Figure 1

Identification of the spe-8 coding sequence. A. The graph shows all the predicted genes on the far left end of Chromosome I, where spe-8 and mex-3 are known to be very close to one another. The Y axis plots the degree of sperm-specific expression measured as fem-3(q23)/fem-l(hcl7) microarray signal intensity [9, 10]. Two genes in the region, F53Gl2.6 and F53Gl2.8, appear spermatogenesis-specific. F53Gl2.8 had been previously eliminated as a candidate based on other results (Jeremy Nance, personal communication). B. Differential PCR analysis was used to confirn the sperm-specific expression of F53G12.6. PCR products were amplified from cDNA libraries derived either from fem-1(hcl7ts) hermaphrodites which produce only oocytes, or from fem-3(q23ts) hermaphrodites which produce only sperm. The PCR reactions were multiplexed for three possible products: (i) a 169 bp product from F53G12.6 cDNA, (ii) a 526 bp product from F25H8.1, a somatic gene included as a non-germline-specific control, and (iii) a 714 bp product from spe-12, included as a sperm-specific control. Both F53G12.6 and spe-12 products appear sperm specific by their appearance only in the fem-3 library, whereas the product from the somatic gene F25H8.1 was found in both libraries as expected. (PCR primers are given in Additional file 1). C. The results of transformation-rescue of spe-8(hc50) with wild-type PCR product containing F53G12.6 and its flanking sequence. Transformant worms produced a mean of 139.8 progeny (n = 12; SEM = 18.0) compared to a mean of 0.1 progeny (n = 15; SEM = 0.1) produced by their non-transformed siblings, indicating that F53G12.6 is spe-8.