Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

Efficient generation of sFat-1 transgenic rabbits rich in n-3 polyunsaturated fatty acids by intracytoplasmic sperm injection

Shun Zhang A , Fenghua Lu A , Qingyou Liu A , Yubing Liu A , Xiaomei Guan A , Yingming Wei A , Shijian Tan A and Deshun Shi A B
+ Author Affliations
- Author Affliations

A State Key Laboratory for Conservation and Utilisation of Subtropical Agro-Bioresources, Guangxi University, Nanning 530004, China.

B Corresponding author. Email: ardsshi@gxu.edu.cn

Reproduction, Fertility and Development 28(3) 310-318 https://doi.org/10.1071/RD13413
Submitted: 14 December 2013  Accepted: 20 May 2014   Published: 16 July 2014

Abstract

N-3 polyunsaturated fatty acids (n-3 PUFAs) have their first double bond at the third carbon from the methyl end of the fatty-acid chain and had been proven to be beneficial to human health. However, mammals cannot produce n-3 PUFAs by themselves because they lack the n-3 fatty-acid desaturase (Fat-1) gene. Thus, the possibility of producing sFat-1 transgenic rabbits was explored in this study. The transgenic cassette of pPGK1–sFat-1–CMV–EGFP was constructed and transgenic rabbit embryos were produced by intracytoplasmic sperm injection (ICSI). When 123 EGFP-positive embryos at the 2–8-cell stage were transplanted into the oviduct of four oestrous-synchronised recipients, two of them became pregnant and gave birth to seven pups. However, transfer of embryos into the uterus of oestrous-synchronised recipients and oviduct or uterus of oocyte donor rabbits did not result in pregnancy. The integration of the sFat-1 gene was confirmed in six of the seven live pups by PCR and Southern blot. The expression of the sFat-1 gene in the six transgenic pups was also detected by reverse transcription polymerase chain reaction (RT-PCR). Gas chromatography–mass spectrometry analysis revealed that transgenic rabbits exhibited an ~15-fold decrease in the ratio of n-6 : n-3 PUFAs in muscle compared with wild-type rabbits and non-transgenic rabbits. These results demonstrate that sFat-1 transgenic rabbits can be produced by ICSI and display a low ratio of n-6 : n-3 PUFAs.

Additional keywords: embryogenesis, embryo transfer, pregnancy, transgenesis.


References

Ali, S., Garg, S. K., Cohen, B. E., Bhave, P., Harris, W. S., and Whooley, M. A. (2009). Association between omega-3 fatty acids and depressive symptoms among patients with established coronary artery disease: data from the Heart and Soul Study. Psychother. Psychosom. 78, 125–127.
Association between omega-3 fatty acids and depressive symptoms among patients with established coronary artery disease: data from the Heart and Soul Study.CrossRef | 19223688PubMed | open url image1

Bousquet, M., Gue, K., Emond, V., Julien, P., Kang, J. X., Cicchetti, F., and Calon, F. (2011). Transgenic conversion of omega-6 into omega-3 fatty acids in a mouse model of Parkinson’s disease. J. Lipid Res. 52, 263–271.
Transgenic conversion of omega-6 into omega-3 fatty acids in a mouse model of Parkinson’s disease.CrossRef | 1:CAS:528:DC%2BC3MXht1alurw%3D&md5=e6ae9d7760aa362459afe5aa5fed4f17CAS | 21115966PubMed | open url image1

Buckley, J. D., and Howe, P. R. (2009). Anti-obesity effects of long-chain omega-3 polyunsaturated fatty acids. Obes. Rev. 10, 648–659.
Anti-obesity effects of long-chain omega-3 polyunsaturated fatty acids.CrossRef | 1:CAS:528:DC%2BD1MXhsFKrsr7N&md5=ea50b8e2f718c9845365947bfb51c501CAS | 19460115PubMed | open url image1

Certik, M., and Shimizu, S. (1999). Biosynthesis and regulation of microbial polyunsaturated fatty-acid production. J. Biosci. Bioeng. 87, 1–14.
Biosynthesis and regulation of microbial polyunsaturated fatty-acid production.CrossRef | 1:CAS:528:DyaK1MXhsl2mtbs%3D&md5=72badc3dbbe72bd0d38b565c44cf18c4CAS | 16232418PubMed | open url image1

