Please wait a minute...
Journal of Integrative Agriculture  2017, Vol. 16 Issue (08): 1789-1799    DOI: 10.1016/S2095-3119(16)61624-6
Animal Science · Veterinary Medicine Advanced Online Publication | Current Issue | Archive | Adv Search |
Cloning, expression, and polymorphism of the ECI1 gene in various pig breeds
LU Yun-feng1, 2, CHEN Ji-bao2, ZHANG Bo1, LI Qing-gang1, 3, WANG Zhi-xiu1, ZHANG Hao1, WU Ke-liang1
1 College of Animal Science and Technology, China Agricultural University, Beijing 100193, P.R.China
2 School of Life Science and Technology, Nanyang Normal University, Nanyang 473061, P.R.China
3 Institute of Animal Sciences and Veterinary Medicine, Anhui Academy of Agricultural Sciences, Hefei 230031, P.R.China
Download:  PDF in ScienceDirect  
Export:  BibTeX | EndNote (RIS)      
Abstract      The enzyme Δ32-dienoyl-CoA isomerase (ECI1) plays a crucial role in the mitochondrial β-oxidation of fatty acids with a double-bond in odd and even positions. The ECI1 gene might be a qualified candidate for studies pertaining to lipid deposition and meat quality in swine. In the present study, ECI1 cDNA of the Tibetan pig was obtained by in silico cloning and verified by PCR analysis. Single-nucleotide polymorphisms (SNPs) of ECI1 were screened by PCR-sequencing and genotypes of those SNPs were tested by PCR-restriction fragment length polymorphism (PCR-RFLP) in Diannan small-ear pigs (DSP, n=40), Tibetan pigs (TP, n=60) and Yorkshire pigs (YP, n=30). The expression levels of ECI1 were analyzed by real-time quantitative PCR and Western blotting in tissues of the liver, backfat, and longissimus dorsi (LD) muscle of DSP (n=8), TP (n=8) and YP (n=8). Single factor linear correlation analysis was applied separately for each breed to evaluate correlations between ECI1 gene expression in the LD muscle and intramuscular fat (IMF) content. We obtained an ECI1 gene length of 1 401 bp from the cDNA that contained a full coding region of 909 bp. Three novel SNPs (g.42425337G>A; g.42424666A>G; and g.42422755A>G) were detected, and only g.42424666A>G exhibited three genotypes among the three breeds. The ECI1 expression levels in the LD muscle of DSP and TP were significantly higher than that of YP (P<0.05). Moreover, TP had the highest ECI1 expression in backfat (P<0.01), and a positive correlation was observed between gene expression and IMF content. The results suggest that differences in ECI1 gene expression might be related to lipid deposition and meat quality in pig.
Keywords:  ECI1 gene        lipid deposition        mRNA expression        pig        single-nucleotide polymorphism        Western blotting  
Received: 31 August 2016   Accepted:

This work was supported by the National Major Special Project on New Varieties Cultivation for Transgenic Organisms, China (2016ZX08009-003-006), the National Key Technology Research and Development Program of China (2012BAD03B03), the Scientific Research Foundation for Advanced Talents, Nanyang Normal University, China (ZX2014071), and the Program for Changjiang Scholar and Innovation Research Team in University, China (IRT1191).

Corresponding Authors:  Correspondence WU Ke-liang, Tel/Fax: +86-10-62734767, E-mail:    
About author:  LU Yun-feng, E-mail:;

Cite this article: 

LU Yun-feng, CHEN Ji-bao, ZHANG Bo, LI Qing-gang, WANG Zhi-xiu, ZHANG Hao, WU Ke-liang . 2017. Cloning, expression, and polymorphism of the ECI1 gene in various pig breeds. Journal of Integrative Agriculture, 16(08): 1789-1799.

Aslan O, Hamill R, Davey G, McBryan J, Mullen A, Gispert M, Sweeney T. 2012. Variation in the IGF2 gene promoter region is associated with intramuscular fat content in porcine skeletal muscle. Molecular Biology Reports, 9, 4101–4110.

Chen Q, Wang H, Zeng Y, Chen W. 2013. Developmental changes and effect on intramuscular fat content of H-FABP and A-FABP mRNA expression in pigs. Journal of Applied Genetics, 54, 119–123.

