The purpose of this study was to first determine the stage when DNA synthesis occurs in microspores of barley and to subsequently assess the influence of pretreatment on DNA synthesis at different stages of microspore development. Microspores in early, mid, and late uninucleate stages were isolated from the barley cultivars Manley and Igri, and subjected to two different pretreatments: 0.3M mannitol plus cold and cold (4°C) alone. During and after pretreatment, the proportion of microspores at the G1 stage was assessed by measuring relative nucleus DNA levels using densitometry and flow cytometry. There was no correlation between two methods and densitometry was considered the most reliable procedure. Relative to DNA content measurement, microspores were collected at three different morphological stages (youngest, middle and oldest) of cvs. Igri and Manley and them pretreated. This research was undertaken to help determine the optimal time for the bombardment of microspores relative to cell cycle in order to produce homozygous transgenic plants. This information will form the basis for subsequent studies including gene transformation by particle bombardment of regenerable tissues, and in understanding the process of development of microspores leading to embryogenesis. (Abstract shortened by UMI.)