氨态氮对菲律宾蛤仔毒理机制研究的内参基因筛选
Alternative TitleSelection of Reference Genes for Toxicological Mechanism Research on Ruditapes philippinarum Exposed to Ammonia Nitrogen
曹滕飞; 丛明; 李兆艳; 赵建民; 吕家森; 李成华
Source Publication生态毒理学报
ISSN1673-5897
2017-04-15
Volume12Issue:2Pages:182-190
Keywordammonia nitrogen Ruditapes philippinarum reference gene 氨氮 菲律宾蛤仔 内参基因
DOIDOI: 10.7524/AJE.1673-5897.20160727001
Contribution Rank(1) 宁波大学海洋学院; (2) 中国科学院烟台海岸带研究所; (3) 烟台大学生命学院
Department海岸带环境过程实验室
AbstractIn order to analyze the transcriptome data accurately and systemically, a suitable reference gene is necessary. In the present study, geNorm and Normfinder softwares were used to analyze the expression stabilities of six candidate reference genes of Ruditapes philippinarum after ammonia nitrogen exposure for 30 days. The reference genes included 18S rRNA (18S), beta-actin (Actin), beta-tubulin (Tubu), elongation factor 1-alpha (EF1alpha), cyclophilin A (CyPA) and Ubiquitin (Ub). Based on a 50-times diluted cDNA in gills as the quantitative real-time PCR template, the geNorm software analysis demonstrated that the expression stabilities of the six candidate reference gene after NH_3-N exposure were as follows: EF1alpha>CyPA>Actin>Tubu>Ub>18S, and EF1alpha and CyPA were the best two ones among the six reference genes. A little different order was detected by the NormFinder software as follows: EF1alpha>Actin>Tubu>CyPA>Ub>18S. And EF1alpha was recommended as the optimal reference gene. As an assay, EF1alpha and CyPA were employed to examine one target unigene by qRT-PCR respectively. Final results showed that the variation trend detected by EF1alpha was more consistent with the foldchange value of the transcriptome. So EF1alpha was selected as the housekeeping gene to analyze the transcriptome data of gills in Ruditapes philippinarum after ammonia nitrogen exposure.
Other Abstract为了准确和系统地分析NH_3-N对菲律宾蛤仔(Ruditapes philippinarum)毒性的分子机制,并对相关基因的转录组学数据进行验证,利用geNorm和Normfinder这2种软件,以暴露30 d中不同时间点的鳃组织cDNA为模板,从18S rRNA(18S)、beta-actin(Actin)、beta-tubulin(Tubu)、elongation factor 1-alpha(EF1alpha)、cyclophilin A(CyPA)、Ubiquitin(Ub)等6个备选管家基因中,筛选可以稳定表达的内参基因,作为转录组学分析的内参基因。以稀释50倍的鳃组织cDNA模板作为检测模板, geNorm软件分析获得6对备选内参基因的表达稳定性依次为: EF1alpha>CyPA>Actin>Tubu>Ub>18S,较优内参为EF1alpha和CyPA; NormFinder软件分析获得6对备选内参基因的表达稳定性为: EF1alpha>Actin>Tubu>CyPA>Ub>18S,最优内参基因为EF1alpha。为了验证上述筛选结果,分别以EF1alpha和CyPA作为内参基因研究unigene1的表达趋势,发现以EF1alpha为内参基因时,unigene1的qRT-PCR的表达趋势与其转录组表达倍数的变化趋势一致。因此确定NH_3-N暴露菲律宾蛤仔后,鳃组织中表达最为稳定的内参基因为EF1alpha。
SubtypeArticle
Indexed ByCSCD
Language中文
WOS KeywordEnvironmental Sciences & Ecology
WOS Research AreaEnvironmental Sciences & Ecology ; 环境科学与生态学
CSCD IDCSCD:6012838
Citation statistics
Document Type期刊论文
Identifierhttp://ir.yic.ac.cn/handle/133337/24244
Collection中科院海岸带环境过程与生态修复重点实验室_海岸带环境过程实验室
Affiliation1.宁波大学海洋学院;
2.中国科学院烟台海岸带研究所;
3.烟台大学生命学院
Recommended Citation
GB/T 7714
曹滕飞,丛明,李兆艳,等. 氨态氮对菲律宾蛤仔毒理机制研究的内参基因筛选[J]. 生态毒理学报,2017,12(2):182-190.
APA 曹滕飞,丛明,李兆艳,赵建民,吕家森,&李成华.(2017).氨态氮对菲律宾蛤仔毒理机制研究的内参基因筛选.生态毒理学报,12(2),182-190.
MLA 曹滕飞,et al."氨态氮对菲律宾蛤仔毒理机制研究的内参基因筛选".生态毒理学报 12.2(2017):182-190.
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