ASSESSMENT OF ENVIRONMENTAL STRESS USING GENE EXPRESSION IN AQUATIC INSECT

利用水生昆虫基因表达评估环境压力

• 批准号: 13680633
• 负责人: MIURA Takashi
• 金额: $ 1.98万
• 依托单位: TOKYO UNIVERSITY OF PHARMACY AND LIFE SCIENCE
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680633/
• 关键词: Environmental risk assessment; Chironomus yoshimatsui; gene expression; Midges in sediment

In order to develop the methods assessing environmental stress using gene expression of aquatic midges, the stress-induced or insecticide-induced gene fragments were isolated and gene expression was examined in insecticide-sensitive and field strains of Chironomus yoshimatsui1) Assessment of environmental stress by expression of stress-induced genesThe gene fragment of heat-shock protein, hsc70 was isolated from mRNA of field strain of the midge by RT-PCR reaction using the primer designed from hsp70 base sequences of four species insects and used for Northern blot analysis. In order to assess environmental stresses, RNA isolated from field strains were hybridized with hsc70 cDNA. The expression of this gene was higher in the midges collected in early MayThe inducible gene of hsp70 was also isolated using RNA from heat-treated midges by PT-PCR reaction using the primer designed from base sequence of Chironomus tentans and determined the sequence of full length gene. Using the probe specific for inducible gene, the expression of hsp70 was examined in RNA of heat-treated midges and revealed to reach the peak within 1hour and decline 6 hours later. These results show that the isolated hsp70 gene is inducible2) Assessment of gene expression induced by insecticide-treatmentIn order to assess insecticide-stress, RNAs were isolated from insecticide-treated midges and gene fragments increased by insecticide-treatment were separated in differential display method. Four fragments were isolated. The expression of these gene were higher in insecticide-treated midges and in field strains

为了建立利用水蠓基因表达评价环境胁迫的方法，分离了胁迫诱导或杀虫剂诱导的水蠓基因片段，并在杀虫剂敏感品系和田间品系中进行了基因表达检测。利用四种昆虫hsp70碱基序列设计的引物，采用RT-PCR方法从蠓野株mRNA中分离hsc70，并进行Northern blot分析。为了评估环境胁迫，从田间菌株分离的RNA与hsc70 cDNA杂交。该基因在5月初采集的蠓中表达量较高。利用从tentans Chironomus碱基序列设计的引物，用RNA从热处理的蠓中分离出hsp70的诱导基因，采用PT-PCR反应，确定了hsp70的全长基因序列。利用诱导基因特异性探针检测热激蛋白70在经热处理的蠓RNA中的表达，发现热激蛋白70在1小时内达到峰值，6小时后下降。结果表明，分离的hsp70基因具有诱导作用。2)杀虫剂诱导的基因表达评估为了评估杀虫剂胁迫，我们从杀虫剂处理的蠓中分离rna，并用差异展示法分离杀虫剂处理后增加的基因片段。分离出4个片段。这些基因在杀虫剂处理的蠓和田间品系中表达较高

项目成果

Yohimi, T., Minowa, K., Karouna-Renier, N., Watanabe, T., Sugaya, Y. and Miura, T.: "Activation of a stress-induced gene by insecticides in the midge, Chironomus yoshimatsui"J. Biochem. Mol. Toxicol.. 16. 10-17 (2002)

Yohimi, T.、Minowa, K.、Karouna-Renier, N.、Watanabe, T.、Sugaya, Y. 和 Miura, T.：“吉蚊摇蚊中杀虫剂对胁迫诱导基因的激活”J

Nakamura, N., Yoshimi, T. and Miura, T.: "Increased expression of lung marker proteins in the homeobox B3-overexpressed fetal lung cell line M3E3/C3"Cell Growth and Differen.,. 13. 195-203 (2002)

Nakamura, N.、Yoshimi, T. 和 Miura, T.：“同源框 B3 过表达的胎儿肺细胞系 M3E3/C3 中肺标记蛋白的表达增加”细胞生长和差异。

Tatsuya Yoshimi et al.: "Activation of a stress induced gene by insecticides in the midge"J. Biochem. Mol. Toxicol.. 16(1)(in press). (2002)

Tatsuya Yoshimi 等人：“杀虫剂对蠓中胁迫诱导基因的激活”J.