A variety-independent protocol was developed for transformation and regeneration of Egyptian cotton plants (Gossypium barbadense L.) from meristemaic tissue of excised embryonic axes and from the shoot apical meristem of seedlings using the Bio-Rad PDS/1000/He gene gun. Tungsten beads coated with plasmid DNA (pBI221) carrying the coding sequence for the GUS protein were used to deliver the gene directly into the meristematic tissue. Transgenic cotton plants have been obtained using techniques of shoot meristem transformation and direct regeneration of shoots and roots. Plants derived from this process were screened for integration of the GUS gene into the plant cell genome using dot blot technique, genomic Southern analysis, immuno blot hybridization and histochemical assay. This work represents the first report for shoot meristems transformation and regeneration of commercial Egyptian cotton varieties, Gossypium barbadense, (Giza 45, Giza 87 and Giza 88 as extra-long staple varieties and Giza 85, Giza 89 and Giza 86 as long staple varieties) using biolistic gun transformation. Molecular and genetic characterization of primary transformants and their progeny proved that GUS gene was successfully integrated in plant cell genome.