stock recombinant zebrafish bmp4 protein  (R&D Systems)

Journal: Neural development

Article Title: Capitella teleta gets left out: possible evolutionary shift causes loss of left tissues rather than increased neural tissue from dominant-negative BMPR1.

doi: 10.1186/s13064-024-00181-7

Figure Lengend Snippet: Fig. 2 Nuclear pSMAD1/5/8 levels approximately 10 h after BMPR1ΔK injection. A Boxplot showing the varying effect of BMPR1ΔK injection on the amount of nuclear pSMAD1/5/8 (measured as the relative brightness of labeling with an anti-pSMAD1/5/8 antibody in the nucleus); uninjected BMP-treated animals had significantly more nuclear pSMAD1/5/8 than animals without added BMP, while BMPR1ΔK-injected, BMP-treated animals have significantly less nuclear pSMAD1/5/8 than all treatments except seawater controls. Letters (a, b, c) indicate significance groups; white dots indicate individual cells. B Uninjected embryo in ASW, Arrows: cyan: higher nuclear pSMAD1/5/8; white: lower/no nuclear pSMAD1/5/8. C BMPR1ΔK-injected embryo in ASW. D Uninjected embryo with 1 h BMP4 pulse. E BMPR1ΔK-injected embryo with 1 h BMP4 pulse. Cyan: pSMAD1/5/8; Magenta: Phalloidin. Scale bar: 50 µm

Article Snippet: Stock recombinant zebrafish BMP4 protein (R&D Systems 1128- BM-010) was reconstituted to 20 μg/mL in 0.1% bovine serum albumin (BSA) and 4 mM HCl in Picopure water, aliquoted and stored at -80°C [11].

Techniques: Injection, Labeling

Journal: Neural development

Article Title: Capitella teleta gets left out: possible evolutionary shift causes loss of left tissues rather than increased neural tissue from dominant-negative BMPR1.

doi: 10.1186/s13064-024-00181-7

Figure Lengend Snippet: Fig. 6 BMPR1ΔK-injected phenotypes after a 12h pulse of BMP4 starting at 1q or 4q stage. A Graphical representation of asymmetries present in BMPR1ΔK-injected, BMP pulse animals. Percentages are rounded to the nearest whole number. B-E BMPR1ΔK-injected, ventral view. F-G uninjected, ventral view. B, D, F 1q BMP 12h pulse. C, E, G 4q BMP 12h pulse B BMPR1ΔK-injected + 1q BMP4 animal with elongated foregut (arrow). C BMPR1ΔK-injected + 4q BMP4 animal with merged hemiganglia (arrowhead). D BMPR1ΔK-injected + 1q BMP4 animal with reduced right foregut, mesoderm and trunk muscles. E BMPR1ΔK-injected + 4q BMP4 animal with reduced left mesoderm and trunk muscles. Nuclei: white or cyan; muscles: yellow; stomodeum: asterisk; scale bar: 50 µm

Article Snippet: Stock recombinant zebrafish BMP4 protein (R&D Systems 1128- BM-010) was reconstituted to 20 μg/mL in 0.1% bovine serum albumin (BSA) and 4 mM HCl in Picopure water, aliquoted and stored at -80°C [11].

Techniques: Injection, Muscles

Journal: BMC Biology

Article Title: Highly distinct genetic programs for peripheral nervous system formation in chordates

doi: 10.1186/s12915-022-01355-7

Figure Lengend Snippet: P. mammillata vPNS candidate genes identification by RNA-seq analysis on whole embryos. A Schematic representation of the experimental design and treatments for DGE analysis at late gastrula, initial tailbud II, mid tailbud I and late tailbud II stages. B Quantification of significantly ( p -value <0.05 calculated by DESeq2) differentially expressed genes (in percentage of the total gene number which is 19,467 genes) from DGE analysis. For each RNA-seq experiment, upregulated genes are indicated in green and downregulated genes in red in the histograms. C Differential gene expression of vPNS genes. Log2 fold change of known (yellow backdrop) or candidate (blue backdrop) vPNS genes are shown for samples treated with BMP4 (light blue to dark blue histograms for early to late stages respectively) or BMP4+DAPT (purple histograms for late tailbud stages) compared to control levels. D–L In situ hybridization at selected developmental stages for P. mammillata candidate genes identified from our RNA-seq data and expressed in the vPNS: Hes.a ( D ), Id.b ( E ), Id.c ( F ), Bmp2/4 ( G ), Phmamm.g00001319 ( H ), Nherf.a ( I ), Selectin ( J ), Slc9a3r1 ( K ) and Slc6a12 ( L ). Genes expressed in the VML are represented by a blue circle and genes expressed in ventral ESNs by a purple circle. Embryos are shown in lateral view with dorsal to the top and anterior to the left. Scale bar: 50 μm

Article Snippet: Phallusia mammillata embryos were treated with 150 ng/ml recombinant zebrafish BMP4 protein (1128-BM, R&D Systems Inc, 100 μg/mL stock solution in HCl 4 mM + BSA 0,1%) or recombinant mouse BMP2 protein (355-BM, R&D Systems Inc, 100 μg/mL stock solution in HCl 4 mM + BSA 0,1%) complemented with 0.1% BSA from either the eight-cell stage (St. 4) or initial gastrula stage (St. 10) (both proteins being as potent in our hands), with the BMP receptor inhibitors Dorsomorphin (S7306, Euromedex, 10 mM stock solution in water) and DMH1 (S7146, Euromedex, 10 mM stock solution in DMSO) at 20 and 2.5 μM respectively from eight-cell stage or initial gastrula stage (St. 10), or with 25 μM of the γ-secretase inhibitor DAPT (D5942, Sigma-Aldrich, 10 mM stock solution in DMSO) from early neurula stage (St. 14).

