ZFIN ID: ZDB-PUB-090112-16
Patterned delivery and expression of gene constructs into zebrafish embryos using microfabricated interfaces
Bansal, T., Lenhart, J., Kim, T., Duan, C., and Maharbiz, M.M.
Date: 2009
Source: Biomedical Microdevices   11(3): 633-641 (Journal)
Registered Authors: Duan, Cunming
Keywords: Spatio-temporal, Electroporation, DNA, Microfabricated, Zebrafish
MeSH Terms:
  • Animals
  • DNA/genetics
  • Electroporation/instrumentation*
  • Embryo, Nonmammalian/metabolism
  • Equipment Design
  • Fluorescein/metabolism
  • Fluorescent Dyes/metabolism
  • Gene Expression Regulation, Developmental
  • Genetic Vectors
  • Glass/chemistry
  • Green Fluorescent Proteins/genetics
  • Microelectrodes
  • Microfluidics/instrumentation
  • Microtechnology
  • Molecular Weight
  • Plasmids
  • RNA, Messenger/metabolism
  • Substrate Specificity
  • Xanthenes/chemistry
  • Xanthenes/metabolism
  • Zebrafish/embryology*
  • Zebrafish/genetics
  • Zebrafish/metabolism
PubMed: 19130242 Full text @ Biomed. Microdevices.
ABSTRACT
We demonstrate a method which uses simple microfabrication and microfluidics to produce custom, shaped electroporators for the patterned delivery of foreign molecules into developing embryos. We show how these electroporators can be used to 'draw' two-dimensional patterns of tracer molecules, DNA and mRNA into the yolk and cells of zebrafish embryos (Danio rerio) at different stages of development. We demonstrate the successful delivery of patterns of Trypan Blue (normal dye), Texas Red (fluorescent dye), GFP-expressing DNA plasmids and GFP expressing mRNA constructs into both chorionated and dechorionated embryos. Both DNA and mRNA were expressed in the desired patterns subsequent to delivery. Square pulses of 10-20 V (0.20-0.40 kV/cm), 50-100 ms width were sufficient to create transient pores and introduce compounds from the late blastula period (3 hpf) to early pharyngula period (24 hpf) embryos. Using 24 hpf dechorionated embryos, we achieved a high survival of 91.3% and 89%, and a delivery efficiency of 38% and 50% for GFP-DNA and GFP-mRNA respectively. Lastly, we demonstrate the simultaneous delivery of different compounds into the developing embryo.
ADDITIONAL INFORMATION No data available