摘要
【目的】筛选5龄飞蝗不同发育时间的最适内参基因,为相关研究提供基础数据。【方法】本文选取β-肌动蛋白(β-actin)、延长因子(EF-1α)、3-磷酸甘油醛脱氢酶(GAPDH)、核糖体蛋白49(RP49)、α-微管蛋白(α-Tubulin)和18S核糖体RNA(18S rRNA)基因作为候选内参基因,运用实时定量PCR(qPCR)方法研究各基因在5龄飞蝗不同发育时间的相对表达量,用geNorm与Normfmder软件分析这6个基因表达稳定性。【结果】geNorm分析结果显示6个内参基因表达稳定度M值顺序为:β-actin(0.3720)>RP49(0.3750)>α-Tubulin(0.4030)>18S rRNA(0.4270)>EF-1α(0.4970)>GAPDH(0.6040)。M值越小表示基因表达稳定度越高,同时geNorm软件以标准化因子配对差异值(Pairwise variations)0.15默认为取舍值,由于V2/3=0.098<0.15,所以最适内参基因数目为2个。运用NormFinder软件也得出相似的结果。【结论】β-actin与RP49为5龄飞蝗不同发育时间的最适内参基因。
[Objectives] To identify the most appropriate reference genes on different days during the development of fifth-instar nymphs of Locusta migratoria to pave the way for relevant studies. [Metllods] Six genes were selected as candidate reference genes, including the beta actin gene （β-actin）, the Elongation factor la gene （EF-la）, the Glyeeraldehyde-3-phosphate dehydrogenase gene （GAPDH）, the Ribosomal protein 49 gene （RP49）, the Alpha tubulin gene （α-Tubulin） and the 18S ribosome RNA gene （18S rRNA）. The relative expression of these genes on different days during the development of L. migratoria was detected with quantitative real-time PCR （qPCR）. The expression stability of these genes was then investigated using geNorm and Normfinder software. [Results] Based on the results of geNorm analysis, the average expression stability （M） of the six genes was, in descending order; fl-actin （0.3720） 〉 RP49 （0.3750） 〉 α-Tubulin （0.4030） 〉 18S rRNA （0.4270） 〉 EF-la （0.4970） 〉 GAPDH （0.6040）. The less the M value the more stable the gene expression. The software took 0.15 as a constant value for pairwise variations. The appropriate number of reference genes was 2 because V2/3=0.098 〈 0.15. Similar results were obtained using NormFinder software. [Conclusion] β-actin and RP49 are the most suitable reference genes during the development of fifth-instar nymph ofL. migratoria.
出处
《应用昆虫学报》
CAS
CSCD
北大核心
2014年第3期733-740,共8页
Chinese Journal of Applied Entomology
基金
国家自然科学基金资助项目(31272380
31201710)
山西省高等学校留学回国人员科研资助项目(201163)
