Tetraploid plants of distinct genotypes viz., Sugar Baby, California Sweet and an advanced breeding line 5255, generated by the application of colchicine were compared with their diploid counterparts. The experiment was laid out in Randomized complete block design replicated thrice, during 2013-2015 at Punjab Agricultural University, Ludhiana. The comparisons were made on the basis of cytological, palynological and various phenotypic characters to confirm the tetraploidy. The average number of chloroplasts per guard cell pair for tetraploid and diploid plants ranged from 11-12 and 6-8, respectively. Pollen grains showed 4 colpi in tetraploids and 3 colpi in diploids; whereas tetraploids revealed the (29.44%) decrease in pollen viability. Among vegetative parameters, tetraploid plants attained the highest leaf length (14.75 cm) and leaf width (16.36 cm) while internode length had (28.14%) decrease than their corresponding diploids. Of the reproductive parts, staminate flower organs were comparatively of larger size in tetraploids; however the increase in size varied across the different tetraploid genotypes. A delay of 8-9 days in the appearance of first female flower was recorded in tetraploids. Among the various fruit traits, fruit weight and TSS content decreased as (59.64%) and (12.33%), respectively in the tetraploids. TSS content of tetraploids was statistically at par with diploid fruits when harvested on delayed maturity without conceding other quality traits. The fruit rind thickness differed significantly with ploidy levels, however was non-significant within different genotypes. The total number of seeds per fruit and developed seeds in tetraploids were 153.39 and 100.47, respectively. In tetraploids, seed length, seed width and 100 seed weight increased (9.92%, 25.92% and 18.52%), respectively over the diploids. Among the diverse genotypes of watermelon, California Sweet had the highest efficiency of generating putative tetraploids (13.55%).