Detailed phenotypic and genetic characterization has been done to select the superior Labeo calbasu stock for ex-situ conservation. Landmark-based morphometrics and Microsatellite DNA marker analysis were applied to evaluate the population status of L. calbasu collected from four wild (the Jamuna, the Padma, the Halda and the Haor) and one hatchery sources. Plotting Discriminant functions (DFs) revealed high isolation of the stocks. The average number of allele (5.6) and observed average heterozygosity (Ho) (0.770) were the highest in the Jamuna population. A simple protocol of refrigerated storage and cryopreservation of L. calbasu sperm was developed in this study. Sperm of L. calbasu diluted in Alsever?s solution at 1:10 dilution ratio stored in refrigerator (4oC) produced desirable results and could be cryopreserved using Alsever?s solution with 10% DMSO (5-10 min equilibration) and 10% methanol (20-25 min equilibration) with a cooling rate of 10oC/min from 5 to-80oC. When fertilized with cryopreserved sperm, 1: (4.13?10 5) egg to sperm ratio yielded better results. Cryopreserved fish sperm can facilitate hatchery operations and for long-term conservation of genetic resources.