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Ectoderm

Q: What are some common usage challenges associated with studying the ectoderm?

One of the key challenges in ectoderm research is identifying the most reproducible and accurate protocols from the vast amount of literature, preprints, and patents. Researchers often struggle to pinpoint the optimal methods that will yield consistent and reliable results for their specific research goals.

Ectoderm is the outermost primary germ layer of an embryo, forming the skin and nervous system.
It plays a crucial role in the development of various structures, including the epidermis, hair, nails, sweat glands, and the central and peripheral nervous systems.
Ectoderm also contributes to the formation of the sensory organs, such as the eyes, ears, and nose.
Understanding the processes involved in ectoderm development is essential for reseach in fields ranging from developmental biology to regenerative medicine.
PubCompare.ai's AI-driven platform can help optimize ectoderm research by identifying the most reproducible and accurate protocols from literature, preprints, and patents, enhancing the efficieny of your ectoderm studies.

Most cited protocols related to «Ectoderm»

Single-Cell Transcriptomics of Hydra

Hydra vulgaris AEP and Hydra vulgaris transgenic lines were dissociated into single cells and were prepared for Drop-seq (49 (link)); FACS was used to enrich for neurons. Sequencing reads were mapped to a de novo assembled transcriptome and a Hydra genome reference and clustering was performed. Subclustering was performed on the following subsets of the data: epithelial ectodermal cells, epithelial endodermal cells, interstitial cells, and neurons and neuronal progenitors. The in situ location of neuron subclusters was determined using in situ hybridization and differential gene expression analysis of separated epithelial layers. URD (24 (link)) was used to build differentiation trajectories for the interstitial and male germline lineages and to analyze the spatial expression of genes in the ectodermal, endodermal, and gland lineages. To analyze regulatory regions, co-expression modules were identified using NMF, ATAC-seq was performed to identify regions of open chromatin, and motif enrichment analysis was used to identify candidate regulators of the gene modules. Colorimetric in situ hybridization, fluorescent in situ hybridization, immunohistochemistry, and generation of transgenic lines was performed and used to validate biomarkers and cell states. For complete methods see supplementary material and methods.

Siebert S., Farrell J.A., Cazet J.F., Abeykoon Y., Primack A.S., Schnitzler C.E, & Juliano C.E. (2019). Stem cell differentiation trajectories in Hydra resolved at single-cell resolution. Science (New York, N.Y.), 365(6451), eaav9314.

Publication 2019

Animals, Transgenic ATAC-Seq Biological Markers Cells Chromatin Colorimetry Ectoderm Endoderm Epithelial Cells Fluorescent in Situ Hybridization Gene Expression Gene Expression Profiling Genes, Regulator Genome Germ Line Hydra Immunohistochemistry In Situ Hybridization Leydig Cells Males Neurons Regulatory Sequences, Nucleic Acid Transcriptome

Quantifying Tissue Deformation Dynamics

We sense the shape of the surface of the embryo in each 3D image stack8 (link), and use these shapes to extract curved image layers of constant radial depth within the embryo. Our methods for quantifying strain rates use distances and velocities calculated across the surface of such curved layers, taking into account the local inclination of the embryo. For Drosophila tissues we extracted layers through the apical zonula adherens of the ectoderm, the site at which much cell-cell interaction is controlled, and for zebrafish tissues we took a surface cutting through the middle of the outermost layer of epiblast cells. Repeating the analysis for different depths of tissues will identify whether the tissue is behaving homogenously in depth, or which layers deform first. The approximate angular curvature spanning an average domain of cells (n_c = 2) are 26.9, 13.3 and 9.0 degrees for our three example tissues respectively. We wrote software to track all cells in these pseudo-2D layers over time (G.B.B. & R.J.A., unpublished), based on the identification of the cell membranes (Supplementary Videos 1-3 and Figure 4a,d,g). The tracking software records for each valid cell, at each time point: the pixelated shape described by the fluorescent cell membrane and hence the cell area; the location of the cell centroid (centre of mass); cell identity; the identity of all touching neighbours. We filter out parts of tracked cell lineages that do not meet criteria for reasonable behaviour, such as anomalously high rates of volume change or cell displacement compared to the immediate cell neighbourhood. This removes 2.5%, 3.9% and 20.9% from our three example tissues respectively. Our analyses are designed to have the essential quality of being robust to occasional missing cells. All cells that touch the edge of the field of view are excluded from the analyses because they may be incomplete. The average number of cells per time point used for strain rate analysis is 87, 522 and 503 respectively.

