DIFFERENTIAL DISPLAY TO IDENTIFY AND ISOLATE NOVEL GENES EXPRESSED DURING SPERMATOGENESIS

Spermatogenesis is a complex differentiation process in which diverse stage-specific proteins are co-ordinately expressed. Previously, subtr active hybridization and differential hybridization have been used in the identification of differentially expressed mRNAs, Although these t echniques have been successfully used they require large amounts of RN A and are time consuming. To overcome these problems we have made use of the recently described mRNA differential display technique. The tec hnique is an effective method which can identify and separate cDNAs th at are differentially expressed between various cell-types. By compari ng RNA from testes of mature (>60 days old) and prepubertal (15-16 day s old) mice we have identified nine differential cDNA bands expressed in mature testes. The differential display cDNA band DDC8 was used to screen a testis cDNA library and the full length cDNA was isolated and sequenced. DDC8 cDNA is 1965 bp with an open reading frame of 533 ami no acids which codes far a predicted hydrophilic protein with a calcul ated molecular weight of 62.04 kDa. RNase protection assays indicate D DC8 to be expressed during the postmeiotic stages of spermatogenesis a nd database searches using both nucleotide and amino acid sequences sh ow DDC8 to have similarities to structural, cytoskeletal and associate d proteins.