AAS ›› 2016, Vol. ›› Issue (2): 284-288.doi: 10.16098/j.issn.0529-1356.2016.02.024

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A method of simultaneously tracing the trajectory of bilateral commissural axons in developing chick spinal cord using dual-color fluorescence

YU Ya-nan1 LIU Yan-li 1,2 YANG Ci-qing 1,2 ZHANG Bi-chao1 XU Zhen-ping 1,2* LIN Jun-tang 1,2*   

  1. 1. Collage of Life Science and Technology, Xinxiang Medical University, He’nan Xinxiang 453003,China; 2. He’nan Key Laboratory of Medical Tissue Regeneration, Xinxiang Medical University, He’nan Xinxiang 453003,China
  • Received:2015-09-18 Revised:2015-11-01 Online:2016-04-06 Published:2016-04-06
  • Contact: XU Zhen-ping; LIN Jun-tang E-mail:linjtlin@126.com
  • Supported by:

    ;Graduate student research and innovation project

Abstract:

Objective To develop a method of simultaneously tracing the development of commissural axons from bilateral chick spinal cord using dual-color fluorescent.
Methods The pCAGGS-GFP plasmid was transfected into the chick spinal cord using ovo electroporation after incubation of fertilized eggs for HH stage17-18. GFP-positive chick embryos were obtained at day 6. At least three samples were collected for each group. The collected specimens were undergone either transverse section or “open-book” processing. In “open-book” processing, the spinal cord was unfolded and placed on the glass slide. After injecting DiI solution into the contralateral side of transfected region, the samples were incubated for 3d at 4℃, and commissural axons were observed under a fluorescent microscopy. Results The results of GFP labeling from both transverse section and open-book processing showed that commissural axons crossed the midline region through the floor plate to the contralateral side of the spinal cord, then they projected rostrally and caudally along the antero-caudal (AP) axis within ventral fasciculus and lateral fasciculus. DiI tracing showed similar trajectory of commissural axons. Conclusion Dual-color fluorescent tracing method is successfully established, which may provide a new method for the study of development of commissural axons of the spinal cord.