|
Antibodies
|
| Rabbit anti-AFP |
Dako |
Cat#: A0008; RRID: AB_2650473 |
| Rabbit anti-CC3 |
Cell Signaling Technology |
Cat#: 9661; RRID: AB_2341188 |
| Goat anti-DCX |
Santa Cruz |
Cat#: sc-8066; RRID: AB_2088494 |
| Rabbit anti-DLX1 |
(Lee et al., 2018) |
N/A |
| Rabbit anti-GFAP |
Dako Agilent |
Cat#: Z033429-2 |
| Rabbit anti-GAPDH (14C10) |
Cell Signaling Technology |
Cat#: 2118S; RRID: AB_561053 |
| Rabbit anti-IBA1 |
Wako |
Cat#: 019-19741; RRID: AB_839504 |
| Mouse anti-ISL1 |
DSHB |
Cat#: 40.3A4; RRID: AB_528313 |
| Rabbit anti-Jak2 (D2E12) XP |
Cell Signaling Technology |
Cat#: 3230S; RRID: AB_2128522 |
| Mouse anti-KI67 |
BD |
Cat#: 550609; RRID: AB_393778 |
| Rabbit anti-KLF4 |
Santa Cruz |
Cat#: sc-20691; RRID: AB_669567 |
| Mouse anti-NESTIN |
Stemcell Technologies |
Cat#: 60091; RRID: AB_2650581 |
| Rabbit anti-NKX2.1 |
Santa Cruz |
Cat#: sc-13040; RRID: AB_793532 |
| Mouse anti-NKX2.2 |
DSHB |
Cat# 74.5A5; RRID:AB_531794 |
| Mouse anti-NEUN |
Millipore |
Cat#: MAB377B; RRID: AB_177621 |
| Rabbit anti-NPY |
Millipore |
Cat#: ab9608; RRID: AB_2153720 |
| Mouse anti-OCT4 |
Santa Cruz |
Cat#: sc-5279; RRID: AB_628051 |
| Guinea Pig anti-OTP |
Clontech |
Cat#: M195 |
| Mouse anti-PAX6 |
BD |
Cat#: 561664; RRID: AB_10895587 |
| Rabbit anti-POMC |
Phoenix Pharmaceuticals |
Cat#: H-029-30; RRID: AB_2307442 |
| Rabbit anti-PV |
Abcam |
Cat#: ab11427; RRID: AB_298032 |
| Rabbit anti-Phospho-Jak2 (Tyr1007/1008) (C80C3) |
Cell Signaling Technology |
Cat#: 3776S; RRID: AB_2617123 |
| Rabbit anti-Phospho-Stat-3 (tyr705) |
Cell Signaling Technology |
Cat#: 9131S; RRID: AB_331586 |
| Mouse anti-RAX |
Santa Cruz |
Cat#: sc-271889; RRID: AB_10708730 |
| Rabbit anti-SMA |
Abcam |
Cat#: AB5694; RRID: AB_2223021 |
| Goat anti-SOX2 |
Santa Cruz |
Cat#: sc-17320; RRID: AB_2286684 |
| Mouse anti-SSEA4 |
Millipore |
Cat#: MAB4304; RRID: AB_177629 |
| Rabbit anti-SST |
Santa Cruz |
Cat#: sc-13099 RRID: AB_2195930 |
| Mouse anti-STEM121 |
Takara |
Cat#: Y40410; RRID: AB_2801314 |
| Mouse anti-S100B (CL2720) |
Sigma-Aldrich |
Cat#: AMAb91038; RRID: AB_2665776 |
| Mouse anti-Stat3 (124H6) |
Cell Signaling Technology |
Cat#: 9139S; RRID: AB_331757 |
| Goat anti-Tbx3 |
Santa Cruz |
Cat#: sc-17871; RRID: AB_661666 |
| Mouse anti-TRA-1-60 |
Millipore |
Cat#: MAB4360; RRID: AB_2119183 |
|
Chemicals, Peptides, and Recombinant Proteins
|
| 2-Mercaptoethanol |
Thermo Fisher Scientific |
21985023 |
| A83-01 |
Stemcell Technologies |
72024 |
| Advantage 2 PCR Kit |
Takara Bio |
639206 |
| Advantage UltraPure PCR deoxynucleotide mix (10mM each dNTP) |
Takara Bio |
639125 |
| AMPure XP beads |
Beckman Coulter |
A63880 |
| Ascorbic acid |
Sigma-Aldrich |
A0278 |
| B-27 Supplement (50X), minus vitamin A |
Thermo Fisher Scientific |
12587010 |
| bFGF |
Peprotech |
100-18B |
| Bovine serum albumin (BSA) |
Sigma-Aldrich |
B6917; CAS# 904846-8 |
