鸭脂肪细胞中circ-FBLN2鉴定与功能预测

doi:10.3969/j.issn.1004-1524.2021.04.05

浙江农业学报 ›› 2021, Vol. 33 ›› Issue (4): 602-609.DOI: 10.3969/j.issn.1004-1524.2021.04.05

鸭脂肪细胞中circ-FBLN2鉴定与功能预测

胡晓丹(), 王来娣, 杨婷, 白皓, 江勇, 王志秀, 王莎莎, 陈国宏, 常国斌^*()

扬州大学动物科学技术学院,江苏扬州 225009

收稿日期:2020-07-17 出版日期:2021-04-25 发布日期:2021-04-25
通讯作者: 常国斌
作者简介:*常国斌,E-mail:passioncgb@163.com
胡晓丹(1995—),女,福建泉州人,硕士研究生,主要从事动物遗传育种与繁殖研究。E-mail:chitahuxd@163.com
基金资助:
国家科技支撑计划(2015BAD03B03);国家自然科学基金(31802057);江苏省333人才工程

Validation and function prediction of circ-FBLN2 in duck adipocytes

HU Xiaodan(), WANG Laidi, YANG Ting, BAI Hao, JIANG Yong, WANG Zhixiu, WANG Shasha, CHEN Guohong, CHANG guobin^*()

College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China

Received:2020-07-17 Online:2021-04-25 Published:2021-04-25
Contact: CHANG guobin

摘要/Abstract

摘要：

Circular RNA(circRNA)是一类在转录过程中形成的首尾相接的环状非编码RNA,其在多种生物学过程以及疾病发生过程中发挥重要作用。基于前期鸭胚原代脂肪细胞分化过程中circRNA测序结果,本研究选择由FBLN2编码的circ-FBLN2进行验证分析。通过PCR扩增和焦磷酸测序技术获得接合序列,采用Rnase R消化实验进一步验证了circ-FBLN2的存在,采用核质分离技术得出circ-FBLN2主要存在于细胞质中,且其在脂肪细胞诱导分化后显著下调,在樱桃谷鸭各组织中的表达具有一定的差异,主要在脂肪和腿肌中表达。生物信息学分析结果显示,circ-FBLN2与预测到的靶标基因主要参与MAPK,Toll样受体,以及细胞因子与受体互作通路等信号通路,揭示其可能通过对靶基因的调控作用间接调控鸭脂肪细胞分化过程。

关键词: circ-FBLN2, 樱桃谷鸭, 脂肪细胞, 组织表达

Abstract:

Circular RNA (circRNA) is a kind of circular non-coding RNA characterized by head to tail connection during the transcription process, which plays an important role in many biological processes and diseases. Based on the results of circRNA sequencing during pre-adipocyte differentiation, circ-FBLN2 encoded by FBLN2 was selected for validation analysis. The conjugated sequence was obtained by PCR amplification and pyrosequencing. And circ-FBLN2 was further verified by RNase R digestion experiment. The results of nuclear and cytoplasmic separation experiment showed that circ-FBLN2 mainly existed in cytoplasm, and its expression level of duck adipocyte was significantly decreased after differentiation. There was a certain difference in the expression level of different tissues in Cherry Valley duck, it was mainly expressed in fat and leg muscle. Bioinformatics analysis showed that the predicted target genes and circ-FBLN2 were involved in MAPK, Toll-like receptor and cytokine-cytokine receptor interaction pathways and other signaling pathways. All these announced that circ-FBLN2 may indirectly regulate the differentiation of duck adipocytes by regulating the target genes.

Key words: circ-FBLN2, Cherry Valley duck, adipocyte, tissues expression

中图分类号:

S834

胡晓丹, 王来娣, 杨婷, 白皓, 江勇, 王志秀, 王莎莎, 陈国宏, 常国斌. 鸭脂肪细胞中circ-FBLN2鉴定与功能预测[J]. 浙江农业学报, 2021, 33(4): 602-609.

HU Xiaodan, WANG Laidi, YANG Ting, BAI Hao, JIANG Yong, WANG Zhixiu, WANG Shasha, CHEN Guohong, CHANG guobin. Validation and function prediction of circ-FBLN2 in duck adipocytes[J]. Acta Agriculturae Zhejiangensis, 2021, 33(4): 602-609.

图/表 10

表1 实验引物设计

Table 1 Design of experimental primers

引物名称Primer names	序列sequences (5'-3')	长度Product length/bp
circ-FBLN2 divergent primer	TTCAGTGCCACAACCTCTCC	194
	TCAGCTCCGAATCTGCAGTG
circ-FBLN2 convergent primer	GGAGAGGTTGTGGCACTGAA	1314
	CACTGCAGATTCGGAGCTGA
FBLN2	TCCACCATCAAGGAGAACAG	217
	CACAGCAGGACAGCATCACA
GAPDH	CACAGCCACACACGAAGACA
	CCTTAGCCAGCCCCAGTAGA	106

图1 circ-FBLN2琼脂糖凝胶电泳

Fig.1 circ-FBLN2 agarose gel electrophoresis

图2 circ-FBLN2模式图及接合序列测序峰图

Fig.2 circ-FBLN2 Pattern maps and conjugated sequence sequencing peaks

图3 circ-FBLN2在RNase R处理组与对照组的相对表达量比较

Fig.3 The relative expression of circ-FBLN2 in the treatment of RNase R and control group

图4 circ-FBLN2亚细胞定位

Fig.4 circ-FBLN2 subcellular localization

图5 樱桃谷鸭脂肪细胞分化过程中circ-FBLN2及主基因表达规律同一基因不同字母表示差异显著(P<0.05)。

Fig.5 circ-FBLN2 and major gene expression during adipocyte differentiation in Cherry Valley duck Different letter of the same gene means siginificant difference(P<0.05).

图6 circ-FBLN2基因在樱桃谷鸭不同组织中的表达情况

Fig.6 Expression of circ-FBLN2 gene in different tissues of Cherry Valley duck

图7 ceRNA(circ-FBLN2-miRNA-mRNA)网络图

Fig.7 ceRNA(circ-FBLN2-miRNA-mRNA) network diagram

图8 一百零七个靶标基因GO 分析结果图

Fig.8 GO analysis of 107 target genes

图9 一百零七个靶标基因KEGG 分析结果图

Fig.9 KEGG analysis of 107 target genes

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鸭脂肪细胞中circ-FBLN2鉴定与功能预测

Validation and function prediction of circ-FBLN2 in duck adipocytes

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