Studies of complicated systems of growth using GH transgenic Amago

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(1)Proteins analysis extracted from intestine, liver and serum using DIGEProteins were extracted from intestine, liver and serum of GH transgenic Amago, and these proteins were performed proteins analysis using DIGE. About 2000 proteins were separated by two-dimensional gel electro-phoresisis, and some of major proteins were extracted from the gel and performed MS/MS-analysis. Most of these were unknown proteins, however, amount of digestive enzymes and G-proteins were significantly greater in transgenic intestine than that of control. Analysis of liver tissue also showed vast differences among transgenic and control. There were proteins having similar to parasites amino sequences in transgenic, and these were about four times grater than control. These proteins were detected only all transgenic used for the experiment.(2)Analysis gene expression by microarrayWe have performed subtractive hybridization using liver tissue from GH transgenic and control, and made microarray plates using these subtracted DNA. Analysis of microarray was performed using RNA extracted from both F2 and F3 of immature and mature GH transgenic. The results of these experiments showed drastic decrease of natural immune system in immature young fish. However, this tendency was getting installed when they were matured. Further, some sex related gene expression was observed in immature transgenic, and these tendency were also showed both F2 and F3.(3)Screening of pituitary genes by subtractionThe completed length of cDNA library was obtained using pituitary of immature GH transgenic fish (F2), and also subtractive hybridization was performed using immature GH transgenic. Each 1000 clones of subtracted genes from both higher expression and lower expression were screened, and these have been sequenced.