Introduction: The banana is the world’s most popular fruit crop. A complex of fungal pathogen is responsible for crown rot diseases of banana. Aims: The present study was designed to detect and characterize the crown rot disease of post-harvest banana (Musa paradisiaca) and also develop an alternative quality improvement approach to improve banana shelf-life during storage period. Study Design: A simplest general factorial experiment that was designed to control crown rot disease of banana using different biological factors, including plant extract, antagonistic agents and commercial fungicide. Place and Duration of Study: Disease infected bananas were collected from Rajshahi city, Rajshahi, Bangladesh in 2017 and the experiment had been conducted from April 2017 to April 2018. Methodology: Different morphological, biochemical and molecular techniques were used to characterize and detect the liable fungi. Responsible fungi were subjected to antifungal activity screening test and in vitro antagonism test. Effect of carbendazim and kanamycin B against the mycelial growth of the isolates was determined by disc diffusion method. Quality parameters including disease incidence and severity, pH, TSS, TTA and AA of the treated banana were also analyzed after application of treatments in the packing stage through standard estimation techniques. Results: Two fungi, isolated from the infected portion were further identified as C. musae and L. theobromae. D. metel and A. sativum extract was better in inhibiting mycelial growth of all the test pathogen in culture. B. cereus and T. harzianum moved and attached to fungal isolates, affecting mycelial growth and A. sativum extract significantly affecting conidial germination on artificial medium. Satisfactory mycelia inhibitory effect was recorded from kanamycin B. Quality analysis after storage of banana showed minor measurable differences among treatments. Conclusion: Post-harvest application of A. sativum extract (Conc. 25% w/v) improve the overall quality of harvested banana fruits and reduced the disease incidence and severity of crown rot to a level significantly lower than in fungicide treated or control fruits.