Identification of genes involved in breast cancer and breast cancer stem cells

Figures & data

Table 1 Primer pairs used in qPCR analysis

Gene	Primer	Sequence (5′–3′)	Amplicon length
18S rRNA	Forward	TGCCCTATCAACTTTCGATGGTAGTC	112 bp
	Reverse	TTGGATGTGGTAGCCGTTTCTCA
NANOG	Forward	TGAGATGCCTCACACGGAGACTG	138 bp
	Reverse	GGGTTGTTTGCCTTTGGGACTG
OCT3/4	Forward	GGTGCCTGCCCTTCTAGGAATG	97 bp
	Reverse	TGCCCCCACCCTTTGTGTTC
SOX2	Forward	CAACGGCAGCTACAGCATGATG	91 bp
	Reverse	GCGAGCTGGTCATGGAGTTGTACT
CD34	Forward	CCCATGCTGGAGGTGACATCTC	130 bp
	Reverse	CCAGGGAGCCGAATGTGTAAAG
HP	Forward	CTGTCATTGCCCTCCTGCTCTG	131 bp
	Reverse	GCGAACCGAGTGCTCCACATAG
SMPD1	Forward	CTCTGTCTGACTCTCGGGTTCTCTG	168 bp
	Reverse	CGAGGTTGATGGCGGTGAATAG
SCRIB	Forward	TGCTGACGACCTGCGGAAGA	158 bp
	Reverse	GTGGCTGCTCATCCTCCTGTTC
KCMF1	Forward	TGGGAGGGCAGTTTTTGGAATG	191 bp
	Reverse	TGAAGAAACGGAGATACACGGACAG
FAM155B	Forward	TGGAGATACGGAAGGGGGAGAG	114 bp
	Reverse	ACCTCAACACTTATCGCCCATCTC
PTGER3	Forward	GCTGGTCTCCGCTCCTGATAATG	91 bp
	Reverse	TCTTTCTGCTTCTCCGTGTGTGTC
GPR3	Forward	GATGCCCACTCTCCACCTCTCTAC	144 bp
	Reverse	GAAGAGGAACAGCAGCAGCAGAC
TMX2	Forward	CATCACCCCTCACCAAGCAACTC	118 bp
	Reverse	TCCTCAGAGAAGGTCCATGAGACAG
DDX49	Forward	GCCAAGTTCAAGTCCAGCATCTACC	87 bp
	Reverse	GACCACCTGTACCGTAGGGATGTC

Abbreviations: qPCR, quantitative polymerase chain reaction; HP, haptoglobin; SMPD1, sphingomyelin phosphodiesterase 1, acid lysosomal; SCRIB, scribbled homolog (Drosophila); KCMF1, potassium channel modulatory factor 1; FAM155B, family with sequence similarity 155, member B; PTGER3, prostaglandin E receptor 3 (subtype EP3); GPR3, G protein-coupled receptor 3; TMX2, thioredoxin-related transmembrane protein 2; DDX49, DEAD (Asp-Glu-Ala-Asp) box polypeptide 49; 18SrRNA, 18S ribosomal RNA; NANOG, Homeobox protein NANOG; OCT3/4, Octamer-binding transcription factor ¾; SOX2, sex determining region Y-box 2; CD34, Hematopoietic progenitor cell antigen CD34.

Figure 1 Microarray clustering analysis.

Notes: The color represents the expression level of the gene. Red represents high expression, while green represents low expression. The expression levels are continuously mapped on the color scale provided at the right of the figure. On the left the experiments are presented as heat maps with the name of samples on the top, and the names of probes on the right.
Abbreviations: ESCs, embryonic stem cells; CSCs, cancer stem cells.

Figure 2 Gene expression among different cancer types.

Notes: Relative gene expression analysis of nine genes that were overexpressed in breast cancer and breast CSCs. The gene expression also was studied in colon and lung cancer, colon CSCs, lung CSCs, embryonic stem cells, and in a reference sample. The experiments were performed in triplicate and a P-value of <0.05 was considered significant.
Abbreviations: CSCs, cancer stem cells; ESCs, embryonic stem cells; HP, haptoglobin; SMPD1, sphingomyelin phosphodiesterase 1, acid lysosomal; SCRIB, scribbled homolog (Drosophila); KCMF1, potassium channel modulatory factor 1; FAM155B, family with sequence similarity 155, member B; PTGER3, prostaglandin E receptor 3 (subtype EP3); GPR3, G protein-coupled receptor 3; TMX2, thioredoxin-related transmembrane protein 2; DDX49, DEAD (Asp-Glu-Ala-Asp) box polypeptide 49.

Figure 3 Breast CSCs pre- and post-siRNA knockdown.

Note: Representative images showing breast CSCs pre- and post-siRNA knockdown.
Abbreviations: CSCs, cancer stem cells; siRNA, small interfering RNA; TMX2, thioredoxin-related transmembrane protein 2; FAM155B, family with sequence similarity 155, member B; PTGER3, prostaglandin E receptor 3 (subtype EP3); GPR3, G protein-coupled receptor 3; DDX49, DEAD (Asp-Glu-Ala-Asp) box polypeptide 49.

Figure 4 Gene expression of stemness transcription factors in breast CSCs.

Notes: Relative gene expression of transcription factors in breast CSCs following knockdown. The ∆∆Ct method was used to perform the analysis. Each bar represents the average of the Ct values. The assays were performed in triplicate and a P-value of <0.05 was considered to be significant. The assays are presented in a log2 scale. Thus, positive values indicate overexpression while negative values indicate underexpression.
Abbreviations: CSCs, cancer stem cells; TMX2, thioredoxin-related transmembrane protein 2; FAM155B, family with sequence similarity 155, member B; PTGER3, prostaglandin E receptor 3 (subtype EP3); GPR3, G protein-coupled receptor 3; DDX49, DEAD (Asp-Glu-Ala-Asp) box polypeptide 49; NANOG, Homeobox protein NANOG; OCT3/4, Octamer-binding transcription factor ¾; SOX2, sex determining region Y-box 2; CD34, Hematopoietic progenitor cell antigen CD34.