Techniques: RNA Sequencing, Gene Expression, Control, In Situ Hybridization

Journal: BMC Biology

Article Title: Highly distinct genetic programs for peripheral nervous system formation in chordates

doi: 10.1186/s12915-022-01355-7

Figure Lengend Snippet: Branchiostoma lanceolatum vPNS candidate genes identification by RNA-seq analysis on explants. A Schematic representation of animal blastomeres explantation of B. lanceolatum and treatments for DGE analysis. Animal blastomeres are indicated in yellow at the eight-cell stage. B Quantification of significantly ( p -value <0.05 calculated by DESeq2) differentially expressed genes (in percentage of the total gene number which is 27,228 genes) from DGE analysis. For each RNA-seq experiment, upregulated genes are indicated in green and downregulated genes in red in the histograms. C Differential gene expression of vPNS genes. Log2 fold change of known (yellow backdrop) or candidate (blue backdrop) vPNS genes are shown for samples treated with BMP4 (light blue to dark blue histograms for early to late stages respectively) or BMP4+DAPT (pink and purple histograms for mid and late neurula stages respectively) compared to control levels. D–O In situ hybridization at selected developmental stages for B. lanceolatum candidate genes identified from our RNA-seq data and expressed in the vPNS: Wnt11 ( D ), Bmper ( E ), Lbx1/2 ( F ), Gata4/5/6 ( G ), Gata1/2/3 ( H ), Znf-like ( I ), Insm ( K ), Nhlh ( K ), BLAG19000137 ( L ), Myt1 ( M ), Slc6a7/9.a ( N ) and Brsk ( O ). Genes expressed in the ventral neurogenic field are represented by a blue circle and genes expressed in ESNs by a purple circle. Embryos are shown in lateral view with dorsal to the top and anterior to the left. Scale bar: 50 μm

Article Snippet: Phallusia mammillata embryos were treated with 150 ng/ml recombinant zebrafish BMP4 protein (1128-BM, R&D Systems Inc, 100 μg/mL stock solution in HCl 4 mM + BSA 0,1%) or recombinant mouse BMP2 protein (355-BM, R&D Systems Inc, 100 μg/mL stock solution in HCl 4 mM + BSA 0,1%) complemented with 0.1% BSA from either the eight-cell stage (St. 4) or initial gastrula stage (St. 10) (both proteins being as potent in our hands), with the BMP receptor inhibitors Dorsomorphin (S7306, Euromedex, 10 mM stock solution in water) and DMH1 (S7146, Euromedex, 10 mM stock solution in DMSO) at 20 and 2.5 μM respectively from eight-cell stage or initial gastrula stage (St. 10), or with 25 μM of the γ-secretase inhibitor DAPT (D5942, Sigma-Aldrich, 10 mM stock solution in DMSO) from early neurula stage (St. 14).

Techniques: RNA Sequencing, Gene Expression, Control, In Situ Hybridization

Journal: BMC Biology

Article Title: Highly distinct genetic programs for peripheral nervous system formation in chordates

doi: 10.1186/s12915-022-01355-7

Figure Lengend Snippet: New vPNS genes are regulated by BMP and Notch signaling pathways in B. lanceolatum embryos. A–L In situ hybridization for candidate genes expressed in ventral epidermis: Wnt11 at early neurula ( A–C ), HairyE ( D–F ), Gata4/5/6 ( G–I ) and Lbx1/2 ( J–L ) at mid neurula stages in control embryos ( A, D, G, J ), following treatment from late gastrula stage (12 hpf) with BMP4 protein ( B, E, H, K ) or with dorsomorphin ( C, F, I, L ). M–B’ In situ hybridization for candidate genes expressed in ESNs: Nhlh ( M–P ), BLAG19000137 ( Q–T ), Islet ( U–X ) and Brsk ( Y–B’ ) at late neurula stages in control embryos ( M, Q, U, Y ), following treatment from late gastrula stage (12 hpf) with BMP4 protein ( N, R, V, Z ), with DAPT ( O, S, W, A’ ) or a combination of BMP4 and DAPT ( P, T, X, B’ ). Embryos are shown in lateral view with dorsal to the top and anterior to the left. Black arrowheads indicate ESNs when not clearly visible and arrows indicate the anterior most epidermis of the embryo presenting an accumulation of ectopic ESNs. Each experiment has been performed twice. The number of embryos examined for each condition is between 15 and 40, and the pictures shown correspond to the phenotype observed in over 90% of the embryos. Scale bar: 50 μm

Article Snippet: Phallusia mammillata embryos were treated with 150 ng/ml recombinant zebrafish BMP4 protein (1128-BM, R&D Systems Inc, 100 μg/mL stock solution in HCl 4 mM + BSA 0,1%) or recombinant mouse BMP2 protein (355-BM, R&D Systems Inc, 100 μg/mL stock solution in HCl 4 mM + BSA 0,1%) complemented with 0.1% BSA from either the eight-cell stage (St. 4) or initial gastrula stage (St. 10) (both proteins being as potent in our hands), with the BMP receptor inhibitors Dorsomorphin (S7306, Euromedex, 10 mM stock solution in water) and DMH1 (S7146, Euromedex, 10 mM stock solution in DMSO) at 20 and 2.5 μM respectively from eight-cell stage or initial gastrula stage (St. 10), or with 25 μM of the γ-secretase inhibitor DAPT (D5942, Sigma-Aldrich, 10 mM stock solution in DMSO) from early neurula stage (St. 14).

Techniques: Protein-Protein interactions, In Situ Hybridization, Control