Fig. 3. Structure of Xag1 upstream region and reporter constructs. (A) Potential transcription factor-binding sites and the start site of transcription (black arrow) are indicated. Cement gland-specific expression is driven at tailbud stages by –102 bp of upstream sequence. For the 8 kb construct, hatching gland expression is also seen. (B) Linkerscan analysis of this short promoter shows linkerscan (LS) regions 1-3 and 5-6 are most important for this expression. (C) In the context of the larger –275 bp promoter, mutation of these regions has no effect on gfp expression, although the distal promoter alone drives expression only very rarely. A large part of the distal compensation can be attributed to the two EBS found in this region, because when these are mutated the frequency of expression in the cement gland falls significantly. Cement gland expression of gfp in more than 25% of embryos is indicated by +++ (18-25%), ++ (9-17%), + (2-8%), +/– (no change) and – (<2%) (see Table 1; Materials and Methods).