猪circ0001651的鉴定及其在卵泡中的表达研究
摘要
本实验旨在探究circRNA在猪卵泡发育过程中的作用,通过生物信息学分析预测其可能的调控机制,为进一步探索circRNA对卵泡发育的调控机制奠定基础。采用RNA-Seq技术对卵泡中差异性表达的circRNA进行筛选,用Real-time PCR进行组织表达谱分析,并分析其在梅山猪和杜洛克猪卵泡中的表达情况。结果表明:circ0001651在肌肉、脂肪、心脏、肝脏、脾脏、肺脏、肾脏、卵巢、输卵管、子宫、子宫角、垂体、下丘脑中均有不同程度表达,其中在下丘脑、垂体、输卵管、卵巢表达量相对较高;circ0001651在梅山猪M2卵泡(M2卵泡直径:5.1～7.0 mm)中的表达量极显著高于杜洛克猪(P <0.01),在杜洛克猪M1卵泡(M1卵泡直径:3.1～5.0 mm)中的表达量极显著高于梅山猪(P<0.01);其潜在靶基因CCR2、EGFL7、TNFRSF12A、CFL1、EGR1、GADD45G、SLIT2等与猪的卵泡发育相关。生物信息学分析显示,circ0001651部分靶标基因参与细胞周期、细胞增殖凋亡及卵巢发育等相关信号通路,提示其可能通过对靶基因的调控作用间接参与卵泡发育过程。
引文
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[2]Sanger HL,Klotz G,Riesner D,et al.Viroids are singlestranded covalently closed circular RNA molecules existing as highly base-paired rod-like structures[J].Proc Natl Acad Sci,1976,73:3852-3256.
[3]马翔,任秀智,刘军龙,等.环状RNA的研究进展[J].中国生物制品学杂志,2016,29(8):881-884.
[4]Memczak S,Jens M,Elefsinioti A,et al.Circular RNAs are a large class of animal RNAs with regulatory potency[J].Nature,2013,495(7441):333-338.
[5]Salzman J,Chen RE,Olsen MN,et al.Cell-type specific features of circular RNA expression[J].PLoS Genet.2013,9(9):e1003777.
[6]宋新强,张丽丽.环状RNA的生物学功能[M].生物学教学,2016,41:72-74.
[7]Hansen T B,Jensen TI,Clausen B H,et al.Natural RNA circles function as efficient microRNA sponges[J].Nature,2013495(7441):384-388
[8]Ven?M T,Hansen T B,Ven?S T,et al.Spatio-temporal regulation of circular RNA expression during porcine embryonic brain development[J].Genome Biol,2015,16(1):245.
[9]Westholm J O,Miura P,Olson S,et al.Genome-wide analysis of drosophila circular RNAs reveals their structural and sequence properties and age-dependent neural accumulation[J].Cell Rep,2014,9(5):1966-1980.
[10]Szabo L,Morey R,Palpant N J,et al.Statistically based splicing detection reveals neural enrichment and tissue-specific induction of circular RNA during human fetal development[J].Genome Biol,2015,16(1):1-26.
[11]Livak K J,Schmittgen T D.Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T))Method[J].Methods,2001,25(4):402-408.
[12]Ye C Y,Chen L,Liu C,et al.Widespread noncoding circular RNAs in plants[J].New Phytologist,2015,208(1):88.
[13]Guo J U,Agarwal V,Guo H,et al.Expanded identification and characterization of mammalian circular RNAs[J].Genome Biol,2014,15(7):409.
[14]Fan X,Zhang X,Wu X,et al.Single-cell RNA-seqtranscriptome analysis of linear and circular RNAs in mouse preimplantation embryos[J].Genome Biol,2015,16(1):148.
[15]Gao Y,Wang J,Zhao F.CIRI:an efficient and unbiased algori thm for de novo,circular RNA identification[J].Genome Biol,2015,16(1):4.
[16]Jeck W R,Sorrentino J A,Wang K,et al.Circular RNAs are abundant,conserved,and associated with ALU repeats[J].RNA,2013,19(2):141-157.
[17]Memczak S,Jens M,Elefsinioti A,et al.Circular RNAs are a large class of animal RNAs with regulatory potency[J].Nature,2013,495(7441):333-338.
[18]Kino T,Hurt D E,Ichijo T,et al.Identification and characterization of circular RNAs as a new class of putative biomarkers in human blood[J].PLoS One,2015,10(10):e0141214.
[19]Memczak S,Papavasileiou P,Peters O,et al.Identification and characterization of circular RNAs as a new class of putative biomarkers in human blood[J].PLoS One,2015,10(10):e0141214.
[20]Li J,Yang J,Zhou P,et al.Circular RNAs in cancer:novel insights into origins,properties,functions and implications[J].Am J Cancer Res,2015,5(2):472-480.
[21]Shi F,Lapolt P S.Relationship between FoxO1 protein levels and folliculardevelopment,atresia,and luteinization in the rat ovary[J].J Endocrinol,2003,179(2):195-203.
[22]丁威.二花脸猪卵泡形成和早期发育的特性及其相关通路的研究[D].南京:南京农业大学,2010.