Blanchard G.B., Kabla A.J., Schultz N.L., Butler L.C., Sanson B., Gorfinkiel N., Mahadevan L, & Adams R.J. (2009). Tissue tectonics: morphogenetic strain rates, cell shape change and intercalation. Nature methods, 6(6), 458-464.

Publication 2009

Cell Communication Cells Drosophila Ectoderm Embryo Epiblast Plasma Membrane Strains Tissues Touch Zebrafish Zonula Adherens

Analyzing hESC Differentiation via DMRs

For analyzing differentiation of hESCs in Fig. 4h, we used a second set of DMRs. We used a pairwise comparison strategy between ESCs and three in vitro derived cell types representative of the three germ layers (mesoderm, endoderm, ectoderm) and performed DMR calling as previously described ⁵². Only DMRs losing more than 30% methylation compared to the ESC state at a significance level of p ≤ 0.01 were retained. Subsequently, we computed weighted methylation levels for all three DMR sets across HUES64, mesoderm, endoderm and ectoderm as well as three consecutive stages of in vitro derived neural progenitors (please see companion⁵² paper for details on the cell types). Finally, we plotted the corresponding distribution using the R function vioplot in the vioplot package. In order to identify potential regulators associated with the loss of DNA methylation at these regions, we determined binding sites of a compendium of transcription factors profiled in distinct cell lines and types (see Ziller, 2013 #45 for details) that overlapped with each set of hypomethylated DMRs. Next, we determined a potential enrichment over a random genomic background by randomly sampling 100 equally sized sets of genomic regions, respecting the chromosomal and size distribution of the different DMR sets and determined their overlap with the same transcription factor binding site compendium to estimate a null distribution. Only transcription factors that showed fewer binding sites across the control regions in 99 of the cases were considered for further analysis. Next, we computed the average enrichment over background for each TF with respect to the 100 sets of random control regions for each germ layer DMR and report this enrichment level in Fig. 4h right, where we capped the relative enrichment at 12.

Kundaje A., Meuleman W., Ernst J., Bilenky M., Yen A., Kheradpour P., Zhang Z., Heravi-Moussavi A., Liu Y., Amin V., Ziller M.J., Whitaker J.W., Schultz M.D., Sandstrom R.S., Eaton M.L., Wu Y.C., Wang J., Ward L.D., Sarkar A., Quon G., Pfenning A., Wang X., Claussnitzer M., Coarfa C., Harris R.A., Shoresh N., Epstein C.B., Gjoneska E., Leung D., Xie W., Hawkins R.D., Lister R., Hong C., Gascard P., Mungall A.J., Moore R., Chuah E., Tam A., Canfield T.K., Hansen R.S., Kaul R., Sabo P.J., Bansal M.S., Carles A., Dixon J.R., Farh K.H., Feizi S., Karlic R., Kim A.R., Kulkarni A., Li D., Lowdon R., Mercer T.R., Neph S.J., Onuchic V., Polak P., Rajagopal N., Ray P., Sallari R.C., Siebenthall K.T., Sinnott-Armstrong N., Stevens M., Thurman R.E., Wu J., Zhang B., Zhou X., Beaudet A.E., Boyer L.A., De Jager P., Farnham P.J., Fisher S.J., Haussler D., Jones S., Li W., Marra M., McManus M.T., Sunyaev S., Thomson J.A., Tlsty T.D., Tsai L.H., Wang W., Waterland R.A., Zhang M., Chadwick L.H., Bernstein B.E., Costello J.F., Ecker J.R., Hirst M., Meissner A., Milosavljevic A., Ren B., Stamatoyannopoulos J.A., Wang T, & Kellis M. (2015). Integrative analysis of 111 reference human epigenomes. Nature, 518(7539), 317-330.

