Long-SAGE Library Construction Of Diffrernt Feeding Habit Silkworm Varieties On Artificial Diet And Differential Expression Genes Analysis

The silkworm (Bombyx mori) feeding habits research is an important part of thesilkworm biology, physiology and genetics study, which Silkworm Artificial forageperforms an essential role of them. The feeding habits of silkworms varieties betweenboth of individuals and lines. The individual feeding habits variety of artificial forageleads to not only the significant differences in silkworms’ development but also themain practical technical obstacles of artificial forage. The physiological genetics andthe molecular mechanism of smell and taste investigation has been carried out bypredecessors extensively and deeply, however, the studies on molecular level of genesabout feeding habit has no breakthrough yet.Velculescu et. al. first proposed the serial analysis of gene expression(SAGE)method in1995. This method is effective, sensitive, high through-put, high abundance,easy to find the unknown sequence, etc., is a powerful and widely applicable tool ofquantitative classification and expressed genes contrast in various states.This thesis, investigated the silkworm varieties (lines) with significantdifferences on artificial forage as experimental materials, constructed the Long-SAGEdigital expression profiling library of the subjects, analyst of the differentiallyexpressed the Tags between high trophic silkworms and low trophic ones bybioinformatics methods systems, mined the artificial forage feeding-related genes ofsilkworms, has following results.1.We constructed8Long-SAGE digital expression profiling libraries using theheads and midguts at the beginning of3rdinstar of Guangshi and Luqi lines forexperimental materials, as well as the larvae at the beginning of2ndinstar of JingsongA and B lines with high and low feeding habits. And we set up the subtractive library according to the Long-SAGE digital expression profiling library of Guangshi andLuqi. Then compared above8Long-SAGE libraries, we gained4comparisonlibraries.2.We screened the Tags which could match the known genes and conformed thedifference standards, and found there were34Tags had common diff expressiongenes among Guangshi and Luqi head difference comparison library, Jingsong Acomparison library and Jingsong B comparison library. There were50genes, had thesame regulation trend, out of119common diff expression genes between Guangshiand Luqi head difference comparison library with Jingsong A comparisonlibrary,when82genes had the same regulation trend in179common diff expressiongenes between Guangshi and Luqi head difference comparison library with JingsongB comparison library. Similarly, there were21genes of the same regulation trend in113common diff expression genes among Jingsong A and B comparison library.3. We constructed the Long-SAGE library, then screened the Tags which did notmatched any genes. And we obtained84Tags which conformed our differencestandard among above four comparison libraries, and Blasted these Tags on NCBI.Then we found that there were39Tags could match the known genes or ESTsequences while45could match noting.4.Then we labeled Tags to above45known genes, extended the Tags(GLGI) andsuccessfully obtained17genes extended.5.And we analyst above45differential expression genes and found that therewere42genes involved their related counterpart metabolic pathways. Analyst of thesepathways’ functions, we concluded that one gene involved in the organism systemconstruction,24genes related to metabolic processes,18genes involved in thegenetic information processes, three genes related the cellular processes and a geneinvolved in the environmental information process.6.We also compared the reported olfactory-sense-related silkworm’s genes withthe constructed8SAGE libraries and4comparison libraries. As a conclusion, there was no significant diversity in most genes expect a few slight-different ones whosehad no common in the variety feeding habits experimental groups.