Construction And Partial Sequence Analysis Of CDNA Library From Mylabris Cichorii Linnaeus

Mylabris Cichorii Linnaeus(Coleoptera: Meloidae) is an important Chinese traditional medicine listed in the edited in 2005 (abbreviated as the Chinese Pharmacopoeia; Ch.P., 2005) which potent content cantharidin is highly poisonous to most of animals. For its extensive using in medicine, the requirement has up increased and the natural resources of Mylabris Cichorii have been dramatically decreased in China. There are a lot of researches achieved about the rearing of Meloidae insects, cantharidin synthesis and application. However, it has little research about Meloidae physiology, biochemistry and molecular biology, and even completely blank in some field. Its transcriptomic data of crest-time of cantharidin biosynthesis would provide invaluable information for molecular entomology and pave a way for the reserch of diapause of Meloidae insects, cantharidin biosynthesis and so on, and it would make foundation for achieving the artificial breeding of Meloidae insects, better utilization the resources of cantharidin even cantharidin artificial synthesis.We selected separated rearing Mylabris Cichorii Linnaeus adults which twenty-five days after eclosion as materials to construct the cDNA library. mRNA was extracted from Insects, then we synthesis the ss-cDNA by RT-PCR. The ds-cDNA were synthesized by the LD-PCR, then we used ds-cDNA more than 200bp to ligate with the carrier, and the products were transformed into XL1-Blue Electric Escherichia coli cells. 30 independent colonies were randomly picked from male and female cDNA library respectively, after treated by software we got 34 ESTs. After compared with the database were found that some genes have important physiological functions.The main results as follows:â‘ We successfully constructed the male and female adult cDNA library of Meloidae, and the quality of libraries meet the requirements. They can be used for further study.â‘¡We got some ESTs from the cDNA libraries, and after a preliminary comparison we found that EST sequences from two cDNA libraries had no significant differences in the length or forecast function. We used BlastX and BlastN programs compared the ESTs with the GeneBank database, the results showed that 41% of the ESTs have high similarity with gene sequences in the database, while 59% of the ESTs have less similarity with gene sequences in the database. It proves that Mylabris Cichorii Linnaeus has especial gene sequence, and construct library is the efficient method to discovery new genes. Including some important genes which significance to Mylabris Cichorii Linnaeus such as gene of cytochrome C oxidase (Accession number: GW342329-GW342333).â‘¢The phylogenetic tree generated from data of Cytochrome oxidase subunitâ… gene sequences revealed that Coleptera Meloidae and Coleptera Chrysomelidae had close relationship, lepidoptera and Coleptera had close relationship too. Diptera alone occupied a node. The results corresponded with the traditional classification criteria.