Microporous polytetrafluoroethylene (PTFE) membrane was widely applied into powder filtering, biomedical and functional breathable apparel areas as an ideal membrane material under harsh environment due to its excellent thermal stability, chemical inertness, low surface energy, high porosity, high strength and so on. But there was few reports about the structure development in PTFE microporous membranes induced by uniaxial stretching operations. In this work, microporous PTFE membranes were prepared from PTFE fine powder by a series of mechanical manipulation, such as extrusion, rolling, and stretching. The morphology of changes of PTFE particles during stretching and the microporous membrane’s unique node-fibril micro-structure were tracked through scanning electron microscopy (SEM) analysis and small angle X-ray scattering (SAXS), which shows that PTFE microporous membranes consisted of island-like node and micro-fibrils domains with the latter highly oriented in the direction of stretching, providing pores in the PTFE membrane as slit-like voids between adjoining fibrils. The crystalline structure and crystallinity of the PTFE were characterized by differential scanning calorimetry (DSC) and wide-angle X-ray diffraction (WAXD), and it was found that the crystallinity of PTFE decreases with the increase of the stretching ratio, which indicates that the stretching operation will damage the PTFE crystal structure. After analyzing and studying the evolution of the morphological structure of PTFE during processing, the relationship between the morphological structure and the relevant properties of the final membrane product was further explored. The effects of stretching processes on the membrane’s structure and pore property were also discussed. The results show that stretching temperature, stretching ratio and stretching rate is the crucial factors affecting the morphology of the membrane, so it can be controlling the morphology by adjusting the above three parameters. This study offers important guidance for the production of PTFE microporous membranes.