Introducing foreign DNA into tiger shrimp (Penaeus monodon) by electroporation.
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.
Tseng
F S
FS
Institutes of Zoology, National Taiwan University, Taipei.
Tsai
H J
HJ
Liao
I C
IC
Song
Y L
YL
D000469:Alkaline Phosphatase / Q000235:genetics; D000818:Animals; D030801:Animals, Genetically Modified; D015139:Blotting, Southern; D003587:Cytomegalovirus / Q000235:genetics; D004274:DNA, Recombinant; D018274:Electroporation; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015151:Immunoblotting; D009030:Mosaicism; D033561:Penaeidae / Q000196:embryology / Q000235:genetics* / Q000254:growth & development; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins
D004274:DNA, Recombinant; D011993:Recombinant Fusion Proteins; D000469:Alkaline Phosphatase