Production of transgenic dwarf surfclams, Mulinia lateralis, with pantropic retroviral vectors.
A pantropic pseudotyped retroviral vector containing the envelope protein of vesicular stomatitis virus was used as a gene transfer vector in the dwarf surfclam, Mulinia lateralis. These pantropic retroviral vectors have an extremely broad host cell range and can infect many nonmammalian species. Newly fertilized dwarf surfclam eggs were electroporated at 700 V in the presence of 1 x 10(4) colony-forming units of pantropic pseudotyped retroviral particles. Infection was well tolerated and did not affect the survival rate of the embryos. Gametes collected from P1 presumptive transgenic animals were analyzed for the presence of provirus by PCR, and in different experiments 13-33% of the gamete pools were positive for the transgene. Dot blot hybridization of DNA samples from the F1 offspring of two different crosses between infected P1 and wild-type individuals revealed that 28% and 31% of F1 offspring were transgenic, respectively. Southern blot analysis of DNA isolated from PCR-positive F1 animals confirmed integration of a single copy of the provirus into the host genome. Thus, the germ lines of these two P1 transgenic animals were mosaic for the transgene. Expression of beta-galactosidase encoded by the provirus was detected in transgenic but not control surfclam embryos. Pantropic pseudotyped retroviral vectors provide a useful method for the stable introduction of foreign genetic information into surfclams and may facilitate the introduction of desirable genetic traits into commercially important shellfish and crustaceans.
Lu
J K
JK
Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, MD 21202, USA.
Chen
T T
TT
Allen
S K
SK
Matsubara
T
T
Burns
J C
JC
D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D049872:Bivalvia / Q000196:embryology / Q000235:genetics*; D003433:Crosses, Genetic; D017931:DNA Primers / Q000235:genetics; D018274:Electroporation; D005260:Female; D015870:Gene Expression; D005822:Genetic Vectors; D008297:Male; D008969:Molecular Sequence Data; D012190:Retroviridae / Q000235:genetics*; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics; D015053:Zygote; D001616:beta-Galactosidase / Q000235:genetics
D017931:DNA Primers; D001616:beta-Galactosidase