Fig. 6

Platypus testis cell separation and gene expression analysis. a Total testis cell suspensions were applied to a continuous 2-4 % BSA gradient, fractions eluted and cell populations from each fraction isolated for RNA extraction and RT-PCR. F6 has the highest pachytene enrichment (70 %) followed by F4 (42 %) and F9 (2.5 %). Scale bar = μ20 m. b Pachytene or late prophase I fraction content was assessed using surface spreads of each fraction stained for SYCP1. c Equivalent cell numbers were used to extract total RNA from each eluted fraction for semi-quantitative RT-PCR. Pachytene enriched fractions have reduced expression of autosomal and sex linked genes. As pachytene content increases the transcript abundance decreases independent of the genes’ genomic location. Controls were total testis from the same animal used for fractions (Testis, lane 4), platypus male genomic DNA (gDNA, lane 5) and no template (Neg., lane 6). Absence of gDNA control is due to primers spanning large intron. Chromosomal location for each gene in parenthesis left of panels. BSA bovine serum albumin