Counting stem cells: methodological constraints.
The number of stem cells contributing to hematopoiesis has been a matter of debate. Many studies use retroviral tagging of stem cells to measure clonal contribution. Here we argue that methodological factors can impact such clonal analyses. Whereas early studies had low resolution, leading to underestimation, recent methods may result in an overestimation of stem-cell counts. We discuss how restriction enzyme choice, PCR bias, high-throughput sequencing depth and tagging method could affect the conclusions of clonal studies.
Laboratory of Stem Cell Research, European Research Institute for the Biology of Ageing, University Medical Center Groningen, University of Groningen, The Netherlands. l.bystrykh@med.umcg.nl
Leonid V
LV
Bystrykh
Evgenia
E
Verovskaya
Erik
E
Zwart
Mathilde
M
Broekhuis
Gerald
G
de Haan
D000818:Animals; D002452:Cell Count / Q000379:methods*; D002999:Clone Cells / Q000166:cytology; D004262:DNA Restriction Enzymes / Q000378:metabolism; D005822:Genetic Vectors; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D051379:Mice; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction; D012190:Retroviridae / Q000235:genetics; D016662:Virus Integration
D004262:DNA Restriction Enzymes