Identification Of Structural Proteins In The Scale Of Silkworm, Bombyx Mori

Lepidoptera is the biggest pest group which threat to agricultural production all over the world. In order to control the pests, large quantities of chemical pesticides have been used to reduce the damage caused by them to agricultural production. However, the abuse of chemical pesticides not only pollute the environment and threaten people's health, but also had a negative impact on Non-target organisms.The typical difference between Lepidoptera insects and other organisms is that Lepidoptera insects have scales which form the surface of colorful Lepidoptera wing patterns. Scales are the main constituent elements of mimicry and camouflage for Lepidoptera insects. Scales are a kind of sophisticated derivatives which have the fine structure and the epidermal derivatives which produced by the epidermal cells. Also scales have a temperature control feature allows the parasites have to adapt to the environment temperature. The structure color is produced by the interaction of their precise structure and the light. People have done a lot of work to research the mechanism of the structure of color, but it remains to be studied.Silkworm (Bombyx mori) is a model organism for Lepidoptera research, we have the genomic data with detailed information. Therefore, this paper started from the perspective of molecular biology to research the silkworm (Bombyx mori) scales.We use the silkworm genome database, EST database and cDNA microarray database to complete the Bioinformatics analysis, The Proteins have been extracted by use a combination method of SDS extraction and trypsin digestion. A LC-MS/MS has been used to investigate the composition of silkworm scales protein. In addition, RNAi technology has been used to study the influence of some epidermial protein gene to the phenotype of silkworm scale. The main results as following:1. The protein composition of silkworm scales: In this study, we make use of a combination of SDS extraction and trypsin digestion as a protein extraction Method, Then we use the LC-MS/MS to investigat the protein composition of silkworm scales. As a result we found 175 peptides in the silkworm scales and found that there are 40 proteins match with these peptides in the local database which based on the silkDB, NCBI and silkworm cDNA database. There are 7 epidermal proteins have been found and these proteins are unsoluble epidermal proteins which are obtained by using trypsin digeste the pellet after the the SDS extracts. This is the first time find proteins in the silkworm scales, and more than half of these epidermal proteins are BmorCPG protein family. In the SDS protein extraction, we found 31 kinds of non-epidermal protein, the protein they belong to 9 categories:(1) muscle protein (2) protease inhibitors (3) transfer protein (4), protein metabolism and carbohydrate metabolism (5) protein secreted proteins (6) heat shock protein (7) redox-related protein (8) storage protein (9) other proteins.2. The function of epidermal protein gene in silkworm scalesAs to expore the influence of the previously identified epidermal protein gene of silkworm on the scales and sockets, we use the RNAi technology in silkworm pupae by injecting siRNA of different gene to do interference experiment. The results show that after siRNA injected, we failed to find any significant differences on molt after silkworm eclosion. By using the fluorescence quantitative PCR, we detected the four epidermal protein Gene expression on mRNA level which are BmCPG1,BmCPR2, BmCPR63, BmCPG11.The detect results compared with the negative control (injection EGFPsiRNA).we found a decline in varying degrees. Then we use the SEM to investigate the phenotypic of scales and sockets after RNAi And found that compared with the control group, the experimental group ridge width and space between ridge has a decreasing trend, while the density of the scales and diameter of sockets tends to decrease. This may be because the mRNA level of the epidermal gene reduced thereby affecting the synthesis of the epidermal protein.The epidermal cells do not have enough raw materials to complete construction of the scales. A very interesting thing is that we can observe in the surface of the ridges of different intervention groups there are a large number of bifurcation and cross which forming braided or beaded ridges.The reseason may be that a developmental disorders happened when the epidermal cells can not adjust their development process ofscales after the epidermal gene were interferedIn summary, inthis study we identified 40 proteins from scales, including 7 epidermal proteins, and we do the the RNAi experiments to interfere four epidermal protein genes which include BmCPGl, BmCPR2, BmCPR63, BmCPG11. The corresponding phenotypes were investigated and analyzed by SEM. We believe that the epidermal proteins is an important component of silkworm scales, they play an important role in the functional integrity and the development of scales.they are key proteins which build the sophisticated structure of scales.