Chen Y, Zhao Y, Wu R.  2001. In silico cloning of mouse Muc5b gene and upregulation of its expression in mouse asthma model. American Journal of Respiratory and Critical Care Medicine, 164, 1059–1066.

Ding S, Schinkel A P, Weber T E, Mersmann H J. 2000. Expression of porcine transcription factors and genes related to fatty acid metabolism in different tissues and genetic populations. Journal of Animal Science, 78, 2127–2134.

Escher P, Wahli W. 2000. Peroxisome proliferator-activated receptors: Insight into multiple cellular functions. Mutation Research, 448, 121–138.

Euler-Bertram S, Stoffel W. 1990. Purification and characterization of bovine liver 3-cis-2-trans-enoyl-CoA isomerase. Biological Chemistry Hoppe-Seyler, 371, 603–610.

Fontanesi L, Scotti E, Buttazzoni L, Dall’Olio S, Davoli R, Russo V. 2010. A single nucleotide polymorphism in the porcine cathepsin K (CTSK) gene is associated with back fat thickness and production traits in Italian Duroc pigs. Molecular Biology Reports, 37, 491–495.

Frantz L A F, Madsen O, Megens H J, Schraiber J G, Paudel Y, Bosse M, Crooijmans R P M A, Larson G, Groenen M. 2015. Evolution of tibetan wild boars. Nature Genetics, 47, 188–189.

Gao X, Gu X, Han J. 2005. Clone full-length cDNA for chicken MIBP by silico cloning. China Journal of Bioinformatics, 2, 58–63. (in Chinese)

Geisbrecht B V, Zhang D, Schulz H, Gould S J. 1999. Characterization of PECI, a novel monofunctional Δ3,Δ2-enoyl-CoA isomerase of mammalian peroxisomes. Journal of Biological Chemistry, 274, 21797–21803.

Gong J, He Z, Li Z, Lv X, Ying S, Chen X. 2007. Investigation on fattening and carcass traits in Tibetan pig and its combinations. Southwest China Journal of Agricultural Science, 20, 1109–1112. (in Chinese)

Gurvitz A, Mursula A M, Firzinger A, Hamilton B, Kilpelainen SH, Hartig A, Ruis H, Hiltunen J K, Rottensteiner H. 1998.Peroxisomal Δ3-cis-Δ2-trans-enoyl-CoA isomerase encoded by ECI1 is required for growth of the yeast Saccharomyces cerevisiae on unsaturated fatty acids. Journal of Biological Chemistry, 273, 31366–31374.

Gurvitz A, Wabnegger L, Yagi A I, Binder M, Hartig A, Ruis H, Hamilton B, Dawes I W, Hiltunen J K, Rottensteiner H. Rottensteiner H. 1999. Function of human mitochondrial 2,4-dienoyl-CoA reductase and rat monofunctional Δ3-Δ2-enoyl-CoA isomerase in β-oxidation of unsaturated fatty acids. Biochemical Journal, 344, 903–914.

Han X L, Jiang T F, Yang H W, Zhang Q D, Wang W M, Fan B, Liu B. 2012. Investigation of four porcine candidate genes (H-FABP, MYOD1, UCP3 and MASTR) for meat quality traits in Large White pigs. Molecular Biology Reports, 39, 6599–6605.

Hiltunen J K, Filppula S A, Koivuranta K T, Siivari K, Qin Y M, Häyrinen H M. 1996. Peroxisomal β-oxidation and polyunsaturated fatty acids. Annals of the New York Academy of Sciences, 804, 116–128.

Hiltunen J K, Qin Y M. 2000. β-Oxidation - strategies for the metabolism of a wide variety of acyl-CoA esters. Biochimica et Biophysica Acta, 1484, 117–128.

Janssen U, Stoffel W. 2002. Disruption of mitochondrial β-oxidation of unsaturated fatty acids in the 3,2-trans-enoyl-CoA isomerase-deficient mouse. Journal of Biological Chemistry, 277, 19579–19584.

Justus J, Weigand E. 2014a. A moderate zinc deficiency does not impair gene expression of PPARa, PPARg, and mitochondrial enoyl-CoA delta isomerase in the liver of growing rats. Nutrition and Metabolic Insights, 7, 29–37.

