【作者】 张阳；

【导师】 任竹梅；

【作者基本信息】 山西大学 ， 遗传学， 2012， 硕士

【摘要】 五倍子蚜属于昆虫纲Insecta同翅目Homoptera瘿绵蚜科Pemphigidae五节根蚜亚科Fordinae昆虫,是具有重要经济意义的蚜虫。其生活周期为转主寄生,且寄主较为单一,寄生性较强。五倍子蚜的夏寄主为漆树科盐肤木属植物,它们为盐肤木、青麸杨和红麸杨；冬寄主以苔藓类植物为主,其中以匐灯藓科、青藓科等苔藓植物居多。五倍子蚜在东亚及北美均有分布,其中在我国分布有6属14种。而角倍蚜Schlechtendalia chinensis是五倍子蚜分布最广、产量最多、应用最广的蚜虫类群,其夏寄主仅为盐肤木一个种,冬季主是以侧枝匐灯藓为主的匐灯藓科苔藓。角倍蚜在分子水平的研究较少,还未见有对其线粒体DNA全基因组序列的研究报道。线粒体是生物体中重要的细胞器,它与有氧呼吸、细胞衰老、能量代谢和细胞凋亡等过程。动物线粒体基因组通常为一个闭合状的环状DNA链,大小通常在15-20kb左右。线粒体基因组通常包含蛋白质编码基因(PCG)、转运RNA(tRNA)、核糖体RNA(rRNA)、非编码控制区(Control Region)或者A+T富含区(A+T-rich region),其中部分蛋白质编码区和A+T富含区较为保守,因此线粒体DNA全基因组的测定对该物种分子方面的研究有着重要意义。本研究以角倍蚜为实验材料,利用常规PCR技术与克隆技术对其线粒体DNA全基因组进行测定,主要结果如下：1.测得角倍蚜线粒体DNA全基因组序列长为15,568bp(其中一段序列还未测通),为一个闭合的环状双链DNA分子。该线粒体基因组含有37个基因,其中包括了13个蛋白质编码区基因、22个tRNA基因、2个rRNA基因以及A+T富含区。在各个编码区域中,AT%的含量也有不同,但是从总体情况来看,AT%的含量还是比较高的,达到了83%,这是昆虫线粒体DNA基因的共有特征。从各个编码区的组成看,非编码区(A+T rich)的AT%为88.6%,是所有编码区中最高的,即使是AT%含量最少的蛋白质编码区也达到了78.0%。2.在角倍蚜线粒体DNA蛋白质编码区中,均是由ATA、ATG、ATT和TAA作为起始密码子,而终止子也是有T、TAA和ATG三种,其中终止密码子T会在转录过程中补全,形成一个完整的密码子。在所有的编码区中,Cytb、COⅠ、COⅡ、 COⅢ保守性较好。3.在rRNA序列中,碱基含量占到线粒体基因组碱基总量的6.5%。利用tRNAscan-SE在线预测得到的角倍蚜线粒体DNA基因组中含有22个tRNA,与其他蚜虫含量一致。在测得到的tRNA中,长度大概在60-70bp之间,总量占到全基因组的14.5%。在所有的tRNA中有5个tRNA基因(Asp、Ala、Glu、 Thr和Trp)定位在轻链(L)上。它们均可折叠为三叶草结构。4.从GenBank下载豌豆蚜与麦二叉蚜线粒体DNA全基因组序列,与角倍蚜进行比较分析,结果显示：在基因组长度结构方面,豌豆蚜全线粒体基因组比角倍蚜和麦二叉蚜长,原因为豌豆蚜线粒体基因组分散着两个A+T富含区；在结构组成方面,比较明显是ND2位置的不同以及豌豆蚜含有两个A+T富含区；在碱基含量方面,三种蚜虫的含量也基本保持一致；在蛋白质编码区的起始、终止密码子方面,三种蚜虫的起始密码子大部分是ATA、ATG、TAA和ATT四种,而终止密码子除了均含有是TAA与ATG两种外,角倍蚜的ND1和ND2还含有T为终止密码子,这也符合昆虫线粒体DNA的特征。对三种蚜虫结构的进化分析得出,最早进化的豌豆蚜含有两个A+T富含区,但在蚜虫在长期的进化过程中编码区发生了缺失,由此造成了进化时间较晚的角倍蚜缺失了一个A+T富含区。另外,ND2和部分tRNA的位置转换也可以对物种的进化起到促进作用。更多还原

【Abstract】 The Rhus gall aphids (Insecta, Homoptera, Pemphigidae, section five root aphid subfamily Fordinae) is the important economic aphids for their applications in the area of medical and chemical industry. In their life cycles, the Rhus gall aphids transfer hosts between the primary hosts-Rhus species and the secondary hosts-moss species. This aphid group includes6genus14species, and mostly distribute in East-Asia exception for one species in North Amarica, among which the species Schlechtendalia chinensis is widely distributed. Its primary host is Rhus chinensis, and the secondary host is Mniaceae. Until now, the research on the complete mitochondrial genome of Schlechtendalia chinensis has not been reported.In cell biology, a mitochondrion (plural mitochondria) is a membrane enclosed organelle found in most eukaryoticcells, whose size is around15-20kb. Mitochondria are sometimes described as "cellular power plants" because they generate most of the cell’s supply ofadenosine triphosphate (ATP), used as a source of chemical energy. Mitochondrial genome usually contain protein coding genes (PCG), transfer RNA (tRNA), ribosomal RNA (rRNA), Control Region or A+T rich. The part of the protein coding region and A+T rich area is more conservative, so the complete mitochondrial genome has important significance for the research of molecular.In this study, we sequenced the complete mitochondrial genome sequence of Schlechtendalia chinensis by PCR technology and cloning method. The main results are as following:1. The whole mitochondrial genome sequence of Schlechtendalia chinensis is15,568bp (one fragment was not obtained), which is a closed circular double-stranded DNA structure. The mitochondrial genome contains37genes, including13protein coding genes,22tRNA genes,2rRNA gene and an A+T rich. In the coding region, the content of AT%is high (reaching83%), which comform the common characteristic of insect mitochondrial gene. From the composition of protein coding region, the AT%of A+T rich is88.6%, which is the highest of the coding regions.2. All the13protein coding genes are observed to use ATA, ATG, ATT and TAA as initiation codon. And the terminators are T, TAA and ATG, respectively. In the coding regions, Cytb, COI, COII, COIII are conserved. The rRNA sequences account for6.5%of the total mitochondrial genome.3. Using the tRNAscan-SE online, the22tRNA are consistent with other aphids. The length of tRNA is about60-70bp. In all the tRNA, there are5tRNA genes (Asp, Ala, Glu, Thr and Trp) positioned in the light chain (L). All the tRNAs are folded in the form of a typical clover leaf structure.4. From the GenBank, we download the whole mitochondrial genome sequences of the two aphids species Acyrthosiphon pisum and Schizaphis graminum. Comparing with the two sequences, the following results were obtained:First, the length of Acyrthosiphon pisum mtDNA genome sequence is longer than the other two. Second, the A+T rich region and ND2of Schlechtendalia chinensis is different with other aphids. Finally, in case of the initiation and termination codon, the three species of aphids are the same, except for the ND1and ND2of Schlechtendalia chinensis using T as termination codon. The genome organization of Acyrthosiphon pisum is different from Schlechtendalia chinensis, suggesting that the evolution of Acyrthosiphon pisum is earlier than others. In addition, the location of ND2and tRNA also played a positive role in the evolution of aphids.更多还原