Publication 2015

Binding Sites Cell Lines Cells Chromosomes DNA Methylation Ectoderm Endoderm Enhanced S-Cone Syndrome Genome Germ Cells Germ Layers Human Embryonic Stem Cells Mesoderm Methylation Nervousness Transcription Factor

Xenopus Embryo Tissue Transplantation

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Publication 2009

ARID1A protein, human CCL7 protein, human Cells Ectoderm Embryo Fertilization in Vitro Genes Hair Microscopy Needles paraform Phosphates RNA, Messenger Saline Solution Silicones Tadpole Tissue, Membrane Tissue Donors Tissue Grafts Xenopus laevis

Comprehensive Pan-Cancer DNA Methylation Landscape

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Publication 2018

Cell Body DNA Methylation Ectoderm Embryonic Stem Cells Endoderm Genome Homo sapiens Human Body Induced Pluripotent Stem Cells Mesoderm Neoplasm Metastasis Neoplasms S-pentachlorobuta-1,3-dien-yl-cysteine Stem, Plant Stem Cells Synapses Tissues

Most recents protocols related to «Ectoderm»

Mitotic Spindle Angle Measurement

Spindle angles were determined using FIJI by finding the distance between the mitotic spindle poles: (i) in the xy axis of the flattened ventrolateral ectoderm in a sum projection of the confocal z stack; (ii) in z axis by determining the z planes that the 2 poles appear. The mitotic spindle orientation angle, φ, was calculated by taking the inverse tan of the difference in z depth divided by the distance in xy between the spindle poles (Fig EV3C and D).

Neville K.E., Finegan T.M., Lowe N., Bellomio P.M., Na D, & Bergstralh D.T. (2023). The Drosophila mitotic spindle orientation machinery requires activation, not just localization. EMBO Reports, 24(3), e56074.

Publication 2023

Ectoderm Epistropheus Mitotic Spindle Apparatus Spindle Poles

Confocal Microscopy of Embryo Morphogenesis

Microscopy was performed using either a Leica SP5 point scanning confocal (63×/1.4 HCX PL Apo CS oil lens) or an Andor Dragonfly Spinning Disk confocal microscope (60× water objective). Images were collected with LAS AF or the Andor Fusion software respectively. Minor processing (Gaussian blur) was performed using FIJI. For live imaging, embryos undergoing germband extension were mounted ventro‐laterally between O₂‐permeable membrane (Sartorius) and a glass coverslip in Voltalef oil (Attachem). Embryos were gently squashed such that the ventrolateral ectoderm was flattened.

Publication 2023

Anisoptera Cell Membrane Permeability Ectoderm Embryo Lens, Crystalline Microscopy Microscopy, Confocal

Quantitative Analysis of Cellular Fate Regulators

Pluripotency, PGC regulator, mesoderm, endoderm, and ectoderm gene sets were curated in the literature (Ding et al. 2015 (link); Hackett et al. 2018 ) and in the R&D database (Table S15). First, for each gene in a specific category, such as pluripotency, the log₂FC of the gene was calculated. Then the mean log₂FC of all genes in a category was considered as the overall activity value.

Li D., Yang J., Ma F., Malik V., Zang R., Shi X., Huang X., Zhou H, & Wang J. (2023). The pluripotency factor Tex10 finetunes Wnt signaling for PGC and male germline development. bioRxiv.

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Publication Preprint 2023

Ectoderm Endoderm Genes Mesoderm

Verifying Pluripotency of iPSCs

To proof pluripotency, iPSCs (CMGANTi003-A passage 16 and CMGANTi004-A passage 15) were differentiated into the three embryonic germ layers (mesoderm, endoderm and ectoderm) using the StemMACS Trilineage Differentiation Kit (Miltenyi Biotec) according to manufacturer’s protocol at 37 °C, 5 % CO₂, 20 % O₂. On day seven, cells were collected for RNA extraction and cDNA synthesis. Expression of the selected germ layer markers (Table 1) was verified using RT-qPCR as described above (Table 2).

