Cloning And Sequence Analyses Of Mitochondrial ATPase8 Gene And ATPase6 Gene Of Eriogyna Pyretorum

The research in Mitochondrial genome (mtDNA) structure, function of the Eriogyna pyretorum haven't been reported up to now, and it is important for the study of Eriogyna pyretoyum at the phylogenesis evolution and the relative relation-ship with other insects. This study used larva of Eriogyna pyretoyum as the experimental materials. We design some primers according to the known Mitochondrial DNA genome sequences of Bombyx mori, Bombyx mandarina and the Antheraea pernyi etc., two fragments of Eriogyna pyretoyum mtDNA was acquired, and it was cloned in pMD-18 T vector.Two fragments were acquired after sequencing. One is a 452 bp sequence. Blast showed that this sequence includes the whole length of Mitochondrial mtDNA ATPase8 gene, upstream to it have 2 RNA genes, they are tRNA- Lys and tRNA- Asp respectively. Another fragment is a 781bp sequence, Blast revealed that, it is Mitochondrial mtDNA ATPase6 gene, the downstream is the part sequence of COXIII. The whole length is 1147bp combined two sequences using DNastar technique, in the sequence, the region from 86bp tol56bp is tRNA- Lys gene, and the region from 179bp to 251bp is tRNA-Asp gene, while from 252bp to 413bp is ATPase8 gene, and from 407bp to 1084bp is ATPase6 gene. We deduced that the acquirable sequence included Mitochondrial DNA ATPase6 gene and ATPase8 gene of Eriogyna pyretorum.Based on the sequences of mtDNA ATPase8 and ATPase6 of Eriogyna pyretorum and other 14 insects which were published on GenBank, comparison of homology gene sequence and amino acid sequence showed that the mtDNA ATPase8 sequence of Eriogyna pyretoyum has identity among 63.5%～91.4% to the other 14 kinds of insects, the highest is to Antheraea pernyi and Antheraea yamamai, and the lowest is to Camponotus. The mtDNA ATPase6 of Eriogyna pyretorum has comparability among 68.5%～90.5% to the other 14 insects, the highest is to Antheraea pernyi and Antheraea yamamai, while the lowest is to Camponotus. For the whole length of ATPase8 and ATPase6, Eriogyna pyretorum showed 68.5% to 90.5% to the other 14 insects. It has the same result that the highest identity is to Antheraea pernyi and Antheraea yamamai, while the lowest is to Camponotus. Compared the amino acid sequence of mtDNA ATPase8 and ATPase6 of Eriogyna pyretorum with other 14 insects, it is showed that mtDNA ATPase8 of Eriogyna pyretorum has identity among 27.5%～79.2% to the other 14 insects, the highest is Samia cynthia ricini, and the lowest is Camponotus, and mtDNA ATPase6 of Eriogyna pyretorum has comparability among 55.9%～96.0% to other 14 insects, the highest is Samia cynthia ricini and Antheraea pernyi, the lowest is Camponotus. For the whole length of ATPase8 and ATPase6, Eriogyna pyretorum showed 50.9% to 92.8% to other 14 insects. It has the same result that the highest identity is to Samia cynthia ricini, while the lowest is to Camponotus.According to the method of Clustal W of Dnastar, phylogenetic tree of amino acid sequences and nucleotide sequences were achieved. Compared the phylogenetic tree of amino acid sequences with that of nucleotide sequence, they almost have the same structure only have some varieties of species distance. The phylogenetic tree indicated that the identity of Bombyx mori to Bombyx mandarina, and the identity of Antheraea pernyi to Antheraea yamamai are high. The identity of Eriogyna pyretorum to Samia cynthia ricini,Antheraea yamamai,Antheraea pernyi is higher than that of Eriogyna pyretoyum to Bombyx mori and Bombyx mandarina,This study acquired the sequence of mtDNA ATPase6 and ATPase8 gene of Eriogyna pyretorum, and according to analysis of homology reviewed the relation in the evolving process and the relative relation of Eriogyna pyretoyum with Bombyx mori,Antheraea pernyi,Samia cynthia ricini,Antheraea yamamai and Bombyx mandarina. It also provides foundation for the origin, evolvement and categorization in silk insects.