A fiberless seed mutant (fl) was identified in a commercial cotton (Gossypium hirsutum L.) variety Xu-Zhou 142 (FL). This phenotype is associated with lack of fiber cell initiation in the outer integument of the ovule, as was characterized by analysis of genes related to fiber differentiation and development. Two genes, fl-E6 and FL-E6, were cloned from fl-integument cells and FL-fiber or integument cells, respectively. Compared with FL-E6, fl-E6 showed a dramatic change in nucleotide sequence: (1). FL-E6 contained a tandem repetitive sequence in which GGCTCA (Gly-Ser) is repeated five times between the 82nd and the 93rd codon from the first ATG codon, while in fl-E6 the same sequence is repeated four times;(2). The fl-E6 gene encodes a polypeptide of 241 amino acids but lacks two codons between the 90th and 93rd codon and three between the 171st and 174th relative to FL-E6;(3). There are also 12 nucleotide substitutions which would result in 7 amino acid differences between fl-E6 and FL-E6. Analysis of RT-PCR and Northern Blot showed that expression of the fl-E6 gene is suppressed in the fl-integument cells, but highly expressed in FL-fiber cells. The difference between fl-E6 and FL-E6 may be associated with lower expression of fl-E6 in the fl-integument cells. Searches of protein databases with the FL-E6 gene sequence showed similarity to the protein backbones of two arabinogalactan-proteins (AGPs), one from the filtrate of suspension-cultured cells of Pyrus communis (AGPPc2) and the other from Nicotiana alata (AGPNa2). Although the function of the FL-E6 protein in differentiation and development of cotton fiber cells is not known, the data indicate that the mutation of fl-E6 gene from FL-E6 gene may inhibit the fiber cell initiation from epidermal cells of the outer integument of the ovule.