Table S1 (Supplementary information).

List of available genetically modified hESC lines

Infection and transfection of human embryonic stem cells

hESC line / Transgene / Technique / Ref.
SNUhES3 / GFP / Transient transfection using Fugene 6 / 1
H1 / GFP / Transient transfection using nucleofection (Amaxa) / 2
H9 / GFP / Transient transfection with Fugene 6 / 3
H1 / GFP / Stable transfection with Fugene 6
(neomycine selection) / 4
H9 / GFP
dsRED2 / Stable transfection with Lipofectamine 2000, Exgen 500
(puromycin selection) / 5
SNUhES3
(D) /

Noggin

/ Stable transfection using Fugene 6
(G418 selection) / 1
H9 / Nodal / Stable transfection with Lipofectamin 2000
(puromycine selection) / 6
H9 / Lefty2
Cerberus-Short
(Constitutive inhibitors of Nodal) / Stable transfection with Lipofectamin 2000
(puromycine selection) / 7
H9 / HSV-TK / Stable transfection with Exgen 500
(neomycin selection) / 8
H1, H9 / EBNA1-neoR
GFP
/ Stable transfection of EBNA1 cDNA with Fugene 6 (neomycin selection)
Episomal EBV-based plasmid expression of GFP / 9
H9 / Pdx1
FoxA2 / Stable transfection with Exgen 500
(Neomycin selection) / 10
H9.2 / hcRed (flanked with loxP sites)
GFP / Stable transfection by electroporation
(neomycin selection) / 11
H7 /
GFP Neo / MSCV Retrovirus / 3
H1, H7, H9
(D) / hTERT / pBABE-MLV Retrovirus / 12
H1, H9
(D) / HOXB4 / MIEV Retrovirus / 13
H9
(D) / HPV16E6E7
hTERT / Retrovirus / 14
H9 / -gal / Adenovirus and adeno-associated virus :
Ad5, AAV2, AAV4 and AAV5 / 15
H1 / GFP / Lentivirus / 16
HES-1 / GFP / Lentivirus / 17
H9 / GFP / Lentivirus / 18
hES-1 / GFP-IRES-PuroR / Lentivirus
(comparison of several IRES and promoters) / 19
H1 / GFP / Lentivirus
(Comparison of different pseudotypes) / 20
H1 / GFP / Lentivirus / 21
(D) indicates DNA transfection to differentiated hESC
Knock-down
hES line / Downregulated gene / Method / Ref.
H9, H1 / RNAi against Oct4 / Transient transfection with Lipofectamin 2000 / 22
H9 / RNAi against GFP
RNAi against dsRED / Transient transfection with Oligofectamine / 5
H1,
hES-NCL1 / RNAi against Nanog / Transient transfection with Lipofectamin 2000 / 23
H1 / RNAi against Oct4 / Transient transfection with Oligofectamin / 24
H7, H14 / RNAi against Oct4
RNAi against
2-microglobulin / Stable transfection with Exgen 500
(puromycine selection) / 25
H9 / RNAi against HPRT / Stable transfection with Oligofectamin, Exgen 500
(Puromycine selection) / 5
H1 / RNAi against GFP / Lentivirus / 26
H9 / RNAi against Oct4
RNAi against Nanog / Lentivirus / 27
Knockout (Homologous recombination)
hES line / Gene / Method / Ref.

H13

/

Knockout of HPRT1

/ Transfection with Exgen 500
(Hygromycin selection) /

28

H1.1

/

Knockout of HPRT1

/ Transfection with electroporation
(Neomycin selection) /

29

Reporter cell lines

hES line / Downregulated gene / Method / Ref.

