Construction of a Full-Length cDNA Library of Gossypium hirsutum L. and Identification of Two MADS-Box Genes

A full-length normalized cDNA library for the flower development stages of short-season cotton (Gossypium hirsutum L.) (CCRI36) was constructed. A total of 3 421 clones were randomly selected for sequencing, with a total of 3 175 effective sequences obtained after removal of empty-carriers and low-quality sequences. Clustering the 3 175 high-quality expressed sequence tags (ESTs) resulted in a set of 2 906 non-redundant sequences comprised of 233 contigs and 2 673 singletons. Comparative analyses indicated that 913 (43.6%) of the unigenes had homologues with function-known genes or function-assumed genes in the National Center for Biotechnology Information. In addition, 763 (36.4%) of the unigenes were functionally classified using Gene Ontology hierarchy. Through EST alignment and the screening method, the full-length cDNA of two MADS-box genes viz., GhMADS11 and GhMADS12 were acquired. These genes may play a role in flower development. Phylogenetic analysis indicated that GhMADS11 and GhMADS12 had high homology and close evolutionary relationship with AGL2/SEP-type and Pi-type genes, respectively. The expression of both GhMADS11 and GhMADS12, genes was high in reproductive organs. In floral organs, GhMADS11 expression was high in petals (whorl2) and ovules, while GhMADS12 expression was high in petals (whorl2) and stamens (whorl3). Results show that the EST strategy based on a normalized cDNA library is an effective method for gene identification. The study provides more insights for future molecular research on the regulation mechanism of cotton flower development.