| Bio-Rad Protein Assay Dye Reagent Concentrate |
Bio-Rad |
500-0006 |
| Collagenase Type IV |
Invitrogen |
17104019 |
| complete™, Mini, EDTA-free Protease Inhibitor Cocktail |
Sigma-Aldrich |
11836170001 |
| DAPI |
Thermo Fisher Scientific |
D1306 |
| Dibutyryl cyclic AMP |
Sigma-Aldrich |
D0627 |
| DL-Dithiothreitol solution |
Millipore Sigma |
43816 |
| Donkey serum |
Millipore |
S30-100ML |
| Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12 (DMEM:F12) |
Thermo Fisher Scientific |
11320033 |
| Dulbecco’s phosphate-buffered saline (DPBS), calcium, magnesium |
Thermo Fisher Scientific |
14040133 |
| Dulbecco’s phosphate-buffered saline (DPBS), no calcium, no magnesium |
Thermo Fisher Scientific |
14190144 |
| Durapore® Membrane Filter, 0.22 μm |
Millipore |
GVWP04700 |
| EZ PREP Nuclei Isolation Kit |
Sigma-Aldrich |
NUC101-1KT |
| EZ-Tn5 Transposase |
Lucigen |
TNP92110 |
| GlutaMAX supplement |
Gibco |
35050061 |
| IWR-1 Endo |
Cayman Chemical |
13659-25 |
| KAPA HiFi Hotstart Readymix |
Emsco/Fisher |
KK2601 |
| KnockOut Serum Replacement |
Thermo Fisher Scientific |
10828028 |
| LDN-193189 |
Stemcell Technologies |
72147 |
| Recombinant Human Leptin |
Peprotech |
300-27 |
| MEM Non-Essential Amino Acids Solution (100X) |
Thermo Fisher Scientific |
11140050 |
| MgCl2 (1M) |
Thermo Fisher Scientific |
AM9530G |
| N-2 Supplement (100X) |
Thermo Fisher Scientific |
17502048 |
| Neurobasal medium |
Thermo Fisher Scientific |
21103049 |
| Nuclease-Free Water (not DEPC-Treated) |
Thermo Fisher Scientific |
AM9937 |
| Formaldehyde, 16%, methanol free, Ultra Pure |
Polysciences |
18814-10 |
| Penicillin-Streptomycin (5,000 U/mL) |
Thermo Fisher Scientific |
15070063 |
| Polyethylene glycol (PEG) solution, 40% |
Sigma-Aldrich |
P1458 |
| Purmorphamine |
Tocris |
455110 |
| PhosSTOP™ |
Sigma-Aldrich |
4906845001 |
| Recombinant Human/Murine/Rat BDNF |
Peprotech |
450-02 |
| Recombinant Human GDNF |
Peprotech |
450-10 |
| Recombinant Human SHH |
BioLegend |
753506 |
| RNA Clean & Concentrator 5 |
Zymo Research |
R1013 |
| RNase Inhibitor, Murine |
New England Biolabs |
M0314S |
| SAG |
Cayman Chemicals |
11914-1 |
| SDS (10% w/v) |
Fisher Scientific |
50-751-7490 |
| SMARTScribe Reverse Transcriptase |
Takara |
639537 |
| Sodium hydroxide solution (1 N) |
Sigma-Aldrich |
1091371000 |
| Sterile saline solution injection |
Midwest Veterinary Supply |
193.74500.3 |
| Sucrose |
Sigma-Aldrich |
S0389 |
| SuperSignal™ West Femto Maximum Sensitivity Substrate |
Thermo Fisher Scientific |
34096 |
| TFM Tissue Freezing Medium |
General Data |
TFM-5 |
| Tris (1 M), pH 8.0, RNase-free |
Thermo Fisher Scientific |
AM9855G |
| Triton X-100 |
Sigma-Aldrich |
T9284 |
| TRIzol reagent |
Thermo Fisher Scientific |
15596026 |
| Trypan blue stain, 0.4% |
Thermo Fisher Scientific |
T10282 |
| TWEEN 20 |
Sigma-Aldrich |
P1379 |
| VECTASHIELD Vibrance Antifade Mounting Medium |
