<title>Abstract</title>Alport syndrome is an inherited chronic human kidney disease, characterized by glomerular basement membrane abnormalities. This disease is caused by mutations in <italic>COL4A3</italic>, <italic>COL4A4</italic>, or <italic>COL4A5</italic> gene. The knockout mice for <italic>Col4</italic>α<italic>3</italic>, <italic>Col4</italic>α<italic>4</italic>, and <italic>Col4</italic>α<italic>5</italic> are developed and well characterized for the study of Alport syndrome. However, disease progression and effects of pharmacological therapy depend on the genetic variability. This model was reliable only to mouse. In this study, we created a novel Alport syndrome rat model utilizing the rGONAD technology, which generated rat with a deletion of the <italic>Col4</italic>α<italic>5</italic> gene. <italic>Col4</italic>α<italic>5</italic> deficient rats showed hematuria, proteinuria, high levels of BUN, Cre, and then died at 18 to 28 weeks of age (Hemizygous mutant males). Histological and ultrastructural analyses displayed the abnormalities including parietal cell hyperplasia, mesangial sclerosis, and interstitial fibrosis. Then, we demonstrated that α3/α4/α5 (IV) and α5/α5/α6 (IV) chains of type IV collagen disrupted in <italic>Col4α5</italic> deficient rats. Thus, <italic>Col4</italic>α<italic>5</italic> mutant rat is a reliable candidate for the Alport syndrome model for underlying the mechanism of kidney diseases and further identifying potential therapeutic targets for human renal diseases.