To understand the molecular mechanism of estrogen and to evaluate the role of the estrogen receptor in mediating estrogen action, the full-length cDNA of estrogen receptor alpha (ER alpha) was cloned from Monopterus a...
To understand the molecular mechanism of estrogen and to evaluate the role of the estrogen receptor in mediating estrogen action, the full-length cDNA of estrogen receptor alpha (ER alpha) was cloned from Monopterus albus, and its expression pattern and distribution were investigated. The ERa cDNA of M. albus includes an open reading frame of 1863 bp, a 140-bp 5'-untranslated region and a 797-bp 3'-untranslated region. Amino acid sequence homology analysis showed that the Monopterus albus ER alpha has a moderate degree of similarity with Sebastes schlegelii, Zoarces viviparus and Haplochromis burtoni (81.1%, 80.7% and 80.4%, respectively). Quantitative PCR results showed that the highest level of ER alpha expression was in the liver;the next highest level of expression was observed in the gonads, where it was expressed at high levels particularly in the ovary in developmental stages IV and V and in the testis in developmental stage II/III. Immunohistochemistry analysis showed that ER alpha was present as slender particles distributed mainly in the membranes of spermatocytes and oocytes in the testis and ovary, whereas no positive signal was observed in the cytoplasm of sperm cells. This report describes the first molecular characterization of full-length ER alpha and its tissue-specific distribution in M. albus.
The normal dose of 17 alpha-methyltestosterone (MT) used in fish farming was 60 mg/L, and now the analysis of residual androgens was carried out in waste water obtained from the Beijing area, which could be detected i...
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The normal dose of 17 alpha-methyltestosterone (MT) used in fish farming was 60 mg/L, and now the analysis of residual androgens was carried out in waste water obtained from the Beijing area, which could be detected in levels ranging from 4.1 to 7.0 ng/L. For the purpose of aquatic early warning, the present study clearly demonstrated that chronic exposure by higher concentration of MT than environmental relevant concentrations could trigger oxidative stress response to juvenile tilapia by modulating hepatic antioxidant enzyme activities and gene transcription. Some antioxidative parameters (T-GSH, GSH/GSSG and MDA) were significant decreased under 0.5 mg/L MT exposure at 7 and 14 days. Some antioxidant enzymes (SOD, CAT and GST) and transcriptional changes (sod and cat) were revealed significant decreases for MT treated groups at 7 days. Total antioxidant capacity was significant increased only in 5 mg/L MT exposure groups, but GR activities were not affected all through the whole exposure period. Almost all of the antioxidant enzymatic genes detected in the present study were showed significant increments for MT exposure both at 14 and 21 days, and the genotoxicity profile of antioxidant enzymatic genes were revealed dose-dependent manner. This study presented evidence that MT could result in oxidative stress response in the early stages of GIFT tilapia.
研究不同时间点(1、2、4、8、16 d)及修复期(26 d)、不同浓度(0、0.06、0.3、1.5 mg/L)二溴海因(DBDMH)对雄性吉富罗非鱼血浆中抗氧化酶基因的基因毒性,筛选稳定的生物标志物。实时荧光定量PCR方法检测DBDMH对超氧化物歧化酶(sod)、谷胱甘肽-S-转移酶(gst)、谷胱甘肽还原酶(gr)、谷胱甘肽过氧化物酶1(gpx1)、过氧化氢酶(cat)基因表达的影响。中浓度组雄性吉富罗非鱼血浆抗氧化酶基因(除gr)表达受DBDMH胁迫诱导显著上调;各DBDMH浓度组1 d(sod,gst,gpx1),2 d(gst,gr)抗氧化基因表达呈现浓度依赖性下降,4 d sod,gpx1基因表达呈现浓度依赖性上升;8 d中浓度DBDMH处理组基因表达(gst,gr,gpx1)显著低于低浓度组;恢复试验阶段,中、高浓度DBDMH组各抗氧化酶基因表达均不能恢复到正常水平。0.3 mg/L DBDMH显著增强雄性吉富罗非鱼血浆所测抗氧化酶基因的表达(除1 d gpx1、cat,8和16 d gr),此浓度下抗氧化酶基因作为标志物较合适。
黄鳝基因组DNA一直采用抽血或剪取肌肉来提取,但是采血或剪取肌肉后黄鳝成活率很低。为了保留亲本,尝试从黄鳝体表粘液中提取DNA,并采用紫外分光光度计对提取到的DNA的纯度和浓度进行了检测。结果表明,用蛋白酶K消化,酚仿抽提后从粘液中得到的DNA的纯度与从肌肉中提取到的相差不大,但是提取到的量较少。为了验证提取的DNA是不是黄鳝基因组DNA,用黄鳝actin引物和18S r RNA基因引物扩增之后检测到了粘液中的较亮的DNA片段。为了进一步确认检测到的是黄鳝的DNA,我们又对得到的DNA进行了扩增,并得到的产物进行了克隆和测序。结果表明,粘液中提取到的DNA是黄鳝的基因组DNA。随后又选择了60尾黄鳝,提取其粘液之后对其成活率进行统计,养殖1个月后黄鳝的成活率都在95%以上。此黄鳝体表粘液提取DNA方法能提取高质量的DNA,可用于用于PCR鉴定。
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