Comparision And Application Of MtDNA Cyt B And CO Ⅰ Gene Sequences In Molecular Phylogeny From Some Species Of Scarabaeoidea

Chafers,belonging to Coleoptera Scarabaeoidea,commonly are called grub for their larva.Many kinds of species of Scarabaeoidea are the important agricultural pests. At present, the research on chafer are mainly focused on the research on morphology,preventing ect.However,the data of molecular level of chafers are quite limited.The mitochondrial cytochrome b gene (Cyt b)and Cytochrome oxidas subunitâ… gene (COâ… ) used as two ideal molecular markers have been extensively applied in study on molecular phylogeny of insects because of having characters of suitable evolutionary rate, few base pairs insertion and deletion,high level polymorphism,and they can be amplified using common primers.But small amounts of data were reported on Scarabaeoidea based on these two genes in domestic. Especially the reports on study based on COâ… sequence from Scarabaeoidea have not found in domestic.In this study,the mtDNA Cyt b and COâ… gene fragments of some species of Scarabeoidea were amplified by PCR and sequenced,the homologus sequences were compared and partial nucleotide sequences of Cyt b and COâ… were obtained.The used frequency of nucleotide was calculated and the molecular phylogenetic tree was reconstructed as well as phylogenetic relationship was analysed.There were three main contents in this experiment:extraction of total genomic DNA; amplification of mtDNA Cyt b and COâ… gene fragments;sequenceing and analyzing these two genes.The research conclusions were as follows:1. We obtained sequence data from the mitochondrial cytochrome b (Cyt b) gene (433bp) and cytochrome oxidase subunitâ… (658bp) for Scarabeoidea (without primers).The results in nucleotide composition showed:The average A+ T and G+C contents were 69.2% and 30.8% respectively for Cyt b (A=30.6%,T=38.6%,G=18.9%,C=11.9%),64.0% and 36.1% for COâ… (A=29.3%,T=34.7%,G=19.5%,C=16.6%). Many sites in both genes exhibited strong A+T nucleotide bias.2. The results in nucleotide divergence showed:the variable sites in Cyt b and CO I gene sequences were mostly concentrated on the third site, accounting for 70.34% and 79.25% respectively.The rates of transition and transversion(TS/TV) were 0.77-1.84 and 0.72-1.85 in Cyt b and COâ… fragments of samples.3.144 AA were coded in Cyt b gene, there were 24 variable AA among them, accounting for 16.67%; 219 AA were coded in COâ… gene and there were 16 variable AA,accounting for 7.31%. Leu was found many times in two sequences.4. Our data on phylogenetic tree by Mega 4.0 biosoftware were consistent with morphological taxonomy of Scarabaeoidae.The results showed that Anomala smaragdina has the closely relationships with Anomala mongolica,has the secondly closely relationships to Anomala corpulenta.The distant relationship beween Rutelidae and Cetoniidae was similar than that beween Rutelidae and Melolonthidae.The conclusion revealed that Cyt b and CO I gene sequences could be suitable as a molecular marker for identifying the genetic distance among Scarabeoidea population.