隆额网翅蝗线粒体基因组全序列的测定与分析
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摘要
昆虫线粒体DNA是共价闭合的环状双链DNA分子，大小一般为15.4～16.3kb，以高拷贝数目存在于线粒体内。线粒体DNA的分子质量小，核酸序列和组成比较保守，基因组中不含间隔区和内含子，无重复序列和不等交换，拷贝数多，易于提取、扩增和分析：在遗传过程中不发生基因重组、倒位、易位等突变，严格遵守母系遗传方式，只需少量个体材料就能反映群体的遗传结构；基因组的替换率比核DNA高5-10倍，适合做进化生物学研究。
本文测定和分析了隆额网翅蝗线粒体全序列，并且将其于已经测出全基因组的2种其他直翅目昆虫的mtDNA进行了比较，并联合其他直翅类昆虫的mtDNA进行了线粒体全基因组水平的系统发育分析。获得的主要结果如下：
1．隆额网翅蝗线粒体基因组基因组成与基因排列
隆额网翅蝗线粒体DNA序列全长15783bp，包括13个蛋白质编码基因，2个核糖体基因，22个转运RNA基因，及1个长894个碱基的A+T丰富区，全线粒体基因组的A+T含量为76.4％。其基因排列顺序与非洲飞蝗的一致。隆额网翅蝗线粒体基因组在8个基因之间共有36bp的重叠区域，重叠范围在1bp到8bp之间变化。有114bp的基因间隔区，它们分散在20个基因之间，最长的间隔区长21bo，短的只有2bp。A+T丰富区位于核糖体12S小亚基和tRNA~(Ile)基因之间，A+T含量为86.0％，包括894个碱基，比其他昆虫的短一些。
2．隆额网翅蝗线粒体基因组蛋白质编码基因
隆额网翅蝗的密码子总数为3718，与非洲飞蝗的密码子(3713)总数接近。隆额网翅蝗编码基因的A+T含量为75.8％，它比非洲飞蝗的编码基因A+T含量高约1.7％。除COⅠ基因外，隆额网翅蝗的蛋白质编码基因的起始密码子都是典型的ATN密码子。其中ND2、COⅡ、ATP6、COⅢ、ND4L、ND6、Cytb这7个基因使用ATG作为起始密码子，ATP8、ND3、ND5基因使用ATT，ND4和ND1基因分别使用ATC和ATA。除COⅡ、ND5、ND4和ND1外，其他基因都是以TAA为终止密码子。亮氨酸Leucine(13.88％)、异亮氨酸isoleucine(10.87％)、丝氨酸serine(9.71％)、苯丙氨酸phenylalanine(9.23％)是隆额网翅蝗线粒体蛋白质中使用频率最高的4种氨基酸，总计占43.69％。
3．隆额网翅蝗线粒体基因组tRNA基因及tRNA二级结构
隆额网翅蝗线粒体DNA编码的22个tRNA基因零散分布于整个基因组中。从预测的二级结构来看，除tRNA~(Ser)(AGN)外，所有tRNA都具有典型的三叶草结构。tRNA~(Set)(AGN)中，双氢尿嘧啶(DHU)臂只是形成了一个简单的环。tRNA的
The mitochondrial DNA (mtDNA), which is typically a single circular duplex molecule with size ranging from approximately 15.4—16.3 kilobases, exists in almost all eukaryotic cells with highly copy numbers. Because of small size, compositional and nucleotide sequence conservation, maternal inheritance, and relatively rapid evolutionary rate, lack of intermolecular genetic recombination, excluding intron and intergenic spacer sequence, being easy to amplification and analysis, mtDNA has been extensively used for studying population structures and phylogenetic relationships at various taxonomic levels.
We determined the complete mitochondrial DNA sequence of Arcyptera coreana (Insecta: Orthoptera: Arcypteridae) and compare it with the other two orthopteran species, Locusta migratoria and Gryllotalpa orientalis. The concatenated nucleotide sequences of the 13 protein-coding genes from 8 polyneoptera insects and an outgroup taxon was analyzed for reconstruction their phylogenetic relationship. From the study we obtained the following conclusion:
1. Composition and gene order of A. coreana mitochondrial genome
A.coreana genome (15,783bp) contained the same 37 genes (two rRNAs, 22 tRNAs, and 13 proteins) plus the putative control region (894bp) as found in other arthropods, and share the identical gene order with Locusta migratoria. A total of 36 bp overlap regions among A.coreana mt genes are found in eight locations with size varied from 1-8 bp. A total of 114 bp of intergenic spacer sequence is spread over in 20 locations with size varied from 2-21 bp. The A+T-rich region known for the initiation of replication in vertebrates and invertebrates is located between the srRNA and tRNA~(Ile) genes, and contains the highest A+T content (86.0%) and spans 894 bp, which is shorter compared with that from other insects.
