Abstract
This paper introduces an innovative method for genotyping 1–2 days old zebrafish embryos, without sacrificing the life/health of the embryos. The method utilizes microfluidic technology to extract and collect a small amount of genetic material from the chorionic fluid or fin tissue of the embryo. Then, using conventional DNA extraction, PCR amplification, and high resolution melt analysis with fluorescent DNA detection techniques, the embryo is genotyped. The chorionic fluid approach was successful 78 % of the time while the fin clipping method was successful 100 % of the time. Chorionic fluid was shown to only contain DNA from the embryo and not from the mother. These results suggest a novel method to genotype zebrafish embryos that can facilitate high-throughput screening, while maintaining 100 % viability of the embryo.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
T. Bansal, J. Lenhart, T. Kim, C. Duan, M.M. Maharbiz, Biomed. Microdevices 11, 633–641 (2009). doi:10.1007/s10544-008-9273-5
D. Choudhury, D. van Noort, C. Iliescu, B. Zheng, K.-L. Poon, S. Korzh, V. Korzh, H. Yu, Lab Chip 12(5), 892–900 (2012). doi:10.1039/c11c20351g
D.C. Duffy, J.C. McDonald, O.J.A. Schueller, G.M. Whitesides, Anal. Chem. 70, 4974–4984 (1998). doi:10.1021/ac980656z
E. Fujimoto, T.J. Stevenson, C.B. Chien, J.L. Bonkowsky, Dev. Biol. 352, 393–404 (2011). doi:10.1016/j.ydbio.2011.01.023
K. Howe et al., Nature 496, 498–503 (2013). doi:10.1038/nature12111
D.-H. Kim, Y. Sun, S. Yun, B. Kim, C.N. Hwang, S.H. Lee, B. Nelson, Conf. Proc. IEEE Eng. Med. Biol. Soc. 7, 5061–5064 (2004)
K.M. Kwan, E. Fujimoto, C. Grabher, B.D. Mangum, M.E. Hardy, D.S. Campbell, J.M. Parant, H.J. Yost, J.P. Kanki, C.-B. Chien, Dev. Dyn. 236, 3088–3099 (2007). doi:10.1002/dvdy.21343
G. Lieschke, P.D. Currie, Nat. Rev. Genet. 8, 353–367 (2007). doi:10.1038/nrg2091
M. Westerfield. 5th ed. University of Oregon Press, Eugene, (2007)
J.M. Parant, S.A. George, R. Pryor, C.T. Wittwer, H.J. Yost, Dev. Dyn. 238, 3168–3174 (2009). doi:10.1002/dvdy.22143
R. Samuel, C.M. Thacker, A.V. Maricq, B.K. Gale, J. Micromech. Microeng. 24, 105007 (2014). doi:10.1088/0960-1317/24/10/105007
S.U. Son, R.L. Garrell, Lab Chip 9, 2398–2401 (2009). doi:10.1039/b906257b
W. Wang, X. Liu, D. Gelinas, B. Ciruna, Y. Sun, PLoS One 2(9), e862 (2007). doi:10.1371/journal.pone.0000862
E.M. Wielhouwer, S. Ali, A. Al-Afandi, M.T. Blom, M.B. Olde Riekerink, C. Poelma, J. Westerweel, J. Oonk, E.X. Vrouwe, W. Buesink, H.G.J. van Mil, J. Chicken, R. van’t Oever, M.K. Richardson, Lab Chip 11, 1815–1824 (2011). doi:10.1039/c01c00443j
F. Yang, Z. Chen, J. Pan, X. Li, J. Feng, H. Yang, Biomicrofluidics 5, 024115 (2011). doi:10.1063/1.3605509
Acknowledgments
JLB and RS were supported by NIH DP2 MH100008. The authors would like to thank the Wittwer Lab, and the State of Utah Center of Excellence in Biomedical Microfluidics at the University of Utah, for providing technical resources and expertise.
Conflict of interest
The authors declare that they have no conflict of interest.
Ethical approval
“All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.”
Author information
Authors and Affiliations
Corresponding authors
Electronic supplementary material
Below is the link to the electronic supplementary material.
ESM 1
(DOCX 2075 kb)
(MOV 968 kb)
(MOV 813 kb)
Rights and permissions
About this article
Cite this article
Samuel, R., Stephenson, R., Roy, P. et al. Microfluidic-aided genotyping of zebrafish in the first 48 h with 100 % viability. Biomed Microdevices 17, 43 (2015). https://doi.org/10.1007/s10544-015-9946-9
Published:
DOI: https://doi.org/10.1007/s10544-015-9946-9