Sequencing And Analysis Of Two Mecopoda Mitochondrial Genomes

The mitochondrial genome (mitogenome) of two long-legged katydids, Mecopodaelongata and Mecopoda niponensis, was determined using the strategy of Sub-PCR basedon L-PCR and the method of PCR direct sequencing combining with clone sequencing.Also we compared the two Mecopoda with other41orthopteran available on GenBankfrom aspects of comparative genomics. The conclusions drew from the study as follows:1. The complete mitogenome of M. elongata and M. niponensis are15284bp and15364bp respectively, including13protein-coding genes,22tRNA genes,2rRNA genes, and anA+T-rich region. The gene order and orientation of the two Mecopoda mitogenome areidentical to those of Drosophila yakuba.2. The nucleotide composition in the two Mecopoda mitogenome show obvious bias, asthe A+T content of M. elongata and M. niponensis are71.8%and72.4%respectively. TheA+T content of A+T-rich region in the two Mecopoda are in lower level than other regions ineach mitogenome, the phenomenon also exist in other Ensifera insects available on GenBank,it may suggest that the highly A+T content is not essential to A+T-rich region.3. In the two Mecopoda mitogenomes, the three protein-coding genes nad1, nad2andcox1use specific but possible initiation codons TTG, TTG and ATGA respectively. The otherten protein-coding genes start with a typical ATN codon. Similarly, the conventionaltermination codons (TAA and TAG) could be assigned to all the protein-coding genesequences of the two species with the exception of three genes (cox2, nad4, nad5) in M.elongata and four genes (cox2, nad4, nad5, Cytb) in M. niponensis, which have incompletetermination codons of a single thymine (T) base.4. There are strong biases in the both codon usage and amino acid usage in the twoMecopoda mitochondrial protein-coding genes. NNU and NNA are the most frequently usedcodons. Leucine, serine, phenylalanine and isoleucine are the most frequent amino acids, andtheir total compositions are42.82%(M. elongata) and42.88%(M. niponensis) respectively.5. In the two Mecopoda mitogenomes, each tRNA could be folded into typical cloverleafsecondary structures except for trnSAGN, which have a reduced DHU arm, as in otherorthopterans. There exist some unmatched base pairs in the two Mecopoda tRNAs, andoverwhelming majority of them are G-U pairs. The phenomenon also exist in otherorthopterans, it seems to infer that G-U pairing is likely to be a kind of correct base pairing in mitogenomes.6. The length of A+T-rich region of M. elongata and M. niponensis were determined tobe294bp and393bp respectively, which were only longer than Ruspolia dubia (70bp)among orthopterans available on GenBank.7. There are three long intergenic regions in the mitogenome of M. elongata and M.niponensis. the first is located between trnA and trnR，with the length of63bp and68bprespectively; the second is located between trnQ and trnM，with the length of55bp and26bprespectively; the third is located between trnSUCNand nad1, both with the length of21bp. Thelargest overlap is both present between trnC and trnW with8bp, the atp8/atp6andnad4/nad4L gene pairs appear to overlap7bp nucleotides. In this article, based on the41sequences available on GenBank plus the two new Mecopoda mitogenomes, we present acomparative analysis of intergenic spacer sequences and overlapping sequences. Conclusionswere as follows: A larger spacer of most species is present between trnSUCNand nad1withabout20bp, and there are no obvious homologies in nucleotide sequence among thesenoncoding regions; the atp8/atp6and nad4/nad4L gene pairs both overlap by conserved7bpnucleotides.