Clayton, E. H., Hanstock, T. L., Hirneth, S. J., Kable, C. J., Garg, M. L., and Hazell, P. L. (2009). Reduced mania and depression in juvenile bipolar disorder associated with long-chain omega-3 polyunsaturated fatty-acid supplementation. Eur. J. Clin. Nutr. 63, 1037–1040.
Reduced mania and depression in juvenile bipolar disorder associated with long-chain omega-3 polyunsaturated fatty-acid supplementation.CrossRef | 1:CAS:528:DC%2BD1MXps1Wls78%3D&md5=cfdee9fdda31cd0da30c2075b7c81b33CAS | 19156158PubMed | open url image1

Conklin, S. M., Runyan, C. A., Leonard, S., Reddy, R. D., Muldoon, M. F., and Yao, J. K. (2010). Age-related changes of n-3 and n-6 polyunsaturated fatty acids in the anterior cingulate cortex of individuals with major depressive disorder. Prostaglandins Leukot. Essent. Fatty Acids 82, 111–119.
Age-related changes of n-3 and n-6 polyunsaturated fatty acids in the anterior cingulate cortex of individuals with major depressive disorder.CrossRef | 1:CAS:528:DC%2BC3cXitVKnurg%3D&md5=d89c23ec6ff7a30bc0b8c9135d4a1fe2CAS | 20060277PubMed | open url image1

Covington, M. B. (2004). Omega-3 fatty acids. Am. Fam. Physician 70, 133–140.
| 15259529PubMed | open url image1

De Truchis, P., Kirstetter, M., Perier, A., Meunier, C., Zucman, D., Force, G., Doll, J., Katlama, C., Rozenbaum, W., Masson, H., Gardette, J., and Melchior, J. C. (2007). Reduction in triglyceride level with N-3 polyunsaturated fatty acids in HIV-infected patients taking potent antiretroviral therapy: a randomised prospective study. J. Acquir. Immune Defic. Syndr. 44, 278–285.
Reduction in triglyceride level with N-3 polyunsaturated fatty acids in HIV-infected patients taking potent antiretroviral therapy: a randomised prospective study.CrossRef | 1:CAS:528:DC%2BD2sXitFyqtr4%3D&md5=abfc865773a77f141f868e352f8fdd6fCAS | 17179770PubMed | open url image1

FAOSTATS 2012. Available at http://faostat.fao.org [Verified 23 June 2014].

Folch, J., Lees, M., and Sloane Stanley, G. H. (1957). A simple method for the isolation and purification of total lipids from animal tissues. J. Biol. Chem. 226, 497–509.
| 1:STN:280:DyaG2s%2FnsFCjtw%3D%3D&md5=7af8b449eecca02927736f072881bbefCAS | 13428781PubMed | open url image1

García-Vázquez, F. A., Garcia-Rosello, E., Gutierrez-Adan, A., and Gadea, J. (2009). Effect of sperm treatment on efficiency of EGFP-expressing porcine embryos produced by ICSI-SMGT. Theriogenology 72, 506–518.
Effect of sperm treatment on efficiency of EGFP-expressing porcine embryos produced by ICSI-SMGT.CrossRef | 19501394PubMed | open url image1

Guo, T., Liu, X. F., Ding, X. B., Yang, F. F., Nie, Y. W., An, Y. J., and Guo, H. (2011). Fat-1 transgenic cattle as a model to study the function of omega-3 fatty acids. Lipids Health Dis. 10, 244.
Fat-1 transgenic cattle as a model to study the function of omega-3 fatty acids.CrossRef | 1:CAS:528:DC%2BC38XisFert7g%3D&md5=c6cee1e3081be24972f6257e50c6c513CAS | 22206437PubMed | open url image1

Innis, S. M. (2007). Dietary (n-3) fatty acids and brain development. J. Nutr. 137, 855–859.
| 1:CAS:528:DC%2BD2sXktVSlt7Y%3D&md5=9e789c927a4827a4481dc5bbfce3a84bCAS | 17374644PubMed | open url image1

