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Cloning and expression of Apis mellifera melittin cDNA in E. Coli

Abstract:

Honey bee venom, known as apitoxin, is composed of several peptides, the most important of which is melittin. This peptide is a current focus of research since it can improve the immune system and act against cancer due to its bactericidal, anti-mutagenic, anti-inflammatory and even contraceptive effects. This makes it very desirable to obtain melittin producing bacteria and for this reason, this study has aimed at the cloning of E. coli with the melittin gene from western bee. In order to do this, the total RNA of the western honey bee (Apis mellifera) has been extracted and a reverse transcription polymerase chain reaction (RT-PCR) has been carried out, at different annealing temperatures (68.0, 68.2, 68.4, 68.6, 68.8 and 69.0 ° C) to amplify the melittin cDNA. The annealing temperature of 68.4 ºC has allowed the highest production. Subsequently, this cDNA has been cloned into the pGEM-T vector, which has transformed E. coli JM109. This transformation has been corroborated by the blue/white test mediated by X-gal.