Justus J, Weigand E. 2014b. The effect of a moderate zinc deficiency and dietary fat source on the activity and expression of the Δ3Δ2-enoyl-CoA isomerase in the liver of growing rats. Biological Trace Element Research, 158, 365–375.

Kilponen J M, Hiltunen J K. 1993. β-Oxidation of unsaturated fatty acids in human. Isoforms of Δ3,Δ2-enoyl-CoA isomerase. FEBS Letters, 322, 299–303.

Klensporfpawlik D, Szydlowski M, Kaczmarek A, Nowackawoszuk J, Switonski M, Jeleń H. 2012. The fatty acid composition of the Longissimus dorsi muscle, subcutaneous and visceral fats differ in four commercial pig breeds. Journal of Animal and Feed Sciences, 21, 661–676.

Lee S H, Lee S K, Paik D, Min K J. 2012. Overexpression of fatty-acid-β-oxidation-related genes extends the lifespan of Drosophila melanogaster. Oxidative Medicine and Cellular Longevity, 2012, 854502.

Li L. 2009. Genomic characterization and polymorphism analysis of genes involved in lipid and energy metabolism in swine. Ph D thesis, Technische Universität München, Germany.

Li Q, Wang Z, Zhang B, Lu Y, Yang Y, Ban D, Wu C, Zhang H. 2014. Single nucleotide polymorphism scanning and expression of the pig PPARGC1A gene in different breeds. Lipids, 49, 1047–1055.

Livak K J, Schmittgen T D. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT methods. Methods, 25, 402–408.

Lu Y, Li Q, Wang Z, Zhang B, Wu K, Zhang H. 2015.Polymorphisms of ECI1 gene and association with backfat thickness in New Huai fattening pig line group. Chinese Journal of Animal Science, 51, 1–3. (in Chinese)

Mursula A M, Aalten D M F V, Hiltunen J K, Wierenga R K. 2001. The crystal structure of Δ3-Δ2-enoyl-CoA isomerase. Journal of Molecular Biology, 309, 845–853.

Palosaari P M, Kilponen J M, Sormunen R T, Hassinen I E, Hiltunen J K. 1990. Δ3-Δ2-enoyl-CoA isomerases. Characterization of the mitochondrial isoenzyme in the rat. Journal of Biological Chemistry, 265, 3347–3353.

Partanen S T, Novikov D K, Popov A N, Mursula A M, Hiltunen J K, Wierenga R K. 2004. The 1.3 Å crystal structure of human mitochondrial Δ3-Δ2-enoyl-CoA isomerase shows a novel mode of binding for the fatty acyl group. Journal of Molecular Biology, 342, 1197–1208.

Plastow G S, Carrión D, Gil M, García-Regueiro J A, Furnols M F I, Gispert M, Oliver M A, Velarde A, Guàrdia M D, Hortós M, Rius M A, Sárraga C, Díaz I, Valero A, Sosnicki A, Klont R, Dornan S, Wilkinson J M, Evans G, Sargent C, et al. 2005. Quality pork genes and meat production. Meat Science, 70, 409–421

Rakhshandehroo M, Hooiveld G, Müller M, Kersten S. 2009. Comparative analysis of gene regulation by the transcription factor PPARα between mouse and human. PLoS One, 4, e6796.

Rakhshandehroo M, Sanderson L M, Matilainen M, Stienstra R, Carlberg C, de Groot P J, Müller M, Kersten S. 2007.Comprehensive analysis of PPARα-dependent regulation of hepatic lipid metabolism by expression pro?ling. PPAR Research, 2007, e26839.

Rasmussen A L, Diamond D L, McDermott J E, Gao X, Metz T O, Matzke M M, Carter V S, Belisle S E, Korth M J, Waters K M, Smith R D, Katze M G. 2011. Systems virology identifies a mitochondrial fatty acid oxidation enzyme, dodecenoyl coenzyme A delta isomerase, required for hepatitis C virus replication and likely pathogenesis. Journal of Virology, 85, 11646–11654.

Reiter S S, Halsey C H C, Stronach B M, Bartosh J L, Owsley W F, Bergen W G. 2007. Lipid metabolism related gene-expression profiling in liver, skeletal muscle and adipose tissue in crossbred Duroc and Pietrain pigs. Comparative Biochemistry and Physiology (Part D: Genomics and Proteomics), 2, 200–206.

