei 1’ were analyzed based on the loss of functional genes, mitogenome rearrangements, RNA editing events, and open reading frames (ORFs).
Results and discussionThe complete mitogenome of L. macranthoides ‘Yulei 1’ was obtained for the first time, with a length of 1,002,202 bp. It contains 48 protein-coding genes (PCGs), 26 tRNA genes, and 3 rRNA genes. Additionally, 79 simple sequence repeats (SSRs), 39 tandem repeats, and 99 dispersed repeats were identified. Among these, two direct repeats (RP1a/1b, RP2a/2b) and two inverse repeats (RP3a/b, RP4a/b) may facilitate mitogenome recombination. Gene transfer analysis revealed that 4.36% and 21.98% of mitogenomic sequences mapped to the chloroplast and nuclear genomes, respectively. Phylogenetic analysis indicated that L. macranthoides is closest to L. japonica at the mitogenome level. Notably, RNA editing events varied across different plant tissues, with 357 editing sites in 30 PCGs in leaves, 138 sites in 24 PCGs in flowers, and 68 sites in 13 PCGs in stems. Finally, all indications of CMS in the mitogenome were screened, including the detection of ORFs, and the findings showed no mutations in the mitogenome that would explain the sterility of ‘Yulei 1’. Overall, our study provides a complete mitogenome of L. macranthoides, which will aid in its genetic marker exploration, evolutionary relationship analysis, and breeding programs.

引言 线粒体是为植物提供能量的必需细胞器，具有半自主、母系遗传的特性，并与植物细胞质雄性不育（cytoplasmic male sterility, CMS）密切相关。灰毡毛忍冬（Lonicera macranthoides）是忍冬科（Caprifoliaceae）广泛应用的药用植物，富含绿原酸（chlorogenic acid, CGA）及其类似物，这类成分具备抗病毒与抗肿瘤活性。但目前针对灰毡毛忍冬线粒体基因组的研究仍较为匮乏。
方法 本研究从灰毡毛忍冬雄性不育品种‘玉蕾1号’中提取全基因组DNA中的线粒体DNA，结合二代测序（next-generation sequencing, NGS）与三代测序（third-generation sequencing, TGS）技术获取其线粒体基因组。通过将线粒体基因组数据与叶片、茎秆、花组织的RNA测序数据整合，鉴定RNA编辑事件。基于功能基因缺失、线粒体基因组重排、RNA编辑事件以及开放阅读框（open reading frames, ORFs），分析‘玉蕾1号’雄性不育的潜在诱因。
结果与讨论 本研究首次获得灰毡毛忍冬‘玉蕾1号’的完整线粒体基因组，其长度为1,002,202 bp。该基因组包含48个蛋白质编码基因（protein-coding genes, PCGs）、26个转运RNA（transfer RNA, tRNA）基因以及3个核糖体RNA（ribosomal RNA, rRNA）基因。此外，共鉴定出79个简单序列重复（simple sequence repeats, SSRs）、39个串联重复序列以及99个散在重复序列。其中，2对正向重复序列（RP1a/1b、RP2a/2b）与2对反向重复序列（RP3a/b、RP4a/b）可能参与介导线粒体基因组重组。基因转移分析显示，分别有4.36%与21.98%的线粒体基因组序列比对至叶绿体基因组与核基因组。系统发育分析表明，在线粒体基因组层面，灰毡毛忍冬与忍冬（Lonicera japonica）亲缘关系最近。值得注意的是，不同组织中的RNA编辑事件存在差异：叶片的30个蛋白质编码基因中共存在357个编辑位点，花的24个蛋白质编码基因中共存在138个编辑位点，茎秆的13个蛋白质编码基因中共存在68个编辑位点。最后，本研究对线粒体基因组中与细胞质雄性不育相关的所有特征进行了筛选（包括开放阅读框的检测），结果未发现可解释‘玉蕾1号’雄性不育的线粒体基因组突变。综上，本研究获得了灰毡毛忍冬的完整线粒体基因组，可为其遗传标记开发、进化关系分析以及育种工作提供重要支撑。