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Abstract  Asymmetric somatic hybrids were obtained between Gossypium hirsutum Coker 201 and wild cotton G. klozschianum Anderss. An investigation on the effect of ultraviolet (UV) irradiation on donor protoplasts was carried out, and the lethal dose was determined to be 38.7 J cm−2. We firstly screened the putative hybrids by the color of the calli produced, followed by morphological, cytological, and molecular analysis of putative hybrid plants. Most regenerated plants derived from fused protoplasts displayed a recipient-like morphology, while some showed an intermediate phenotype between Coker 201 and G. klozschianum. Chromosome numbers in these somatic hybrids ranged from 54 to 74. The hybrids were verified by random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR). Absence or co-existence of parents’ genome DNA fragments was identified through molecular analysis. The heredity of cytoplasm was investigated by cleaved amplified polymorphic sequence (CAPS) analysis using mitochondrial and chloroplast universal primer pairs. The results indicated that recombination and rearrangements might have occurred in some regions of mitochondria (mt) and chloroplast (cp) DNA. To our knowledge, this is the first report about asymmetric protoplast fusion in cotton, and the hybrids obtained would be useful for breeding programs. [ABSTRACT FROM AUTHOR]