H1.1

/

Oct-4 knock-in reporterOct4+/IRES-GFP-IRES-NeoR

/

Transfection with electroporation(Homologous recombination, selection with Neomycin)

/

29

H9 / Rex1 reporter
Rex1::GFP / Stable transfection with Exgen 500
(Neomycin selection) / 30
H9 / Albumin reporter
ALB::GFP / Stable transfection with Exgen 500
(Neomycin selection) / 31
H9 / Rat Insulin1
and mouse Pdx1 reporter
Ins::GFP
Pdx1::GFP / Stable transfection with Exgen 500
(Neomycin selection) / 10
H1 / Oct4 reporter
Oct4::GFP / Stable transfection with Fugene 6
(Neomycin selection) / 24

References for Table S1

1Kim JH, Do HJ, Choi SJ, Cho HJ, Park KH, Yang HM et al. Efficient gene delivery in differentiated human embryonic stem cells. Exp Mol Med 2005; 37: 36-44.

2Lakshmipathy U, Pelacho B, Sudo K, Linehan JL, Coucouvanis E, Kaufman DS et al. Efficient transfection of embryonic and adult stem cells. Stem Cells 2004; 22: 531-543.

3Lebkowski JS, Gold J, Xu C, Funk W, Chiu CP, Carpenter MK. Human embryonic stem cells: culture, differentiation, and genetic modification for regenerative medicine applications. Cancer J 2001; 7 Suppl 2: S83-93.

4Liu YP, Dovzhenko OV, Garthwaite MA, Dambaeva SV, Durning M, Pollastrini LM et al. Maintenance of pluripotency in human embryonic stem cells stably over-expressing enhanced green fluorescent protein. Stem Cells Dev 2004; 13: 636-645.

5Vallier L, Rugg-Gunn PJ, Bouhon IA, Andersson FK, Sadler AJ, Pedersen RA. Enhancing and diminishing gene function in human embryonic stem cells. Stem Cells 2004; 22: 2-11.

6Vallier L, Reynolds D, Pedersen RA. Nodal inhibits differentiation of human embryonic stem cells along the neuroectodermal default pathway. Dev Biol 2004; 275: 403-421.

7Vallier L, Alexander M, Pedersen RA. Activin/Nodal and FGF pathways cooperate to maintain pluripotency of human embryonic stem cells. J Cell Sci 2005; 118: 4495-4509.

8Schuldiner M, Itskovitz-Eldor J, Benvenisty N. Selective ablation of human embryonic stem cells expressing a "suicide" gene. Stem Cells 2003; 21: 257-265.

9Ren C, Zhao M, Yang X, Li D, Jiang X, Wang L et al. Establishment and Applications of Epstein-Barr Virus-based Episomal Vectors in Human Embryonic Stem Cells. Stem Cells 2006.

10Lavon N, Yanuka O, Benvenisty N. The effect of over expression of Pdx1 and Foxa2 on the differentiation of human embryonic stem cells into pancreatic cells. Stem Cells 2006.

11Nolden L, Edenhofer F, Haupt S, Koch P, Wunderlich FT, Siemen H et al. Site-specific recombination in human embryonic stem cells induced by cell-permeant Cre recombinase. Nat Methods 2006; 3: 461-467.

12Xu C, Jiang J, Sottile V, McWhir J, Lebkowski J, Carpenter MK. Immortalized fibroblast-like cells derived from human embryonic stem cells support undifferentiated cell growth. Stem Cells 2004; 22: 972-980.

13Wang L, Menendez P, Shojaei F, Li L, Mazurier F, Dick JE et al. Generation of hematopoietic repopulating cells from human embryonic stem cells independent of ectopic HOXB4 expression. J Exp Med 2005; 201: 1603-1614.

14Iuchi S, Dabelsteen S, Easley K, Rheinwald JG, Green H. Immortalized keratinocyte lines derived from human embryonic stem cells. Proc Natl Acad Sci U S A 2006; 103: 1792-1797.

15Smith-Arica JR, Thomson AJ, Ansell R, Chiorini J, Davidson B, McWhir J. Infection efficiency of human and mouse embryonic stem cells using adenoviral and adeno-associated viral vectors. Cloning Stem Cells 2003; 5: 51-62.

