Transcript Slide 1
Comparison of Gene Expression in H9 and iPS Cells
Authentic Science Research Program
Manchester-Essex Regional High School, Manchester-by-the-Sea, MA 01944
Comparison of Gene Expression in H9 and
Manchester Essex High School, Manchester, MA
Teacher: Dr. Maria Lonnett Burgess, Manchester Essex
Mentor: Dr. George Murphy, Center for Regenerative
Medicine, Boston University, Boston, MA
Induced pluripotent stem cells [iPSCs] are regular somatic
cells that have been “reprogrammed” to dedifferentiate
back into a pluripotent state. The use of induced iPSCs
instead of embryonic H9 stem cells could be advantageous
for a variety of ethical and financial reasons. Many studies
have shown there is identical functionality between iPSCs
and H9 cell, yet researchers are still skeptical of whether
iPSCs and H9 cells are epigenetically identical. This
experiment focuses on the gene expression levels of
known stem cell markers in H9 and iPS cells. We
hypothesize that the iPSCs and the H9 cells should show
similar expression levels of these markers.
iPSC,H9, K562 (hematopoietic cells), and HFF
GAPDH, Nanog, Oct4, Gata1, HBG, Scl, and VWF
• RNA is extracted from the cells.
• cDNA is produced from the RNA with reverse
• The cDNA is run in a polymerase chain reaction
[PCR]. PCR amplifies certain sequences of DNA or
RNA. Through repeated cycles of heating and
cooling, the target sequence is duplicated
• The GAPDH gene (Fig. 5G), a housekeeping gene,
was expressed by all cell types. This means that the
primers were functioning properly.
• The HFF cells expressed only the GAPDH gene,
which means the chances of the samples being
contaminated are unlikely.
• There was more expression of Gata1, VWF, HGB,
and Scl, the hematopoietic markers (Fig. 5C-F) in the
K562 cells than in the H9 or iPS cells.
• There was expression of Nanog and Oct4, the
stem cell markers (Fig. 5A-B) in the iPSC and H9
cells but not in the K562 cells.
• Both the H9 cells and iPSCs showed similar trends
of expression in regards to all genes tested.
• Neither cell type showed the control genes
associated with differentiated cells.
• The results support the hypothesis that iPSCs and
H9 cells are epigenetically identical.
• In future experiments, more markers should be
tested. In addition, experiments should be designed
to compare the expression levels of multiple genes in
one cells type and the expression of one gene across
multiple cell types.
Fig 5. A-G
Stem Cell Markers
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RESEARCH POSTER PRESENTATION DESIGN © 2011
Fig 4. Human iPSC Colony
Fig 3. Mouse iPSC Colonies
•The resulting amplified sequence of cDNA is then
run in real-time polymerase chain reaction [qPCR].
qPCR reactions require a fluorescence marker, Sybr
Green, to measure the level of expression of RNA or
DNA after each individual cycle [CT].
• The original amount of RNA can be computed
based on the rate at which the cDNA proliferated,
and represents a relative expression level the gene.
University Medical Campus Center for Regenerative
Medicine. Boston University, 12 Mar. 2012. Web. 12 Mar.
• Stem cells are defined by their pluripotent and
indefinite self-renewal properties.
• Embryonic stem cells are isolated from an embryo.
This experiment used embryonic cells from the H9
• Induced pluripotent stem cells [iPSCs] are stem
cells that have been dedifferentiated from somatic
vs. embryonic cells.
• Somatic cells are dedifferentiated back into a
pluripotent state by utilizing a viral vector to insert
four key genes (Oct4, Klf4, Sox2, cMyc) into the
1. Amoh Y, Katsuoka K, Hoffman RM. The advantages of
hair follicle pluripotent stem cells over embryonic stem
cells and induced pluripotent stem cells for regenerative
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2. "Center for Regenerative Medicine University Medical
CampusCenter for Regenerative Medicine." Boston
Thank you to George Murphy, the members of his lab Shirley, Brendan,
Amy, and Sarah, and Dr. Maria Lonnett Burgess.
Contact [email protected] for further information.