REAGENT or RESOURCE	SOURCE	IDENTIFIER
Antibodies
Goat polyclonal antibody to NANOG	R&D Systems	AF1997
Mouse monoclonal antibody to FLAG	Sigma	F3165
Hoechst 33258	Sigma	94403
Bacterial and virus strains
DH5α Electroporation-Competent Cell	Weidi	DE1001
Chemicals, peptides, and recombinant proteins
5×PrimeSTAR® Buffer	Takara	9158A
dNTPs (10 mM)	Takara	4019
PrimeSTAR-GXL DNA polymerase	Takara	R050A
Ampicillin	Sigma	A9518
SOC Medium	Sigma	S1797
Agarose	Amresco	9002-18-0
Puromycin	Sigma	540411
Dispase	Gibco	17105-041
Trypsin	Gibco	25200072
Doxycycline	MCE	HY-N0565B
1×phosphate buffer (PBS)	Corning	21-040-CV
FBS	ExCell Bio	FSP500
DMEM/F-12	Gibco	11330-032
DMEM (1×) for MEF growth medium	Gibco	11965-092
DMEM (1×) for HEK293FT medium	Corning	10-013-CVRC
Nonessential amino acids (NEAA)	Gibco	11140050
GlutaMAX	Gibco	35050061
KnockOut Serum Replacement	Gibco	A3181502
bFGF	PeproTech	100-18B
Glucose	Biodee	G8270
HEPES	Gibco	11344041
NaCl	SCR	10019318
Na2HPO4	SCR	7558-79-4
CaCl2	Amresco	1631C052
NaOH	SCR	1310-73-2
Glycerol	Amresco	56-81-5
KCl	Sigma	V900068
Critical commercial assays
QIAGEN Plasmid Maxi Kit	QIAGEN	12163
Universal DNA Purification Kit	TIANGEN	DP214-03
QuickExtract DNA Extraction Solution 1.0	Epicentre	QE09050
Deposited data
Raw data	Xu et al., 2021	GEO: GSE132309
Experimental models: cell lines
hPSCs (H9 hESCs)	WiCell	CVCL_9773
HEK293FT cells	Invitrogen	R70007
Oligonucleotides
Primer: sgRNA-PCR-F: AATGG ACTATCATATGCTTACCGTAA CTTGAAAGTATTTCG	This paper	N/A
Primer: sgRNA-PCR-R: CTTTAGT TTGTATGTCTGTTGCTATTATG TCTACTATTCTTTCC	This paper	N/A
Primer: sgRNApool-PCR-F: AATGA TACGGCGACCACCGAGATCTAC ACTCTTTCCCTACACGACGCTCT TCCGATCTTAAGTAGAGGCTTTATAT ATCTTGTGGAAAGGACGAAACACC	This paper	N/A
Primer: sgRNApool-PCR-R: CAAGC AGAAGACGGCATACGAGATTCGC CTTGGTGACTGGAGTTCAGACGTG TGCTCTTCCGATCTCCGACTCGGT GCCCTTTTTCAA	This paper	N/A
Recombinant DNA
CRISPR-Cas9 knockout library	Addgene	1000000049
Cas9-GFP plasmid	Addgene	44719
3×TRE-FLAG-Cas9 plasmid	Addgene	60843
psPAX2 plasmid	Addgene	12260
pMD2.G (VSVG) plasmid	Addgene	12259
AAVS1-TALEN-L plasmid	Addgene	59025
AAVS1-TALEN-R plasmid	Addgene	59026
Software and algorithms
MAGeCK	Li et al., 2014	https://sourceforge.net/p/mageck/wiki/Home/
Cutadapt (version 1.11)	Martin, 2011	https://cutadapt.readthedocs.io/en/stable/
Bowtie2 (version 2.3.4.3)	Langmead and Salzberg, 2012	http://bowtie-bio.sourceforge.net/bowtie2/index.shtml
ImageJ	N/A	https://imagej.nih.gov/ij/
Other
Gene Pulser Xcell system	Bio-Rad	N/A
BD FACSVerse™ System	BD Biosciences	N/A
NanoDrop UV spectrophotometer	Thermo Fisher Scientific	N/A
Illumina HiSeq/NovaSeq system	Illumina	N/A
37°C incubator	Eppendorf	Galaxy 170 R
20-mm Cuvette	Bio-Rad	165-2082
0.22 μm Vacuum driven filter	Corning	431097
0.45 μm Filter	Millipore	HVHP02500
5 mL Round Bottom Polystyrene Test Tube with Cell Strainer Snap Cap	BD	352235
Centrifuge tube	Beckman	344058
Falcon 5 mL Round Bottom Polystyrene Test Tube	Falcon	352054
10 mL Serological pipette	Corning	4488