Improved genetic manipulation of human embryonic stem cells.
Low efficiency of transfection limits the ability to genetically manipulate human embryonic stem cells (hESCs), and differences in cell derivation and culture methods require optimization of transfection protocols. We transiently transferred multiple independent hESC lines with different growth requirements to standardized feeder-free culture, and optimized conditions for clonal growth and efficient gene transfer without loss of pluripotency. Stably transfected lines retained differentiation potential, and most lines displayed normal karyotypes.
Braam
Stefan R
SR
Hubrecht Institute, Developmental Biology and Stem Cell Research, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands.
Denning
Chris
C
van den Brink
Stieneke
S
Kats
Peter
P
Hochstenbach
Ron
R
Passier
Robert
R
Mummery
Christine L
CL
D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002460:Cell Line; D003094:Collagen; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004338:Drug Combinations; D053595:Embryonic Stem Cells / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D020868:Gene Silencing; D018014:Gene Transfer Techniques; D024002:HMGB Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007797:Laminin; D011509:Proteoglycans; D034741:RNA, Small Interfering / Q000235:genetics; D015203:Reproducibility of Results; D055748:SOXB1 Transcription Factors; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014357:Trypsin
D004268:DNA-Binding Proteins; D004338:Drug Combinations; D024002:HMGB Proteins; D007797:Laminin; D011509:Proteoglycans; D034741:RNA, Small Interfering; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; D014157:Transcription Factors; C059485:matrigel; D003094:Collagen; D014357:Trypsin