Construction of a cDNA library and identification of genes from Liaoning cashmere goat

Abstract

We constructed a skin cDNA library of the Liaoning cashmere goat during follicle anagen. The recombination was over 89%. A total of 1986 expressed sequence tags (ESTs) were sequenced. After assembly, 1126 ESTs that were homologous to functionally characterized or hypothetical proteins were obtained. BLASTX searches found 904 ESTs that had strong matches to known proteins. More than 50% of the ESTs were similar to proteins that were annotated either as related to enzyme/catalytic activity (26.5%) or as being involved in cell activities, such as growth differentiation, proliferation, apoptosis, transportation, and signal transmission (25.9%). 1.2% of the ESTs were similar to proteins that were related to hair/hair follicle. By analyzing the structure of the unisequences, 81 full-length cDNA clones were identified, none of which had yet been identified in goat. We selected two full-length cDNA clones with predicted proteins, which were similar to dimethyladenosine transferase 1-like (DIMT1L) and coatomer protein complex, subunit zeta (ζ-COP). An analysis of the tertiary structure of the DIMT1L homolog showed that it had a methyltransferase domain. The ζ-COP was a short chain and had a simple tertiary structure. In order to study DIMT1L and ζ-COP genes expression, the semi-quantitative RT-PCR was used to detect whether DIMT1L and ζ-COP genes were expressed in heart, liver, spleen, lung, kidney tissues and in situ hybridization (ISH) to detect DIMT1L and ζ-COP genes expression location. Both DIMT1L and ζ-COP genes had high expression levels in heart and skin, and low levels in liver, kidney, lung and spleen. Moreover, DIMT1L and ζ-COP genes had a strong expression in the inner root sheath of the primary hair follicles (PF) and the secondary hair follicles (SF). The ζ-COP gene also had a strong expression in the surrounding tissue. In conclusion, we have successfully constructed a skin cDNA library from Liaoning cashmere goat during follicle anagen and characterized two genes. The cDNA library could become a valuable source to study the biology genetic characteristics of Liaoning cashmere goat and may provide an important platform for protecting genomic resources and improving the breed.