Notochord CRMs controlled by Ci-Bra through two cooperative binding sites.

<p>Identification of the minimal sequences necessary for notochord activity of the <i>Ci-FCol1</i> and <i>Ci-Noto5</i> notochord CRMs, through sequence-unbiased truncation analyses and site-directed mutations. All schematics are as in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001697#pbio-1001697-g002" target="_blank">Figure 2</a>. (A) Sequence-unbiased truncation and site-directed mutation analyses of the <i>Ci-FCol1</i> notochord CRM. (B–F) Low-magnification group microphotographs of embryos electroporated in parallel with either the wild-type 65-bp <i>Ci-FCol1</i> minimal notochord CRM (B) or with mutated versions of this construct carrying individual mutations in each Ci-Bra binding site, as shown at the bottom of each panel (C–E) or a double mutation in the two proximal Ci-Bra binding sites (F). (G) Quantification of the activity of the constructs shown in (B–F) in different tissues, as in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001697#pbio-1001697-g002" target="_blank">Figure 2</a>. (H) Sequence-unbiased truncation and site-directed mutation analyses of the <i>Ci-Noto5</i> notochord CRM. (I–M) Low-magnification group microphotographs of embryos electroporated in parallel with either the wild-type 83-bp <i>Ci-Noto5</i> minimal notochord CRM (I) or with mutated versions of this construct carrying the individual mutations in each Ci-Bra binding site shown at the bottom of each panel (J–L) or a double mutation in the two proximal Ci-Bra binding sites (M). (N) Quantification of the activity of the constructs shown in (I–M) in different tissues, plotted as described in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001697#pbio-1001697-g002" target="_blank">Figure 2</a>. Insets show higher magnifications of representative stained embryos.