Lu Youxue, Liu Naiyi, Xu Liuxiang, Fang Jie, Wang Shuyan
School of Life Science, Anhui University, Hefei, China.
Genes Genomics. 2018 Oct;40(10):1011-1022. doi: 10.1007/s13258-018-0709-x. Epub 2018 Jun 14.
Vanessa indica is a small butterfly lacking historical molecular and biological research. Vanessa indica belongs to the family Nymphalidae (Lepidoptera: Papilionoidea), which is the largest group of butterflies and are nearly ubiquitous. However, after more than a century of taxonomic and molecular studies, there is no consensus for family classification, and the phylogenetic relationships within Nymphalidae are controversial. The first objective was to sequence and characterize the complete mitochondrial genome of V. indica. The most important objective was to completely reconstruct the phylogenetic relationships for family members within Nymphalidae. The mitochondrial genomic DNA (mtDNA) of V. indica was extracted and amplified by polymerase chain reaction. The complete mitochondrial sequence was annotated and characterized by analyzing sequences with SeqMan program. The phylogenetic analyses were conducted on thirteen protein coding genes (PCGs) in 95 mtDNA of Nymphalidae downloaded from GenBank for reference using the maximum likelihood method and Bayesian inference to ensure the validity of the results. The complete mitogenome was a circular molecule with 15,191 bp consisting of 13 protein coding genes, two ribosomal RNA genes (16S rRNA and 12S rRNA), 22 transfer RNA (tRNA) genes, and an A + T-rich region (D-loop). The nucleotide composition of the genome was highly biased for A + T content, which accounts for 80.0% of the nucleotides. All the tRNAs have putative secondary structures that are characteristic of mitochondrial tRNAs, except tRNA. All the PCGs started with ATN codons, except cytochrome c oxidase subunit 1 (COX1), which was found to start with an unusual CGA codon. Four genes were observed to have unusual codons: COX1 terminated with atypical TT and the other three genes terminated with a single T. The A + T rich region of 327 bp consisted of repetitive sequences, including a ATAGA motif, a 19-bp poly-T stretch, and two microsatellite-like regions (TA). The phylogenetic analyses consistently placed Biblidinae as a sister cluster to Heliconiinae and Calinaginae as a sister clade to Satyrinae. Moreover, the phylogenetic tree identified Libytheinae as a monophyletic group within Nymphalidae. The complete mitogenome of V. indica was 15,191 bp with mitochondrial characterizations common for lepidopteran species, which enriched the mitochondria data of Nymphalid species. And the phylogenetic analysis revealed different classifications and relationships than those previously described. Our results are significant because they would be useful in further understanding of the evolutionary biology of Nymphalidae.
苎麻珍蝶是一种缺乏历史分子和生物学研究的小型蝴蝶。苎麻珍蝶属于蛱蝶科（鳞翅目：凤蝶总科），蛱蝶科是蝴蝶中最大的类群，几乎分布广泛。然而，经过一个多世纪的分类学和分子研究，对于科的分类尚无共识，蛱蝶科内部的系统发育关系也存在争议。第一个目标是对苎麻珍蝶的完整线粒体基因组进行测序和特征分析。最重要的目标是完全重建蛱蝶科内各成员的系统发育关系。提取苎麻珍蝶的线粒体基因组DNA（mtDNA），并通过聚合酶链反应进行扩增。通过使用SeqMan程序分析序列，对完整的线粒体序列进行注释和特征分析。利用最大似然法和贝叶斯推断，对从GenBank下载的95个蛱蝶科线粒体DNA中的13个蛋白质编码基因（PCGs）进行系统发育分析，以确保结果的有效性。完整的线粒体基因组是一个环状分子，长度为15191 bp，由13个蛋白质编码基因、两个核糖体RNA基因（16S rRNA和12S rRNA）、22个转移RNA（tRNA）基因和一个富含A+T的区域（D环）组成。基因组的核苷酸组成在A+T含量上高度偏向，占核苷酸的80.0%。除了tRNA^Ser(AGN)外，所有tRNA都具有线粒体tRNA特有的推定二级结构。所有PCGs均以ATN密码子起始，除了细胞色素c氧化酶亚基1（COX1），它被发现以不寻常的CGA密码子起始。观察到四个基因具有不寻常的密码子：COX1以非典型的TT结尾，其他三个基因以单个T结尾。327 bp的富含A+T的区域由重复序列组成，包括一个ATAGA基序、一个19 bp的聚T序列和两个微卫星样区域（TA）。系统发育分析一致将喙蝶亚科作为蚬蝶亚科的姐妹类群，将绢蛱蝶亚科作为眼蝶亚科的姐妹分支。此外，系统发育树将珍蝶亚科确定为蛱蝶科内的一个单系类群。苎麻珍蝶的完整线粒体基因组为15191 bp，具有鳞翅目物种常见的线粒体特征，丰富了蛱蝶科物种的线粒体数据。并且系统发育分析揭示了与先前描述不同的分类和关系。我们的结果具有重要意义，因为它们将有助于进一步理解蛱蝶科的进化生物学。
