Green fluorescent protein (GFP) transgenic animals are widely used in biomedical research. We observed that the commonly used β-actin-GFP transgenic mouse has renal defects with proteinuria starting as early as 5 weeks of age. Histological analysis reveals a widespread increase in glomerular extracellular matrix, occasional mesangiolysis, and secondary tubulointerstitial injury. Electron microscopic (EM) analysis reveals dramatic thickening of the glomerular basement membrane (GBM). Several other transgenic strains with GFP on ubiquitous promoters including β-actin (with insertion in a different location) and ubiquitin C show no renal abnormalities. Western blot analysis on crude glomerular preparations from several GFP transgenic strains revealed that higher levels of GFP expression might be responsible for the observed pathogenesis. Mapping of the transgene insertion site by inverse PCR indicates that the β-actin GFP transgene does not cause insertional mutagenesis nor does it modify the transcription level of adjacent genes. Taken together, this strain of β-actin-GFP transgenic mouse may be used to study the mechanism of GBM expansion. Moreover, experiments using this strain of GFP mouse should be hereafter carefully planned because its renal pathology may interfere with data interpretation.