Vector Laboratories |
H170010 |
|
Critical Commercial Assays
|
| BrainPhys™ Neuronal Medium N2-A & SM1 Kit |
STEMCELL Technologies |
05793 |
| CytoView MEA 12 |
Axion Biosystems |
M768-GL1-30Pt200 |
| ELISA Kit for Alpha-Melanocyte Stimulating Hormone (aMSH) |
Cloud-Clone Corp |
CEA239Hu |
| Qubit dsDNA HS Assay Kit |
Thermo Fisher Scientific |
Q33231 |
| KAPA Library Quantification Kit for Illumina NGS |
Kapa Biosystems |
KK4835 |
| Neural Tissue Dissociation Kit |
Miltenyi Biotech |
130-092-628 |
| NextSeq High Output v2 75 Cycles |
Illumina |
TG-160-2005 |
| Nextera XT Library Prep Kit |
Illumina |
FC-131-1024 |
|
Deposited Data
|
| Bulk and single-cell RNA-sequencing of ARCOs |
This paper |
GEO: GSE164102
|
| Single-nucleus RNA-sequencing of human neonatal hypothalamus |
This paper |
GEO: GSE164101
|
|
Experimental models: Cell lines
|
| Human C3-1 iPSC line |
(Wen et al., 2014) |
DOI: 10.1038/nature13716 |
| Human C65 iPSC line |
This study |
|
| PWS Major Deletion line |
This study |
(Kuslich et al., 1999) |
| PWS Minor Deletion line |
This study |
(Schüle et al., 2005) |
| Software and Algorithms |
| Adobe Illustrator CC |
Adobe |
https://www.adobe.com/products/illustrator.html; RRID:SCR_010279 |
| Adobe Photoshop CC |
Adobe |
https://www.adobe.com/products/photoshop.html; RRID:SCR_014199 |
| bcl2fastq v2.17.1.14 |
Illumina |
https://support.illumina.com/sequencing/sequencing_software/bcl2fastq-conversion-software.html; RRID:SCR_015058 |
| DESeq2 v1.24.0 |
(Love et al., 2014) |
https://bioconductor.org/packages/release/bioc/html/DESeq2.html; RRID:SCR_015687 |
| Drop-seq tools v.2.1.0 |
(Saunders et al., 2018) |
https://github.com/broadinstitute/Drop-seq; RRID:SCR_018142 |
| EnhancedVolcano v1.7.8 |
GitHub |
https://github.com/kevinblighe/EnhancedVolcano; RRID:SCR_018931 |
| GenomicFeatures v1.36.4 |
(Lawrence et al., 2013) |
http://bioconductor.org/packages/release/bioc/html/GenomicFeatures.html; RRID:SCR_016960 |
| GraphPad Prism |
GraphPad Software |
https://www.graphpad.com/scientific-software/prism/; RRID:SCR_002798 |
| Harmonizome |
(Rouillard et al., 2016) |
http://amp.pharm.mssm.edu/Harmonizome/; RRID:SCR_016176 |
| Human genome release 28 (GRCh38.p12) |
GENCODE |
https://www.gencodegenes.org/human/release_28.html
|
| Imaris |
Bitplane |
http://www.bitplane.com/imaris; RRID:SCR_007370 |
| Monocle v2.14.0 |
(Qiu et al., 2017a; Qiu et al., 2017b; Trapnell et al., 2014) |
http://cole-trapnell-lab.github.io/monocle-release/docs/; RRID:SCR_016339 |
| PANTHER v15.0 |
(Mi et al., 2019) |
http://www.pantherdb.org/; RRID:SCR_004869 |
| pathwayPCA v1.4.0 |
(Odom et al., 2019) |
https://www.bioconductor.org/packages/release/bioc/html/pathwayPCA.html
|
| R Project v3.6.0 |
Open source |
https://www.r-project.org/; RRID:SCR_001905 |
| RStudio 1.2.1335 |
Open source |
https://www.rstudio.com/; RRID:SCR_000432 |
| scater v1.12.2 |
