| The spado venom is secreted by venom glands with aromatic scents, it is a transparent liquid stored in the capsule drug, stung by a pad thorn from the needle. It is the most developed one of the bee products, on the 20th century.Foreign scholars have been studying bee venom of Apis mellifera, including the composition of bee venom purified activity Components Analysis and pharmacological role. As early as 1888, people used bee venom to treat rheumatism, early chelation therapy using gill direct manner, with the development of science and technology, it has emerged in natural bee venom injection produced by the injections. With the rapid molecular biology technology development, a series of bee venom functional genes have been successfully cloned, their prospects for development and use of genetic engineering rather optimistic. Researchs almost in a state of blank should be pay attention to the bee venom function and molecular biology rarely. The development use of bee venom resource can promote social and economic benefits.Bee venom is a transparent liquid, which has a special aromatic smell, bitter, acid reaction, pH5.0 to 5.5, the proportion is 1.13. At room temperature, it dry quickly and volatile liquid weight of the original 30% to 40%, the composition of volatile compounds containing at least 12 kinds of gas available over the composition of identification, including isoamyl acetatebased alarm hormone, Since its acquisition and easily lost in the refining process, they are usually referred to in the chemical composition of bee venom was ignored. Bee venom can easily dissolve in water, acid and glycerin, do not dissolve in alcohol. Bee venom is a complex mixture, which contains a lot of water in addition to, but also the hard work of containing protein peptide, enzymes, histamine, acids, such as amino acids and trace elements. In the material peptide, about 50 percent is dry, it neuropeptide dry bee venom is 3%. The bee venom enzymes up more than 55 species, A dry bee venom content of 12%, hyaluronic acid content of dry bee venom about 2% to 3%. The bee venom peptide (Apamin) is the second important peptides, and the dry weight of bee venom about 1% to 2%. In 1965, the first time that Habermann & Reitz, Haux, etc. At the same time set its primary structure, which is a 18 amino acid peptide composed of alkaline. Study found that the peptide is contained 46 amino acid peptide precursor of the bee venom in a certain enzymes in the cutting role, into the bee venom sac in the basic peptides.The molecular weight of 2.03 kDa peptide of the dry weight of the entire bee venom 1% to 2%.Apamin in bee venom peptides of the most notable features can be strong blocking calcium-activated potassium channel, is a strong neurotoxin. We mainly done the following work:1. We use RT technology to Apis mellifera bee venom for the base, and through the bee venom total RNA extraction,reverse transcription cDNA synthesis,in the application of RT-PCR method was that bee venom peptide gene fragments.2. The bee venom peptide DNA as a template to design primers for PCR amplification with Bam HΙand Eco RΙ, the PCR product was digested, access to the target gene fragment, then the target gene fragment with the same enzyme and the pGEX-2T plasmid vector, the recombinant plasmid transformed into E.coli BL21, and then under optimum conditions soluble expression was induced by IPTG. Collect cell, using SDS-PAGE preliminary identification. Conclusion:RT-PCR gene was cloned and identified.Construction Melittin GST fusion expression vector pGEX-AMapamin, recombinant plasmid was transformed into E.coli BL21 fusion expression. The GST affinity chromatography after SDS-PAGE analysis, the 31 kDa has a specific band, with the expected molecular weight of the same size that bee venom peptide and GST fusion protein in E.coli species have been expressed. By optimizing conditions of the expression can express the soluble protein.
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