Introducing foreign DNA into tiger shrimp (Penaeus monodon) by electroporation

Theriogenology. 2000 Dec 1;54(9):1421-32. doi: 10.1016/s0093-691x(00)00464-7.

Abstract

Electroporation was used to introduce pFLAG-CMV-1-BAP, a DNA fragment that includes a bacterial alkaline phosphatase gene driven by a human cytomegalovirus (CMV) promoter, into Penaeus monodon zygotes. The transgenic tiger shrimp was achievedby using 10kV, 28 pulses, 120 g sec pulse time, 10 cycles, and a DNA concentration of 37.5 microg/mL. The hatching rate of electroporated zygotes (46%) was significantly lower than that of zygotes in the untreated group (89%). The survival rate of postlarvae in the electroporated group using a DNA concentration of 37.5 microg/mL decreased from 0.6% for postlarva 45 to 0.4% for postlarva 120. Based on dot blot analysis, the rate of gene transfer was 37% in mysis-stage, 23% postlarva 15(PL15), 19% postlarva 45(PL45), and 21% 4-month-old (about PL120). Genomic Southern blotting demonstrated that DNA from transgenic tiger shrimp contained fragments of exogenous DNA that were smaller, larger and of the same molecular size as pFLAG-CMV-1-BAP. Transferred DNA fragments were integrated into the genomes of 31% of the transgenic tiger shrimp. The exogenous DNA was mosaically distributed in a wide variety of tissues. Immunohistochemical staining revealed that the FLAG-BAP fused-protein encoded by pFLAG-CMV-1-BAP was present in the ovaries of some transgenic tiger shrimp.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkaline Phosphatase / genetics
  • Animals
  • Animals, Genetically Modified*
  • Blotting, Southern
  • Cytomegalovirus / genetics
  • DNA, Recombinant
  • Electroporation*
  • Gene Expression
  • Gene Transfer Techniques*
  • Immunoblotting
  • Mosaicism
  • Penaeidae / embryology
  • Penaeidae / genetics*
  • Penaeidae / growth & development
  • Promoter Regions, Genetic
  • Recombinant Fusion Proteins

Substances

  • DNA, Recombinant
  • Recombinant Fusion Proteins
  • Alkaline Phosphatase