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Cenchrus ciliaris L. (Poaceae) commonly known as buffelgrass is an important perennial grass that grows successfully in arid and semi-arid areas, particularly in varying habitats within the Cholistan Desert of Pakistan. The present study was carried out to assess the effect of limited availability of water on morpho-anatomical features of roots of some diverse ecotypes of C. ciliaris collected from different habitats within the Cholistan Desert and to investigate the relationship between soil physico-chemical and morpho-anatomical characteristics. The highly drought tolerant ecotypes maintained their root and shoot growth even under severe drought stress. Root cross sectional area, size of dermal tissue (epidermis and endodermis), vascular tissue and parenchymatous tissues increased significantly under stressful environments. The drought tolerant ecotypes showed an increased root number and the development of epidermis, endodermis and cortical parenchyma, whereas the drought sensitive ecotypes showed a considerable decrease in almost all morphological and anatomical characteristics. The drought tolerant ecotypes were suitable for revegetation of drought-affected areas of the arid and semi-arid regions.