[논문]Circular RNA expression profiles in the porcine liver of two distinct phenotype pig breeds

Huang, Minjie; Shen, Yifei; Mao, Haiguang; Chen, Lixing; Chen, Jiucheng; Guo, Xiaoling; Xu, Ningying

doi:10.5713/ajas.17.0651

Circular RNA expression profiles in the porcine liver of two distinct phenotype pig breeds 원문보기

Asian-Australasian journal of animal sciences, v.31 no.6, 2018년, pp.812 - 819

Huang, Minjie (College of Animal Science, Zhejiang University) , Shen, Yifei (College of Animal Science, Zhejiang University) , Mao, Haiguang (College of Animal Science, Zhejiang University) , Chen, Lixing (College of Animal Science, Zhejiang University) , Chen, Jiucheng (College of Animal Science, Zhejiang University) , Guo, Xiaoling (College of Animal Science, Zhejiang University) , Xu, Ningying (College of Animal Science, Zhejiang University)

Abstract ▼ AI-Helper

Objective: An experiment was conducted to identify and characterize the circular RNA expression and metabolic characteristics in the liver of Jinhua pigs and Landrace pigs. Methods: Three Jinhua pigs and three Landrace pigs respectively at 70-day were slaughtered to collect the liver tissue samples. Immediately after slaughter, blood samples were taken to detect serum biochemical indicators. Total RNA extracted from liver tissue samples were used to prepare the library and then sequence on HiSeq 2500. Bioinformatic methods were employed to analyze sequence data to identify the circRNAs and predict the potential roles of differentially expressed circRNAs between the two breeds. Results: Significant differences in physiological and biochemical traits were observed between growing Jinhua and Landrace pigs. We identified 84,864 circRNA candidates in two breeds and 366 circRNAs were detected as significantly differentially expressed. Their host genes are involved in lipid biosynthetic and metabolic processes according to the gene ontology analysis and associated with metabolic pathways. Conclusion: Our research represents the first description of circRNA profiles in the porcine liver from two divergent phenotype pigs. The predicted miRNA-circRNA interaction provides important basis for miRNA-circRNA relationships in the porcine liver. These data expand the repertories of porcine circRNA and are conducive to understanding the possible molecular mechanisms involved in miRNA and circRNA. Our study provides basic data for further research of the biological functions of circRNAs in the porcine liver.

주제어

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문제 정의

In this study, we report the identification and characterization of circRNAs in the liver between the two distinct pig breeds. The intention of this study is to extend the circRNAs data in pig species and analyze putative circRNAs that may be related in breed-specific growth and fat deposition.

제안 방법

81% of candidate back-spliced junction reads were assembled by TopHat. Based on the sequence characteristics of splice site, 0.28% to 0.52% of clean reads respectively in each individual were realigned to determine circRNAs (Table 2). We annotated these circRNA candidates based on the NCBI database and UCSC genome browser (Supplementary Table S1) and identified 84,864 circRNAs in two breeds (Figure 1).
The liver plays a major role in carbohydrate, protein, amino acid and lipid metabolism [20]. In this study, we conducted a primary investigation of circRNA expression profiles in the liver of LD and JH pigs to detect the potential interaction with miRNA related to porcine growth and fat deposition. Ribosome RNA and linear RNA sequences were first removed to detect porcine liver transcriptome.
Quantitative real-time polymerase chain reaction (PCR) was performed on an ABI Step One Plus system (Applied Biosystem, Carlsbad, CA, USA) using GoTaq qPCR Master Mix (Promega, Madison, WI, USA) with specific primers. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was chosen as a control to normalize the technical variations.
RNA quantity and purity were analyzed with the ND-1000 Nanodrop (Thermo Fisher, Wilmington, DE, USA) and RNA integrity number (RIN) was analyzed with the Agilent2200 (Agilent, Palo Alto, CA, USA) with RIN>7.0.
The purified library products were evaluated using the Agilent 2200 Tape Station and Qubit2.0 (Life Technologies, Carlsbad, CA, USA) and then diluted to 10 pM for cluster generation in situ on the HiSeq2500 pair-end flow cell followed by sequencing (2×100 bp) on HiSeq 2500 (Illumina, USA).

이론/모형

13. Then the unmapped reads were compared to fusion genes of reference genome using TopHat-fusion algorithm. Back-spliced junction reads which corresponded to splicing sites of exonic circRNA forms were selected.

성능/효과

The larger absolute value of free energy indicated the stronger binding ability. The total matching score of ssc-miR-30c-5p and circ:chr5: 105610075-105629122 was the highest, and also had maximum free energy at the same time. Ssc-miR-184 and ssc_circ:chr7:53408460-53472958, ssc-miR-140-5p and ssc_circ:chr2: 162425884-162509599, ssc-miR-122 and ssc_circ:chr2:162425884- 162509599, ssc-miR-30c-5p and ssc_ circ:chr5:105610075-105629122 also had high scores and free energy according to the prediction results of miRanda (Supplementary Table S4).

참고문헌 (30)
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