Issa, A. M., Mojica, W. A., Morton, S. C., Traina, S., Newberry, S. J., Hilton, L. G., Garland, R. H., and Maclean, C. H. (2006). The efficacy of omega-3 fatty acids on cognitive function in ageing and dementia: a systematic review. Dement. Geriatr. Cogn. Disord. 21, 88–96.
The efficacy of omega-3 fatty acids on cognitive function in ageing and dementia: a systematic review.CrossRef | 1:CAS:528:DC%2BD28XksVyguw%3D%3D&md5=98df4c6b4ba43595106cac77f6ccb208CAS | 16340205PubMed | open url image1

Kang, J. X., Wang, J., Wu, L., and Kang, Z. B. (2004). Transgenic mice: fat-1 mice convert n-6 to n-3 fatty acids. Nature 427, 504.
Transgenic mice: fat-1 mice convert n-6 to n-3 fatty acids.CrossRef | 1:CAS:528:DC%2BD2cXpsFWhtA%3D%3D&md5=1304c18a8fbf0ce471d1c450bc6f4b90CAS | 14765186PubMed | open url image1

Kannass, K. N., Colombo, J., and Carlson, S. E. (2009). Maternal DHA levels and toddler free-play attention. Dev. Neuropsychol. 34, 159–174.
Maternal DHA levels and toddler free-play attention.CrossRef | 19267293PubMed | open url image1

Kuo, Y. T., So, P. W., Parkinson, J. R., Yu, W. S., Hankir, M., Herlihy, A. H., Goldstone, A. P., Frost, G. S., Wasserfall, C., and Bell, J. D. (2010). The combined effects on neuronal activation and blood–brain barrier permeability of time and n-3 polyunsaturated fatty acids in mice, as measured in vivo using MEMRI. Neuroimage 50, 1384–1391.
The combined effects on neuronal activation and blood–brain barrier permeability of time and n-3 polyunsaturated fatty acids in mice, as measured in vivo using MEMRI.CrossRef | 1:CAS:528:DC%2BC3cXltFGgs7s%3D&md5=89878d0d540104f22990e4e5129c09ccCAS | 20097292PubMed | open url image1

Lacham-Kaplan, O., Shaw, J., Sanchez-Partida, L. G., and Trounson, A. (2003). Oocyte activation after intracytoplasmic injection with sperm frozen without cryoprotectants results in live offspring from inbred and hybrid mouse strains. Biol. Reprod. 69, 1683–1689.
Oocyte activation after intracytoplasmic injection with sperm frozen without cryoprotectants results in live offspring from inbred and hybrid mouse strains.CrossRef | 1:CAS:528:DC%2BD3sXosV2rs74%3D&md5=3b60b3f3450b1ec5236749cd2e025541CAS | 12890731PubMed | open url image1

Lai, L., Kang, J. X., Li, R., Wang, J., Witt, W. T., Yong, H. Y., Hao, Y., Wax, D. M., Murphy, C. N., Rieke, A., Samuel, M., Linville, M. L., Korte, S. W., Evans, R. W., Starzl, T. E., Prather, R. S., and Dai, Y. (2006). Generation of cloned transgenic pigs rich in omega-3 fatty acids. Nat. Biotechnol. 24, 435–436.
Generation of cloned transgenic pigs rich in omega-3 fatty acids.CrossRef | 1:CAS:528:DC%2BD28Xjt1Wisb4%3D&md5=0ed7f01e0027fd57446c8bf68b9a1391CAS | 16565727PubMed | open url image1

Li, Q., Hou, J., Wang, S., Chen, Y., and An, X. R. (2010). Production of transgenic rabbit embryos through intracytoplasmic sperm injection. Zygote 18, 301–307.
Production of transgenic rabbit embryos through intracytoplasmic sperm injection.CrossRef | 20663236PubMed | open url image1

Lim, G. P., Calon, F., Morihara, T., Yang, F., Teter, B., Ubeda, O., Salem, N., Frautschy, S. A., and Cole, G. M. (2005). A diet enriched with the omega-3 fatty acid docosahexaenoic acid reduces amyloid burden in an aged Alzheimer mouse model. J. Neurosci. 25, 3032–3040.
A diet enriched with the omega-3 fatty acid docosahexaenoic acid reduces amyloid burden in an aged Alzheimer mouse model.CrossRef | 1:CAS:528:DC%2BD2MXivFGhs70%3D&md5=f9eb44e4a11d032c9ac482d12d9fe9ffCAS | 15788759PubMed | open url image1

