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Kafe 바로가기주관연구기관 | 경상대학교 GyeongSang National University |
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보고서유형 | 2단계보고서 |
발행국가 | 대한민국 |
언어 | 한국어 |
발행년월 | 2015-07 |
과제시작연도 | 2014 |
주관부처 | 미래창조과학부 Ministry of Science, ICT and Future Planning |
등록번호 | TRKO201600009274 |
과제고유번호 | 1711015711 |
사업명 | 해양극지기초원천기술개발 |
DB 구축일자 | 2016-10-01 |
키워드 | 돌돔.혈구.ESTs 해석.Rock bream.Blood cell.ESTs analysis.cDNA microarray. |
DOI | https://doi.org/10.23000/TRKO201600009274 |
1. 3종 (LPS, 15℃, 25℃)의 적혈구 cDNA library를 제작하였음.
2. 총 5,310 클론 (LPS: 2,150, 15℃: 1,555, 25℃: 1,605) 획득.
3. 30개 이상 full length cDNA 확보, 유전자 구조 및 발현 해석.
4. ESTs 분석에서 얻어진 적혈구 유래 유전자를 spotting 된 chip 제작.
5. LPS 자극, 저온 자극 후 Microarray 분석을 통해 환경변화 및 병원체 침입시 각 혈구간의 상호작용 현상 및 적혈구의 기능 유추.
IV. Results of Research and Development
We have constructed three RBC cDNA library from LPS, cold or heat (15 and/or 25℃)-stimulation and performed the ESTs analysis from total of 5,310 (LPS-2,150; 15℃-1,555; 25℃-1,605) clones. We have investigated the molecular characterization and expressio
IV. Results of Research and Development
We have constructed three RBC cDNA library from LPS, cold or heat (15 and/or 25℃)-stimulation and performed the ESTs analysis from total of 5,310 (LPS-2,150; 15℃-1,555; 25℃-1,605) clones. We have investigated the molecular characterization and expression analysis of more than 30 genes which are obtained as full-length cDNA.
Furthermore, we have constructed the cDNA microarray chips spotted with RBC related genes. Finally, we performed functional analysis with cDNA microarray chips in RBC via water temperature and/or mitogen (LPS)-stimulation. Microarray analysis results indicated that between WBC and RBC have significant interaction relationship in rock bream.
In the result of the second step research, we have obtained more than 30 full-length cDNAs from rock bream leukocytes. Subsequently, we have investigated its immunological functions by molecular characterization and expression analysis. Moreover, we produced 12 kinds of synthetic peptide using its amino acid sequences. On the basis of the results, we have conducted more intensive study for functional analysis of leukocyte related genes by in vitro using these synthetic peptide.
In this study, we have accomplished immunofluorescence analysis using specific antibodies and confocal microscopy for confirmation of particular leukocyte presence in several tissues, such as liver, kidney, spleen, and gill from rock bream. As a result, we observed the leukocytes which are bind to specific antibodies in several tissues.
In addition, to analyze the function of leukocyte cell surface molecules in rock bream, we have isolated the pure leukocytes from rock bream blood. Subsequently, the leukocytes were bind to specific antibodies and then the leukocytes were stimulated with LPS. After that, the gene expression was investigated using cDNA microarray analysis in rock bream leukocyte group which are binding or non-binding with specific antibodies. The control groups were used non-binding antibody leukocytes. Consequently, we have observed total 1,649 of up- or down-regulated genes.
Finally, for functional analysis of fish lymphocytes, we have conducted lymphocyte cell sorting to identify the specific antigen presence lymphocytes, such as T-cell, B-cell, pre B-cell using antibodies and Flow cytometry.
과제명(ProjectTitle) : | - |
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연구책임자(Manager) : | - |
과제기간(DetailSeriesProject) : | - |
총연구비 (DetailSeriesProject) : | - |
키워드(keyword) : | - |
과제수행기간(LeadAgency) : | - |
연구목표(Goal) : | - |
연구내용(Abstract) : | - |
기대효과(Effect) : | - |
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