It is felt that selection needs to be initiated as early as possible in a generation where the constitution would have reached a satisfactory level of uniformity (homozygosity) which minimizes the labour, land and other resource requirement for the maintenance unproductive materials. Keeping this in a mind, Line X tester analysis was carried out involving thirty seven F4 lines and three testers in cotton for assessing the combining ability with respect to twelve yield and yield attributing traits. The objective of this research was to determine the combining ability among segregant F4 lines of within diverse populations. Individual plants in the F4 generation are selfed and crossed with a tester to initiate the selection of improved inbreds. Many of the characters had the preponderance of variances due to specific combining ability indicating the non-additive gene action for these characters. among the 37 lines, 15 lines considered as good parents could be used in further breeding programme. twenty eight crosses had good specific combining ability for seed cotton yield, these crosses were derived from high x low and high x medium general combining ability of lines and would be used for further selection to obtain high yielding lines in the segregating generation.