De Kinderen P., Peeters S., Rabaut L., Mortier G., Ponsaerts P., Loeys B., Verstraeten A, & Meester J.A. (2023). IPSC reprogramming of two patients with spondyloepimetaphyseal dysplasia (SEMD, biglycan type). Stem Cell Research, 67, 103024.

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Publication 2023

Anabolism Cells DNA, Complementary Ectoderm Embryo Endoderm Germ Layers Induced Pluripotent Stem Cells Mesoderm

Characterization and Validation of iPSC Lines

RNA was extracted from both fibroblast and iPSC cell pellets (passage 10) using the Quick-RNA^TM Miniprep Kit (ZYMO Research). Subsequently, cDNA was synthesized using the SuperScript^TM III First-Strand Synthesis System (Life Technologies). Expression of the selected pluripotency markers (Table 1) was confirmed using RT-qPCR TaqMan® probes (Life Technologies) (Table 2) using a BioRad CFX384 Real-Time system (50 °C 2′, 95 °C 10′, 40x (95 °C 15′', 60 °C 1′)).Table 1

Characterization and validation.

Classification	Test	Result	Data
Morphology	Photography Bright field	Normal	Fig. 1 panel F
Phenotype	Qualitative analysis(Immunocytochemistry)	Staining/expression of pluripotency markers: Oct3/4, Nanog, Sox2, Tra1-60, Tra1-80.	Fig. 1 panel A
Phenotype	Quantitative analysis (RT-qPCR)	Expression of DNMT3B, NANOG, POU5F1 and SOX2	Fig. 1 panel B
Genotype	HumanCytoSNP-12 array	Resolution 72 kb, no major copy number variations	Fig. 1 panel D
Identity	HumanCytoSNP-12 arrayOR	> 99.9 % identical SNPs	Table 3
Identity	STR analysis	N/A	N/A
Mutation analysis (IF APPLICABLE)	Sequencing	Hemizygous BGN c.776G > T	Fig. 1 panel C
Mutation analysis (IF APPLICABLE)	Southern Blot OR WGS	N/A	N/A
Microbiology and virology	Mycoplasma	Negative	Supplementary Fig. 2, Supplementary Fig. 3
Differentiation potential	Trilineage differentiation	Expression of appropriate markers of the respective germ layers, i.e. ectoderm, mesoderm and endoderm.	Fig. 1 panel E
List of recommended germ layer markers	Expression of these markers has to be demonstrated at mRNA (RT PCR) or protein (IF) levels, at least 2 markers need to be shown per germ layer	Endoderm: CXCR4, FOXA2, SOX17Mesoderm: NKX2.5, αSMA (ACTA2), HAND1Ectoderm: HES5, MAP2, PAX6	Fig. 1 panel E
Donor screening (OPTIONAL)	HIV 1 + 2 Hepatitis B, Hepatitis C	N/A	N/A
Genotype additional info (OPTIONAL)	Blood group genotyping	N/A	N/A
Genotype additional info (OPTIONAL)	HLA tissue typing	N/A	N/A

Table 2

Reagents details.

	Antibodies used for immunocytochemistry/flow-cytometry
	Antibody	Dilution	Company Cat #	RRID
Pluripotency Markers	Mouse anti-TRA1-60	1:200	Cell Signaling Technology Cat#4746S	AB_2119059
	Rabbit anti-OCT4	1:100	Thermo Fisher Scientific Cat#PA596860	AB_2808662
	Rabbit anti-SOX2	1:500	Merck Millipore Cat#AB5603	AB_2286686
	Mouse anti-TRA1-81	1:200	Cell Signaling Technology Cat#4745S	AB_2119060
	Rabbit anti-NANOG	1:500	ThermoFisher Scientific Cat#PA1-097	AB_2539867
Secondary antibodies	AF555 Goat anti-Mouse, IgM	1:500	Thermo Fisher Scientific Cat#A21426	AB_2535847
Secondary antibodies	AF488 Goat anti-Rabbit, IgG	1:500	Thermo Fisher scientific Cat#A11034	AB_2576217