Sambrook J, Russell D. 2001. Molecular Cloning: A Laboratry Manual. 3rd ed. Cold Spring Harbor Laboratory Press, New York.

Stoffel W, Ecker W. 1979. Δ3-cis,Δ2-trans-Enoyl-CoA isomerase from rat liver mitochondria. Methods in Enzymology, 14, 99–105.

Suzuki A, Kojima N, Ikeuchi Y, Ikarashi S, Moriyama N, Ishizuka T, Tokushige H. 1991. Carcass composition and meat quality of Chinese purebred and European×Chinese crossbred pigs. Meat Science, 29, 31–41.

Suzuki K, Irie M, Kadowaki H, Shibata T, Kumagai M, Nishida A. 2005. Genetic parameter estimates of meat quality traits in Duroc pigs selected for average daily gain, longissimus muscle area, backfat thickness, and intramuscular fat content. Journal of Animal Science, 83, 2058–2065.

Wang Z, Li Q G, Zhang B, Lu Y F, Yang Y Z, Ban D M, Zhang H. 2014. Single nucleotide polymorphism scanning and expression of the FRZB gene in pig populations. Gene, 543, 198–203.

van Weeghel M, te Brinke H, van Lenthe H, Kulik W, Minkler P E, Stoll MSK, Sass J O, Janssen U, Stoffel W, Schwab K O, Wanders R J A, Hoppel C L, Houten S M. 2012. Functional redundancy of mitochondrial enoyl-CoA isomerases in the oxidation of unsaturated fatty acids. FASEB Journal, 26, 4316–4326.

Yan D, Zhao G, Gou X, Wang T, Du Y, Li G, Lian L. 2007. Study on characteristics of Diqing Tibetan pig’s meat quality. Journal of Yunnan Agricultural University, 22, 86–89. (in Chinese)

Yang S Y, He X Y. 1999. Molecular mechanisms of fatty acid β-oxidation enzyme catalysis. In: Quant P A, Eaton S, eds., Current Views of Fatty Acid Oxidation and Ketogenesis: From Organelles to Point Mutations (Advances in Experimental Medicine and Biology).  E-Publishing, New York. pp. 133–143.

Zhou X, Huang W, Cao Z, Yang X, Zhang X, Gan W, Tao L. 2016. Nutrient composition comparison of longissimus dorsi between Diannan small-ear pigs and DLY commercial pigs. China Animal Husbandry & Veterinary Medicine, 43, 1743–1748. (in Chinese)