2. Protein-coding genes and codon usage
The total number of codons in the A.coreana protein coding genes was 3718, excluding termination codons in each gene, the number is close to L.migratoria (3713). The A+T content of the 13 A.coreana protein-coding genes is 75.8 %, which is a little higher than that of L. migratoria (77.5%). Four genes (COII> ND5、 ND4 和 ND1) have
本文测定和分析了隆额网翅蝗线粒体全序列，并且将其于已经测出全基因组的2种其他直翅目昆虫的mtDNA进行了比较，并联合其他直翅类昆虫的mtDNA进行了线粒体全基因组水平的系统发育分析。获得的主要结果如下：
1．隆额网翅蝗线粒体基因组基因组成与基因排列
隆额网翅蝗线粒体DNA序列全长15783bp，包括13个蛋白质编码基因，2个核糖体基因，22个转运RNA基因，及1个长894个碱基的A+T丰富区，全线粒体基因组的A+T含量为76.4％。其基因排列顺序与非洲飞蝗的一致。隆额网翅蝗线粒体基因组在8个基因之间共有36bp的重叠区域，重叠范围在1bp到8bp之间变化。有114bp的基因间隔区，它们分散在20个基因之间，最长的间隔区长21bo，短的只有2bp。A+T丰富区位于核糖体12S小亚基和tRNA~(Ile)基因之间，A+T含量为86.0％，包括894个碱基，比其他昆虫的短一些。
2．隆额网翅蝗线粒体基因组蛋白质编码基因
隆额网翅蝗的密码子总数为3718，与非洲飞蝗的密码子(3713)总数接近。隆额网翅蝗编码基因的A+T含量为75.8％，它比非洲飞蝗的编码基因A+T含量高约1.7％。除COⅠ基因外，隆额网翅蝗的蛋白质编码基因的起始密码子都是典型的ATN密码子。其中ND2、COⅡ、ATP6、COⅢ、ND4L、ND6、Cytb这7个基因使用ATG作为起始密码子，ATP8、ND3、ND5基因使用ATT，ND4和ND1基因分别使用ATC和ATA。除COⅡ、ND5、ND4和ND1外，其他基因都是以TAA为终止密码子。亮氨酸Leucine(13.88％)、异亮氨酸isoleucine(10.87％)、丝氨酸serine(9.71％)、苯丙氨酸phenylalanine(9.23％)是隆额网翅蝗线粒体蛋白质中使用频率最高的4种氨基酸，总计占43.69％。
3．隆额网翅蝗线粒体基因组tRNA基因及tRNA二级结构
隆额网翅蝗线粒体DNA编码的22个tRNA基因零散分布于整个基因组中。从预测的二级结构来看，除tRNA~(Ser)(AGN)外，所有tRNA都具有典型的三叶草结构。tRNA~(Set)(AGN)中，双氢尿嘧啶(DHU)臂只是形成了一个简单的环。tRNA的
The mitochondrial DNA (mtDNA), which is typically a single circular duplex molecule with size ranging from approximately 15.4—16.3 kilobases, exists in almost all eukaryotic cells with highly copy numbers. Because of small size, compositional and nucleotide sequence conservation, maternal inheritance, and relatively rapid evolutionary rate, lack of intermolecular genetic recombination, excluding intron and intergenic spacer sequence, being easy to amplification and analysis, mtDNA has been extensively used for studying population structures and phylogenetic relationships at various taxonomic levels.
We determined the complete mitochondrial DNA sequence of Arcyptera coreana (Insecta: Orthoptera: Arcypteridae) and compare it with the other two orthopteran species, Locusta migratoria and Gryllotalpa orientalis. The concatenated nucleotide sequences of the 13 protein-coding genes from 8 polyneoptera insects and an outgroup taxon was analyzed for reconstruction their phylogenetic relationship. From the study we obtained the following conclusion:
1. Composition and gene order of A. coreana mitochondrial genome
A.coreana genome (15,783bp) contained the same 37 genes (two rRNAs, 22 tRNAs, and 13 proteins) plus the putative control region (894bp) as found in other arthropods, and share the identical gene order with Locusta migratoria. A total of 36 bp overlap regions among A.coreana mt genes are found in eight locations with size varied from 1-8 bp. A total of 114 bp of intergenic spacer sequence is spread over in 20 locations with size varied from 2-21 bp. The A+T-rich region known for the initiation of replication in vertebrates and invertebrates is located between the srRNA and tRNA~(Ile) genes, and contains the highest A+T content (86.0%) and spans 894 bp, which is shorter compared with that from other insects.
2. Protein-coding genes and codon usage
The total number of codons in the A.coreana protein coding genes was 3718, excluding termination codons in each gene, the number is close to L.migratoria (3713). The A+T content of the 13 A.coreana protein-coding genes is 75.8 %, which is a little higher than that of L. migratoria (77.5%). Four genes (COII> ND5、 ND4 和 ND1) have
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