Liu, A., Chang, J., Lin, Y., Shen, Z., and Bernstein, P. S. (2010). Long-chain and very long-chain polyunsaturated fatty acids in ocular ageing and age-related macular degeneration. J. Lipid Res. 51, 3217–3229.
Long-chain and very long-chain polyunsaturated fatty acids in ocular ageing and age-related macular degeneration.CrossRef | 1:CAS:528:DC%2BC3cXhsVWgt7rI&md5=e6e08658832c934b19f48fe7ccb5f172CAS | 20688753PubMed | open url image1

Ma, D. W., Ngo, V., Huot, P. S., and Kang, J. X. (2006). N-3 polyunsaturated fatty acids endogenously synthesised in fat-1 mice are enriched in the mammary gland. Lipids 41, 35–39.
N-3 polyunsaturated fatty acids endogenously synthesised in fat-1 mice are enriched in the mammary gland.CrossRef | 1:CAS:528:DC%2BD28XitFCmsLk%3D&md5=4bbd733344e39ea6b46f0949d994fe60CAS | 16555469PubMed | open url image1

Marik, P. E., and Varon, J. (2009). Omega-3 dietary supplements and the risk of cardiovascular events: a systematic review. Clin. Cardiol. 32, 365–372.
Omega-3 dietary supplements and the risk of cardiovascular events: a systematic review.CrossRef | 19609891PubMed | open url image1

Ménesi, D., Kitajka, K., Molnár, E., Kis, Z., Belleger, J., Narce, M., Kang, J. X., Puskás, L. G., and Das, U. N. (2009). Gene and protein expression profiling of the fat-1 mouse brain. Prostaglandins Leukot. Essent. Fatty Acids 80, 33–42.
Gene and protein expression profiling of the fat-1 mouse brain.CrossRef | 19138887PubMed | open url image1

Moreira, P. N., Pozueta, J., Perez-Crespo, M., Valdivieso, F., Gutierrez-Adan, A., and Montoliu, L. (2007). Improving the generation of genomic-type transgenic mice by ICSI. Transgenic Res. 16, 163–168.
Improving the generation of genomic-type transgenic mice by ICSI.CrossRef | 1:CAS:528:DC%2BD2sXjt1Sqt7Y%3D&md5=fed5f42533442c920fcc1192d8ea5fa2CAS | 17372844PubMed | open url image1

Niu, Y., and Liang, S. (2008). Progress in gene transfer by germ cells in mammals. J. Genet. Genomics 35, 701–714.
Progress in gene transfer by germ cells in mammals.CrossRef | 1:CAS:528:DC%2BD1MXjsVegs70%3D&md5=19ee45723431c36b4aff70ee17a5a29cCAS | 19103425PubMed | open url image1

Pan, D., Zhang, L., Zhou, Y., Feng, C., Long, C., Liu, X., Wan, R., Zhang, J., Lin, A., Dong, E., Wang, S., Xu, H., and Chen, H. (2010). Efficient production of omega-3 fatty-acid desaturase (sFat-1)-transgenic pigs by somatic cell nuclear transfer. Sci. China Life Sci. 53, 517–523.
Efficient production of omega-3 fatty-acid desaturase (sFat-1)-transgenic pigs by somatic cell nuclear transfer.CrossRef | 1:CAS:528:DC%2BC3cXlslWntLw%3D&md5=bf97492a190be57ed5ce0bda5b5175a8CAS | 20596920PubMed | open url image1

Perry, A. C., Wakayama, T., Kishikawa, H., Kasai, T., Okabe, M., Toyoda, Y., and Yanagimachi, R. (1999). Mammalian transgenesis by intracytoplasmic sperm injection. Science 284, 1180–1183.
Mammalian transgenesis by intracytoplasmic sperm injection.CrossRef | 1:CAS:528:DyaK1MXjtlajsbk%3D&md5=1b1f4e6c2c8eb03fd9654b469839af01CAS | 10325231PubMed | open url image1