(McCarthy et al., 2017) |
https://bioconductor.org/packages/release/bioc/html/scater.html; RRID:SCR_015954 |
| Seurat v3.1.5 |
(Stuart et al., 2019) |
https://github.com/satijalab/seurat; RRID:SCR_007322 |
| STAR v2.5.2a |
(Dobin et al., 2013) |
https://github.com/alexdobin/STAR; RRID:SCR_015899 |
| Trimmomatic v0.32 |
(Bolger et al., 2014) |
http://www.usadellab.org/cms/index.php?page=trimmomatic; RRID:SCR_011848 |
| VennDiagram v1.6.20 |
(Chen and Boutros, 2011) |
https://cran.r-project.org/web/packages/VennDiagram/; RRID:SCR_002414 |
| Zen Blue |
Carl Zeiss |
https://www.zeiss.com/microscopy/us/products/microscope-software/zen-lite.html; RRID:SCR_013672 |
|
Experimental models: Organisms/Strains
|
| CD-1 Mus musculus
|
Charles River |
CRL:022; RRID: IMSR_CRL:022 |
| NU/J Mus musculus, female |
The Jackson Laboratory |
Cat# 002019; RRID: IMSR_JAX:002019 |
|
Oligonucleotides
|
| CDS Primer sequence: 5′-AAGCAGTGGTATCAACGCAGAGTACT30VN-3′ |
IDT |
N/A |
| TSO Primer sequence: 5′-AAGCAGTGGTATCAACGCAGAGTACATrGrGrG-3′ |
IDT |
N/A |
| LS PCR Primer sequence: 5′-AAGCAGTGGTATCAACGCAGAGT-3′ |
IDT |
N/A |
|
Others
|
| 23G blunt needle |
Neta Scientific |
B23-100 |
| 6-well ultra-low attachment culture plate |
Fisher Scientific |
07200601 |
| Bioanalyzer 2100 |
Agilent |
G2939BA |
| C&B Metabond Quick adhesive cement system |
Benco Dental |
1681-343 |
| Cell counting slides |
Thermo Fisher Scientific |
C10228 |
| Charged microscope slides |
Fisher Scientific |
22-035813 |
| Countess II Automated Cell Counter |
Thermo Fisher Scientific |
AMQAX1000 |
| Disposable pellet pestle |
Fisher |
12-141-368 |
| Fine Forceps - Curved/Serrated |
Fine Science Tools |
11065-07 |
| Forma Steri-Cult CO2 Incubator |
Thermo Fisher Scientific |
3310 |
| Gelfoam Sponge |
Pfizer |
031508 |
| Hydrophobic barrier PAP pen |
Vector Laboratories |
H-4000 |
| In-VitroCell ES Direct Heat CO2 Incubator |
Nuaire |
NU-5710 |
| MACS SmartStrainer, 70 μM |
Miltenyi Biotech |
130-098-462 |
| Maxi-Cure Super Glue |
Bob Smith Industries |
BSI-113 |
| MaxQ CO2 Plus Shaker |
Thermo Fisher Scientific |
88881102 |
| Microfluidic chips for drop-seq |
FlowJEM |
Custom order |
| Micromotor drill |
Stoelting |
51449 |
| NanoDrop 2000 |
Thermo Fisher Scientific |
ND-2000 |
| NextSeq550 |
Illumina |
SY-415-1002 |
| PCR tube strips |
Emsco/Fisher |
AB0490 |
| Pellet pestle cordless motor |
Fisher |
12-141-361 |
| Plastic cryomold |
Electron Microscopy Sciences |
62534-10 |
| Qubit 3 Fluorimeter |
Thermo Fisher Scientific |
Q33216 |
| Tube rotator |
Boekel |
UX-51202-07 |
| Round Bottom Polystyrene Test Tube, with Cell Strainer Snap Cap |
Corning |
352235 |
| T100 Thermal Cycler |
Bio-rad |
1861096EDU |
| TB Syringe (26 G × 3/8 in, 1 ml) |
BD Biosciences |
309625 |
| Tissue homogenizer |
Sigma |
D8938 |
| Vannas Spring Scissors - Curved/3mm Cutting Edge |
Fine Science Tools |
15000-10 |