	Primers
	Target	Size of band	Forward/Reverse primer (5′-3′)

Pluripotency Markers (RT-qPCR)	DNMT3B	55 bp	Hs00171876_m1
	NANOG	99 bp	Hs04260366_g1
	POU5F1	77 bp	Hs04260367_gH
	SOX2	91 bp	Hs01053049_s1
House-Keeping Genes (RT-qPCR)	GAPDH	93 bp	Hs02758991_g1
House-Keeping Genes (RT-qPCR)	ACTB	63 bp	Hs01060665_g1
Differentiation markers (RT-qPCR)	CXCR4	153 bp	Hs00607978_s1
	FOXA2	66 bp	Hs00232764_m1
	SOX17	149 bp	Hs00751752_s1
	NKX2.5	64 bp	Hs00231763_m1
	αSMA (ACTA2)	105 bp	Hs00426835_g1
	HAND1	54 bp	Hs00231848_m1
	HES5	62 bp	Hs01387463_g1
	MAP2	98 bp	Hs00258900_m1
	PAX6	76 bp	Hs00240871_m1
Targeted mutation sequencing	BGN c.776G > T	319 bp	GTTTTCCCAGTCACGACAAGGGTGATGCCAGAGTCC/ CAGGAAACAGCTATGACGACTGAGGGACTGCCCG
Sendai virus Plasmids (PCR)	SeV	181 bp	GGATCACTAGGTGATATCGAGC/ACCAGACAAGAGTTTAAGAGATATGTATC
	KOS	528 bp	ATGCACCGCTACGACGTGAGCGC/ ACCTTGACAATCCTGATGTGGyc
	Klf4	410 bp	TTCCTGCATGCCAGAGGAGCCC/AATGTATCGAAGGTGCTCAA
	c-Myc	532 bp	TAACTGACTAGCAGGCTTGTCG/ TCCACATACAGTCCTGGATGATGATG

Table 3

Cell line identity testing.

iPSC line	total count	correct count	errors	% identical
CMGANTi003-A P10	288,134	288,131	3	>99.9 %
CMGANTi004-A P10	287,779	287,769	10	>99.9 %

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Publication 2023

ACTA2 protein, human Anabolism Antibodies Cells CXCR4 protein, human DNA, Complementary DNMT3B protein, human Ectoderm Endoderm Fibroblasts Genes Germ Layers Goat Hepatitis A Hepatitis B HIV-1 IgG1 IgM1 Immunocytochemistry Induced Pluripotent Stem Cells MAP2 protein, human Mesoderm Mus Mutation Oligonucleotide Primers Pellets, Drug Plasmids POU5F1 protein, human Proteins Rabbits Reverse Transcriptase Polymerase Chain Reaction RNA, Messenger SOX2 protein, human SOX21 protein, human Tissues Virus

Top products related to «Ectoderm»

Stemdiff trilineage differentiation kit by STEMCELL

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The STEMdiff Trilineage Differentiation Kit is a cell culture medium designed for the in vitro differentiation of human pluripotent stem cells into the three primary germ layers: ectoderm, mesoderm, and endoderm. The kit provides the necessary components to support the stepwise differentiation of stem cells towards these lineages.

Human pluripotent stem cell functional identification kit by R&D Systems

Sourced in United States

The Human Pluripotent Stem Cell Functional Identification Kit is a laboratory tool designed to assess the functional characteristics of human pluripotent stem cells. The kit provides essential components and protocols to evaluate the differentiation potential and functionality of these cells.

Dmem f12 by Thermo Fisher Scientific

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DMEM/F12 is a cell culture medium developed by Thermo Fisher Scientific. It is a balanced salt solution that provides nutrients and growth factors essential for the cultivation of a variety of cell types, including adherent and suspension cells. The medium is formulated to support the proliferation and maintenance of cells in vitro.

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Non essential amino acids (neaa) by Thermo Fisher Scientific

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GlutaMAX is a chemically defined, L-glutamine substitute for cell culture media. It is a stable source of L-glutamine that does not degrade over time like L-glutamine. GlutaMAX helps maintain consistent cell growth and performance in cell culture applications.