Pohlmeier, W. E., Hovey, R. C., and Van Eenennaam, A. L. (2011). Reproductive abnormalities in mice expressing omega-3 fatty-acid desaturase in their mammary glands. Transgenic Res. 20, 283–292.
Reproductive abnormalities in mice expressing omega-3 fatty-acid desaturase in their mammary glands.CrossRef | 1:CAS:528:DC%2BC3MXivVGhsrk%3D&md5=3d40cd97f5dab9f970b5e8cc969753c7CAS | 20532624PubMed | open url image1

Richards, M. P., Kathirvel, P., Gong, Y., Lopez-Hernandez, A., Walters, E. M., and Prather, R. S. (2011). Long-chain omega-3 fatty-acid levels in loin muscle from transgenic (fat-1 gene) pigs and effects on lipid oxidation during storage. Food Biotechnology 25, 103–114.
Long-chain omega-3 fatty-acid levels in loin muscle from transgenic (fat-1 gene) pigs and effects on lipid oxidation during storage.CrossRef | 1:CAS:528:DC%2BC3MXmtlejtro%3D&md5=d200357a8b96412dbb75546e8e8fa332CAS | open url image1

Rubin, M. G., Kim, K., and Logan, A. C. (2008). Acne vulgaris, mental health and omega-3 fatty acids: a report of cases. Lipids Health Dis. 7, 36.
Acne vulgaris, mental health and omega-3 fatty acids: a report of cases.CrossRef | 18851733PubMed | open url image1

Saeki, K., Matsumoto, K., Kinoshita, M., Suzuki, I., Tasaka, Y., Kano, K., Taguchi, Y., Mikami, K., Hirabayashi, M., Kashiwazaki, N., Hosoi, Y., Murata, N., and Iritani, A. (2004). Functional expression of a Delta12 fatty-acid desaturase gene from spinach in transgenic pigs. Proc. Natl. Acad. Sci. USA 101, 6361–6366.
Functional expression of a Delta12 fatty-acid desaturase gene from spinach in transgenic pigs.CrossRef | 1:CAS:528:DC%2BD2cXjvVyitrY%3D&md5=06353dff56d760adacb058e5a31f4a1fCAS | 15067141PubMed | open url image1

Sessler, A. M., and Ntambi, J. M. (1998). Polyunsaturated fatty-acid regulation of gene expression. J. Nutr. 128, 923–926.
| 1:CAS:528:DyaK1cXjs1Wgs70%3D&md5=cd42bf1ced15c028be4d2f4a8545388cCAS | 9614148PubMed | open url image1

Simopoulos, A. P. (2002). The importance of the ratio of omega-6/omega-3 essential fatty acids. Biomed. Pharmacother. 56, 365–379.
The importance of the ratio of omega-6/omega-3 essential fatty acids.CrossRef | 1:CAS:528:DC%2BD38XotVKrtbY%3D&md5=7f7e3a094b81234fb2299ea438c9d261CAS | 12442909PubMed | open url image1

Simopoulos, A. P., Leaf, A., and Salem, N. (2000). Workshop statement on the essentiality of and recommended dietary intakes for omega-6 and omega-3 fatty acids. Prostaglandins Leukot. Essent. Fatty Acids 63, 119–121.
Workshop statement on the essentiality of and recommended dietary intakes for omega-6 and omega-3 fatty acids.CrossRef | 1:CAS:528:DC%2BD3cXnslCrt7w%3D&md5=7f7e105faabd1e5323d63dbe2a05e1beCAS | 10991764PubMed | open url image1

Smith, B. K., Holloway, G. P., Reza-Lopez, S., Jeram, S. M., Kang, J. X., and Ma, D. W. (2010). A decreased n-6/n-3 ratio in the fat-1 mouse is associated with improved glucose tolerance. Appl. Physiol. Nutr. Metab. 35, 699–706.
A decreased n-6/n-3 ratio in the fat-1 mouse is associated with improved glucose tolerance.CrossRef | 1:CAS:528:DC%2BC3cXhtlWlsr3J&md5=7a60c67f681e821aaf08e43a41b5c3aeCAS | 20962926PubMed | open url image1

