Additional Materials of the Thesis "Decoding the Notch signal: investigating dynamics of transcription directed by Notch responsive enhancers". Julia Falo Sanjuan, January 2020

Movies and plasmid sequences described in the Thesis "Activity and specificity of Notch regulated enhancers". Julia Falo Sanjuan, January 2020 Time-lapse confocal microscopy of Drosophila embryos in which transcription from Notch responsive was imaged live. Movie 01 - Expression of m5/m8. Movie showing transcription from m5/m8-peve in the mesectoderm stripe during nc14. Also note earlier, Notch independent transcription in broad domains in nc10-13 and in few scattered cells during the first few minutes of nc14, followed by a long period (approximately 20 min) of inactivity before MSE stripe cells initiate transcription. Maximum intensity projection (19x1\textmu m stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 10s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 02 - Expression of sim. Movie showing transcription from sim-peve in the mesectoderm stripe and some mesodermal cells during nc14. Also note activity in scattered cells before nc14. Maximum intensity projection (29x1um stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 03 - Ectopic expression of m5/m8 with eve2-NICD. Movie showing ectopic transcription from m5/m8-peve in the eve2 domain during nc14. Maximum intensity projection (29x1\textmu m stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 04 - Ectopic expression of sim with eve2-NICD. Movie showing ectopic transcription from sim-peve in the eve2 domain during nc14. Maximum intensity projection (29x1um stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 05 - Regions of ectopic expression of m5/m8 and sim with eve2-NICD. Combined movie of m5/m8-peve (left) and sim-peve (right) showing ectopic transcription in the eve2 stripe. The maximum projection of the MCP-GFP signal is overlaid with tracked nuclei false colored with the maximum intensity pixel in each nucleus. Active nuclei in each of the analyzed regions is marked with a different color: red (mesoderm), purple (mesectoderm), blue (neuroectoderm) and green (dorsal ectoderm). Anterior to the left; embryo imaged from the ventral side. Movie 06 - Transcription of two m5/m8-peve MS2 reporters in the presence of ectopic NICD. Movie showing ectopic transcription from m5/m8-peve x2 in the eve2 domain during nc14. Maximum intensity projection (29x1um stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 07 - Changes in nuclear size and shape during nc14. Movie showing maximum projection of medial slices and orthogonal views of Nup107-GFP. 0.18 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 08 - Expression of m5/m8 starts during cellularization. Movie showing cellularizing membranes using the marker Gap43-mCherry (maximum intensity projection of medial slices and orthogonal views, left) and transcription from m5/m8-peve (maximum intensity projection (29x1um stacks) of the MCP-GFP channel, right). 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 09 - Localization of Notch and Delta during cellularization. Movies showing maximum projection of medial slices and orthogonal views of Ni-GFP (left) and Dl-mScarlet (right). 0.36 um/px XY resolution and final time resolution of 60s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 10 - Expression of vnd-peve. Movie showing transcription from vnd-peve in the neuroectoderm. Maximum intensity projection (29x1um stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Movie 11 - Transcription of m5/m8 MS2 in a fog mutant background. Mesoderm invagination is delayed and does not proceed properly, resulting in a folded gastrulation phenotype. Maximum intensity projection (29x1um stacks) of the MCP-GFP (grey in left panel, green in right panel) and His2Av-RFP (blue in right panel) channels. 0.36 um/px XY resolution and final time resolution of 15s/frame. Anterior to the left; embryo imaged from the ventral side. Plasmid sequences - annotated sequences of all generated plasmids: pattB-m5/m8-peve-MS2-LacZ pattB-m5/m8-hsp70-MS2-LacZ pattB-m5/m8-pm5-MS2-LacZ pattB-m5/m8-pm6-MS2-LacZ pattB-m5/m8-pm7-MS2-LacZ pattB-m5/m8-pm8-MS2-LacZ pattB-m5/m8-psimE-MS2-LacZ pattB-sim-peve-MS2-LacZ pattB-sim-psimE-MS2-LacZ pattB-m8NE-peve-MS2-LacZ pattB-m8NE-pm8-MS2-LacZ pattB-vnd-peve-MS2-LacZ pattB-pUbi-MS2-LacZ pattB-m5/m8_insSPS-peve-MS2-LacZ pattB-sim_SPS-peve-MS2-LacZ pattB-m5/m8_Dtwi-peve-MS2-LacZ pattB-m5/m8_Ddl-peve-MS2-LacZ pattB-m5/m8_DtwiDdl-peve-MS2-LacZ pCR4-E-m5m8 pCR4-NE-sim pCR4-NE-m8NE' pCR4-E-m5m8_insSPS pCR4-NE-sim_SPS pCR4-E-m5m8_Ddl pCR4-E-m5m8_Dtwi pCR4-E-m5m8_DtwiDdl pattB-eve2x2-peve-FRT-STOP-FRT-NICD-eve3'UTR