Germ-line Transformation of the South American Malaria Vector, Anopheles Albimanus, With a piggyBac/EGFP Transposon Vector Is Routine and Highly Efficient

Insect Mol Biol. 2002 Aug;11(4):291-7. doi: 10.1046/j.1365-2583.2002.00336.x.

Abstract

Stable and efficient germ-line transformation was achieved in the South American malaria vector, Anopheles albimanus, using a piggyBac vector marked with an enhanced green fluorescent protein gene regulated by the Drosophila melanogaster polyubiquitin promoter. Transgenic mosquitoes were identified from four independent experiments at frequencies ranging from 20 to 43% per fertile G0. Fluorescence was observable throughout the body of larvae and pupae, and abdominal segments of adults. Transgenic lines analysed by Southern hybridization had one to six germ-line integrations, with most lines having three or more integrations. Hybridized transposon vector fragments and insertion site sequences were consistent with precise piggyBac-mediated integrations, although this was not verified for all lines. The piggyBac/PUbnlsEGFP vector appears to be a robust transformation system for this anopheline species, in contrast to the use of a piggyBac vector in An. gambiae. Further tests are needed to determine if differences in anopheline transformation efficiency are due to the marker systems or to organismal or cellular factors specific to the species.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Anopheles / genetics*
  • Anopheles / metabolism
  • Base Sequence
  • Blotting, Southern / veterinary
  • DNA Transposable Elements
  • Female
  • Genetic Vectors / genetics*
  • Genetic Vectors / metabolism
  • Green Fluorescent Proteins
  • Insect Vectors / genetics*
  • Luminescent Proteins / analysis
  • Luminescent Proteins / biosynthesis
  • Luminescent Proteins / genetics
  • Male
  • Microscopy, Fluorescence / veterinary
  • Molecular Sequence Data
  • Organisms, Genetically Modified / genetics
  • Organisms, Genetically Modified / metabolism
  • Polymerase Chain Reaction / veterinary
  • Sequence Analysis, DNA
  • Transformation, Genetic / genetics*

Substances

  • DNA Transposable Elements
  • Luminescent Proteins
  • Green Fluorescent Proteins