Song, C., Li, X., Leonard, B. E., and Horrobin, D. F. (2003). Effects of dietary n-3 or n-6 fatty acids on interleukin-1beta-induced anxiety, stress and inflammatory responses in rats. J. Lipid Res. 44, 1984–1991.
Effects of dietary n-3 or n-6 fatty acids on interleukin-1beta-induced anxiety, stress and inflammatory responses in rats.CrossRef | 1:CAS:528:DC%2BD3sXos1Giu7Y%3D&md5=3f01716cd767bd64bd2378e7920c1b86CAS | 12837849PubMed | open url image1

Spychalla, J. P., Kinney, A. J., and Browse, J. (1997). Identification of an animal omega-3 fatty-acid desaturase by heterologous expression in Arabidopsis. Proc. Natl. Acad. Sci. USA 94, 1142–1147.
Identification of an animal omega-3 fatty-acid desaturase by heterologous expression in Arabidopsis.CrossRef | 1:CAS:528:DyaK2sXhtlGhsb4%3D&md5=d00a3d8517437aad22371011a89882f0CAS | 9037020PubMed | open url image1

van Gelder, B. M., Tijhuis, M., Kalmijn, S., and Kromhout, D. (2007). Fish consumption, n-3 fatty acids and subsequent 5-y cognitive decline in elderly men: the Zutphen Elderly Study. Am. J. Clin. Nutr. 85, 1142–1147.
| 1:CAS:528:DC%2BD2sXks1OgsLc%3D&md5=9c9cdf6518f14f28085f056478022a48CAS | 17413117PubMed | open url image1

Watanabe, M., Kurome, M., Matsunari, H., Nakano, K., Umeyema, K., Shiota, A., Nakauchi, H., and Nagashima, H. (2012). The creation of transgenic pigs expressing human proteins using BAC-derived, full-length genes and intracytoplasmic sperm injection-mediated gene transfer. Transgenic Res. 21, 605–618.
The creation of transgenic pigs expressing human proteins using BAC-derived, full-length genes and intracytoplasmic sperm injection-mediated gene transfer.CrossRef | 1:CAS:528:DC%2BC38Xms12gtLw%3D&md5=45b4228e718ce9cc1230f0e100777512CAS | 22038447PubMed | open url image1

Wright, R. W., Bondioli, K., Grammer, J., Kuzan, F., and Menino, A. (1981). FSH or FSH plus LH superovulation in ewes following oestrus synchronisation with medoxyprogesterone acetate pessaries. J. Anim. Sci. 52, 115–118.
| 1:CAS:528:DyaL3MXhtFCqurs%3D&md5=d84dd39744e7f2275a9e7ec7a3c36113CAS | 6787006PubMed | open url image1

Wu, X., Ouyang, H., Duan, B., Pang, D., Zhang, L., Yuan, T., Xue, L., Ni, D., Cheng, L., Dong, S., Wei, Z., Li, L., Yu, M., Sun, Q. Y., Chen, D. Y., Lai, L., Dai, Y., and Li, G. P. (2012). Production of cloned transgenic cow expressing omega-3 fatty acids. Transgenic Res. 21, 537–543.
Production of cloned transgenic cow expressing omega-3 fatty acids.CrossRef | 1:CAS:528:DC%2BC38Xms12gsbs%3D&md5=902014dc05f88ba7f5b2aafa458473ceCAS | 21918821PubMed | open url image1

Zhu, G., Chen, H., Wu, X., Zhou, Y., Lu, J., Chen, H., and Deng, J. (2008). A modified n-3 fatty-acid desaturase gene from Caenorhabditis briggsae produced high proportion of DHA and DPA in transgenic mice. Transgenic Res. 17, 717–725.
A modified n-3 fatty-acid desaturase gene from Caenorhabditis briggsae produced high proportion of DHA and DPA in transgenic mice.CrossRef | 1:CAS:528:DC%2BD1cXnvFyitb0%3D&md5=a416fcaf1d761da501f276d552f93b27CAS | 18322818PubMed | open url image1



Rent Article (via Deepdyve) Export Citation