Zhou Y, Zhang X, Chen L, Wu J, Dang H, Wei M, Fan Y, Zhang Y, Zhu Y, Wang N, Breyer M D , Guan Y. 2008. Expression profiling of hepatic genes associated with lipid metabolism in nephritic rats. American Journal of Physiology (Renal Physiology), 295, 662–671.
[1] WANG Peng-fei, WANG Ming, SHI Zhi-bin, SUN Zhen-zhao, WEI Li-li, LIU Zai-si, WANG Shi-da, HE Xi-jun, WANG Jing-fei. Development of a recombinant pB602L-based indirect ELISA assay for detecting antibodies against African swine fever virus in pigs[J]. >Journal of Integrative Agriculture, 2022, 21(3): 819-825.
[2] SUN Li-na, LIU Yan-di, ZHANG Huai-jiang, YAN Wen-tao, YUE Qiang, QIU Gui-sheng. Molecular characterization of the ryanodine receptor from Adoxophyes orana and its response to lethal and sublethal doses of chlorantraniliprole[J]. >Journal of Integrative Agriculture, 2021, 20(6): 1585-1595.
[3] WANG Xiao-bo, WU Nan, CAI Rui-jie, GENG Wei-na, XU Xiao-yan. Changes in speciation, mobility and bioavailability of Cd, Cr and As during the transformation process of pig manure by black soldier fly larvae (Hermetia illucens)[J]. >Journal of Integrative Agriculture, 2021, 20(5): 1157-1166.
[4] ZHU Mei-chen, HU Ran, ZHAO Hui-yan, TANG Yun-shan, SHI Xiang-tian, JIANG Hai-yan, ZHANG Zhi-yuan, FU Fu-you, XU Xin-fu, TANG Zhang-lin, LIU Lie-zhao, LU Kun, LI Jia-na, QU Cun-min. Identification of quantitative trait loci and candidate genes controlling seed pigments of rapeseed[J]. >Journal of Integrative Agriculture, 2021, 20(11): 2862-2879.
[5] Iram SHAFIQ, Sajad HUSSAIN, Muhammad Ali RAZA, Nasir IQBAL, Muhammad Ahsan ASGHAR, Ali RAZA, FAN Yuan-fang, Maryam MUMTAZ, Muhammad SHOAIB, Muhammad ANSAR, Abdul MANAF, YANG Wen-yu, YANG Feng. Crop photosynthetic response to light quality and light intensity[J]. >Journal of Integrative Agriculture, 2021, 20(1): 4-23.
[6] LI Cen-cen, YU Shu-long, REN Hai-feng, WU Wei, WANG Ya-ling, HAN Qiu, XU Hai-xia, XU Yong-jie, ZHANG Peng-peng. Identification and functional prediction of long intergenic noncoding RNAs in fetal porcine longissimus dorsi muscle[J]. >Journal of Integrative Agriculture, 2021, 20(1): 201-211.
[7] DAI Xiao-wen, Zhanli SUN, Daniel MÜLLER. Driving factors of direct greenhouse gas emissions from China’s pig industry from 1976 to 2016[J]. >Journal of Integrative Agriculture, 2021, 20(1): 319-329.
[8] SHAN Yan-ju, JI Gai-ge, ZOU Jian-min, ZHANG Ming, TU Yun-jie, LIU Yi-fan, JU Xiao-jun, SHU Jing-ting. PGC-1α differentially regulates the mRNA expression profiles of genes related to myofiber type specificity in chicken[J]. >Journal of Integrative Agriculture, 2020, 19(8): 2083-2094.
[9] NAN Jiu-hong, YIN Li-lin, TANG Zhen-shuang, CHEN Jian-hai, ZHANG Jie, WANG Hai-yan, DU Xiao-yong, LIU Xiang-dong . Genetic parameter estimation and genome-wide association study (GWAS) of red blood cell count at three stages in a Duroc×Erhualian pig population[J]. >Journal of Integrative Agriculture, 2020, 19(3): 793-799.
[10] ZHUO Ni, JI Chen, DING Jing-yu. Pig farmers’ willingness to recover their production under COVID-19 pandemic shock in China-Empirical evidence from a farm survey[J]. >Journal of Integrative Agriculture, 2020, 19(12): 2891-2902.
[11] ZHENG Yao, CHEN Cai, CHEN Wei, WANG Xiao-yan, WANG Wei, GAO Bo, Klaus WIMMERS, MAO Jiu-de, SONG Cheng-yi. Two new SINE insertion polymorphisms in pig Vertnin (VRTN) gene revealed by comparative genomic alignment[J]. >Journal of Integrative Agriculture, 2020, 19(10): 2514-2522.
[12] WANG Man, YU Bing, HE Jun, YU Jie, LUO Yu-heng, LUO Jun-qiu, MAO Xiang-bin, CHEN Dai-wen. The toxicological effect of dietary excess of saccharicterpenin, the extract of camellia seed meal, in piglets[J]. >Journal of Integrative Agriculture, 2020, 19(1): 211-224.
[13] ZENG Zhu, JIANG Jun-jie, YU Jie, MAO Xiang-bing, YU Bing, CHEN Dai-wen. Effect of dietary supplementation with mulberry (Morus alba L.) leaves on the growth performance, meat quality and antioxidative capacity of finishing pigs[J]. >Journal of Integrative Agriculture, 2019, 18(1): 143-151.
[14] SHAN Nan, LI Hu, LI Jian-zheng, Ee Ling Ng, MA Yan, WANG Li-gang, CHEN Qing. A major pathway for carbon and nitrogen losses- Gas emissions during storage of solid pig manure in China[J]. >Journal of Integrative Agriculture, 2019, 18(1): 190-200.
[15] JI Chen, CHEN Qin, Jacques Trienekens, WANG Hai-tao. Determinants of cooperative pig farmers' safe production behaviour in China – Evidences from perspective of cooperatives' services[J]. >Journal of Integrative Agriculture, 2018, 17(10): 2345-2355.
No Suggested Reading articles found!