16Xue T, Cho HC, Akar FG, Tsang SY, Jones SP, Marban E et al. Functional integration of electrically active cardiac derivatives from genetically engineered human embryonic stem cells with quiescent recipient ventricular cardiomyocytes: insights into the development of cell-based pacemakers. Circulation 2005; 111: 11-20.

17Gropp M, Itsykson P, Singer O, Ben-Hur T, Reinhartz E, Galun E et al. Stable genetic modification of human embryonic stem cells by lentiviral vectors. Mol Ther 2003; 7: 281-287.

18Ma Y, Ramezani A, Lewis R, Hawley RG, Thomson JA. High-level sustained transgene expression in human embryonic stem cells using lentiviral vectors. Stem Cells 2003; 21: 111-117.

19Ben-Dor I, Itsykson P, Goldenberg D, Galun E, Reubinoff BE. Lentiviral Vectors Harboring a Dual-Gene System Allow High and Homogeneous Transgene Expression in Selected Polyclonal Human Embryonic Stem Cells. Mol Ther 2006.

20Jang JE, Shaw K, Yu XJ, Petersen D, Pepper K, Lutzko C et al. Specific and stable gene transfer to human embryonic stem cells using pseudotyped lentiviral vectors. Stem Cells Dev 2006; 15: 109-117.

21Pfeifer A, Ikawa M, Dayn Y, Verma IM. Transgenesis by lentiviral vectors: lack of gene silencing in mammalian embryonic stem cells and preimplantation embryos. Proc Natl Acad Sci U S A 2002; 99: 2140-2145.

22Hay DC, Sutherland L, Clark J, Burdon T. Oct-4 knockdown induces similar patterns of endoderm and trophoblast differentiation markers in human and mouse embryonic stem cells. Stem Cells 2004; 22: 225-235.

23Hyslop L, Stojkovic M, Armstrong L, Walter T, Stojkovic P, Przyborski S et al. Downregulation of NANOG induces differentiation of human embryonic stem cells to extraembryonic lineages. Stem Cells 2005; 23: 1035-1043.

24Gerrard L, Zhao D, Clark AJ, Cui W. Stably transfected human embryonic stem cell clones express OCT4-specific green fluorescent protein and maintain self-renewal and pluripotency. Stem Cells 2005; 23: 124-133.

25Matin MM, Walsh JR, Gokhale PJ, Draper JS, Bahrami AR, Morton I et al. Specific knockdown of Oct4 and beta2-microglobulin expression by RNA interference in human embryonic stem cells and embryonic carcinoma cells. Stem Cells 2004; 22: 659-668.

26Xiong C, Tang DQ, Xie CQ, Zhang L, Xu KF, Thompson WE et al. Genetic engineering of human embryonic stem cells with lentiviral vectors. Stem Cells Dev 2005; 14: 367-377.

27Zaehres H, Lensch MW, Daheron L, Stewart SA, Itskovitz-Eldor J, Daley GQ. High-efficiency RNA interference in human embryonic stem cells. Stem Cells 2005; 23: 299-305.

28Urbach A, Schuldiner M, Benvenisty N. Modeling for Lesch-Nyhan disease by gene targeting in human embryonic stem cells. Stem Cells 2004; 22: 635-641.

29Zwaka TP, Thomson JA. Homologous recombination in human embryonic stem cells. Nat Biotechnol 2003; 21: 319-321.

30Eiges R, Schuldiner M, Drukker M, Yanuka O, Itskovitz-Eldor J, Benvenisty N. Establishment of human embryonic stem cell-transfected clones carrying a marker for undifferentiated cells. Curr Biol 2001; 11: 514-518.

31Lavon N, Yanuka O, Benvenisty N. Differentiation and isolation of hepatic-like cells from human embryonic stem cells. Differentiation 2004; 72: 230-238.