猪circ0015292的鉴定及其在卵泡中的表达研究

试验旨在通过高通量测序及生物信息学分析circRNA在猪卵泡发育过程中的作用及其可能的机制,为进一步探索circRNA对猪卵泡发育的调控机理奠定生物学基础。采集梅山和杜洛克猪不同级别的卵泡及各组织样本,提取总RNA,采用RNA-Seq技术对卵泡中差异表达的circRNA进行筛选,同时验证并分析其在梅山猪和杜洛克猪卵泡中的表达情况,并用实时荧光定量PCR进行组织表达谱分析。结果显示:试验证实了circ0015292的存在,且其在肌肉、心脏、肝脏、脾脏、肺脏、肾脏、卵巢、输卵管、子宫、垂体及下丘脑中均有不同程度的表达,在心脏、脾脏、肺脏、卵巢中的表达量相对较高;circ0015292在梅山猪S卵泡中的表达量极显著高于杜洛克猪(P<0.01),在杜洛克猪M1、M2、L卵泡中的表达量均显著高于梅山猪(P<0.05);其潜在靶基因GADD45G、CCR2、IL20RB、CFL1、LAMB3与猪的卵泡发育相关。综合试验结果,circ0015292在不同组织、不同时期卵泡中均有不同水平的表达;部分靶标基因参与细胞周期及卵巢发育相关信号通路,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,这为猪繁殖机理的研究提供了新思路。

Identification of Porcine circ0015292 and Its Expression Pattern in Ovarian Follicles

High throughput sequencing and bioinformatics were used to explore the role of circRNA in porcine follicular development and its possible mechanism,in order to provide the biological foundation for further exploring the regulatory mechanism of circRNA on porcine follicular development.Different levels of follicles and tissue samples from Meishan and Duroc pigs were collected,and then the total RNA was extracted.The differential expression of circRNA in follicles was screened by RNA-Seq technology.Meanwhile,the expression of RNA-Seq in the follicles of Meishan and Duroc pigs was validated and analyzed.At the same time,Real-time PCR was used to make tissue expression profiles.The results confirmed the existence of circ0015292,and it was expressed differently in muscle,heart,liver,spleen,lung,kidney,ovary,fallopian tube,uterus,pituitary gland and hypothalamus,and the expression level was relatively high in heart,spleen,lung and ovary.The expression level of circ0015292 in follicle S of Meishan pigs was extremely significantly higher than that of Duroc pigs (P<0.01),and the expression levels of follicles M1,M2 and L in Duroc pigs were significantly higher than those in Meishan pigs (P<0.05).The potential target genes of circ0015292 (GADD45G,CCR2,IL20RB,CFL1,LAMB3) were related to the development of porcine follicles.The results indicated that circ0015292 was expressed at different levels in different tissues and follicles at different stages;some target genes were involved in cell cycle and ovarian development-related signaling pathways,suggesting that circ0015292 might indirectly participate in follicular development through the regulation of target genes.This result would provide a new idea for the study of pig reproductive mechanism.