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Journal of Ophthalmology
Volume 2015, Article ID 816329, 7 pages
Research Article

Knocking Down Snrnp200 Initiates Demorphogenesis of Rod Photoreceptors in Zebrafish

1Department of Ophthalmology, The First Affiliated Hospital of Nanjing Medical University and State Key Laboratory of Reproductive Medicine, Nanjing Medical University, Nanjing 210029, China
2Department of Ophthalmology, The First People’s Hospital of Suqian, Suqian 223000, China
3Department of Ophthalmology, Tianjin Medical University, Tianjin Eye Hospital, Tianjin Key Laboratory of Ophthalmology and Visual Science, Tianjin 300040, China
4Model Animal Research Center, MOE Key Laboratory of Model Animal for Disease Study, Nanjing University, Nanjing 210061, China
5Department of Molecular Genetics and Cell Biology, University of Chicago, 920 E. 58th Street, Chicago, IL 60637, USA

Received 31 October 2014; Revised 17 December 2014; Accepted 29 December 2014

Academic Editor: Patrik Schatz

Copyright © 2015 Yuan Liu et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Supplementary Material

Morpholino Oligos and Knockdown of Snrnp200 Standard control morpholino oligos (control-MO: 5’-CCTCTTACCTCAGTTACAATTTATA-3’) and zebrafish Snrnp200 splicing-blocking MO targeting exon 25 (Snrnp200-MO: 5’-TCAACATCAAGACAACTCACATCCT-3’) were purchased from Gene Tools (Philomath, OR, USA). Zebrafish embryos were injected 3 at the 1- or 2-cell stage (0 day postfertilization (dpf)) with ∼ 1 nL of purified with ∼ 1 nl of purified MOs dissolved in water. To get the best adjusted dosage for Snrnp200-MO, zebrafish embryos were divided into three groups, and injected with 2 (n=67), 4 (n=68), and 8 ng (n=65) of MOs, respectively. The counting and the percentage calculation of deformation and death in each injected group were conducted from 2 to 4 dpf as described previously. Embryos died within 24 hr post fertilization were excluded because such death was likely resulted from unspecific causes. At 4 dpf, embryos with relatively normal appearance were collected from each injected group for further investigations.

RT-PCR: RNA was isolated from 10 zebrafish embryos from each injection group, followed by reverse-transcriptase polymerase chain reaction (RT-PCR) to generate cDNA template. PCR was subsequently conducted on the obtained cDNA to verify the effectiveness of Snrnp200-MO.

  1. Supplementary Material