The main aim of this study was to transform the enhanced green fluorescent protein gene (egfp) into biocontrol fungus Paecilomyces lilacinus strain 9410. We constructed the expression vector pUPNGT of the fusion gene nptII-egfp using pcDNA3.1(-) as a helper plasmid. The egfp gene was then transformed into P. lilacinus strain 9410 via Agrobacterium tumefaciens-mediated transformation. PCR and Southern blotting analysis showed that the egfp gene was integrated into the genomes of the tested transformants and the integration manner was single-copy. The transformants could generate green fluorescence when they were excited by 488 nm blue laser. These results indicated that the egfp gene had been successfully transformed into P. lilacinus 9410 and expressed in the tested transformants. Our work may provide a new approach to assess environmental safety and practical biocontrol efficacy of P. lilacinus under different conditions.