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Bovine serum albumin (BSA) is a common laboratory reagent derived from bovine blood plasma. It is a protein that serves as a stabilizer and blocking agent in various biochemical and immunological applications. BSA is widely used to maintain the activity and solubility of enzymes, proteins, and other biomolecules in experimental settings.

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Fetal Bovine Serum (FBS) is a cell culture supplement derived from the blood of bovine fetuses. FBS provides a source of proteins, growth factors, and other components that support the growth and maintenance of various cell types in in vitro cell culture applications.

Chir99021 by Selleck Chemicals

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CHIR99021 is a small molecule chemical compound used in laboratory research. It functions as a selective inhibitor of the glycogen synthase kinase-3 (GSK-3) enzyme.

What is the role of the ectoderm in embryonic development?

The ectoderm is the outermost primary germ layer of an embryo and plays a crucial role in the development of various structures, including the epidermis, hair, nails, sweat glands, and the central and peripheral nervous systems. It also contributes to the formation of the sensory organs, such as the eyes, ears, and nose. Understanding the processes involved in ectoderm development is essential for research in fields ranging from developmental biology to regenerative medicine.

What are some common usage challenges associated with studying the ectoderm?

How can PubCompare.ai help with ectoderm research?

PubCompare.ai's AI-driven platform can assist researchers in optimizing their ectoderm studies by helping them screen protocol literature more efficiently and leverage AI to pinpoint critical insights. The platform's advanced comparison tools can identify the most reproducible and accurate methods, enabling researchers to choose the best option for their specific research goals and enhance the efficiency of their ectoderm studies.

Are there different variations or types of ectoderm that researchers should be aware of?

Yes, the ectoderm can be further divided into different regions or cell types, each with its own unique characteristics and developmental pathways. For example, the neural ectoderm gives rise to the central nervous system, while the surface ectoderm forms the epidermis and associated structures like hair and nails. Understanding these variations is important for tailoring ectoderm research to specific applications.

What are some specific applications of ectoderm research?

Ectoderm research has a wide range of applications, from developmental biology to regenerative medicine. Studies on ectoderm development can provide insights into the formation of the skin, nervous system, and sensory organs, which is crucial for understanding birth defects and degenerative diseases. Additionally, ectoderm-derived cells and tissues have potential uses in tissue engineering and regenerative therapies, such as the treatment of skin injuries or neurological disorders.

More about "Ectoderm"

Ectoderm, the outermost primary germ layer of an embryo, plays a crucial role in the development of various structures, including the epidermis, hair, nails, sweat glands, and the central and peripheral nervous systems.
It also contributes to the formation of the sensory organs, such as the eyes, ears, and nose.
Understanding the processes involved in ectoderm development is essential for research in fields ranging from developmental biology to regenerative medicine.
Pluripotent stem cells, derived from sources like human embryos or induced pluripotent stem cells (iPSCs), can be differentiated into ectoderm-derived cell types using specialized culture media and protocols.
The STEMdiff Trilineage Differentiation Kit and the Human Pluripotent Stem Cell Functional Identification Kit are examples of tools that can be used to facilitate ectoderm differentiation.
The culture media used for ectoderm differentiation typically includes DMEM/F12, a nutrient-rich medium that supports cell growth and proliferation.
Matrigel, a basement membrane extract, is often used as a substrate to provide a suitable extracellular matrix for cell attachment and differentiation.
Non-essential amino acids, GlutaMAX, and L-glutamine are also common supplementts that provide essential nutrients for the cells.
Bovine serum albumin (BSA) and fetal bovine serum (FBS) are sometimes added to the culture medium to provide additional growth factors and proteins.
The small molecule inhibitor CHIR99021, which targets the Wnt/β-catenin signaling pathway, has been shown to promote ectoderm differentiation in some protocols.
By leveraging the insights gained from research on ectoderm development and the use of specialized tools and culture conditions, scientists can optimize the efficiency and reproducibility of their ectoderm studies, ultimately advancing our understanding of this critical germ layer and its role in human development and regenerative medicine.