Table 1.

Information * on the biological functions of 20 candidate genes and on the primer sequences used for RT-qPCR

Gene Symbol	Gene Name and Functions	Primer Sequence
VANGL1 (KITENIN)	VANGL planar cell polarity protein 1 (located on chromosome 1) 1. It encodes a member of the tetraspanin family. 2. It may be involved in mediating intestinal trefoil factor-induced wound healing in the intestinal mucosa.	F: 5′-GACACAAGTCACCCCGGAATA-3′ R: 5′-TCCTCTGTCCGAGTAGAATCATT-3′ Amplicon length: 109 bp
IL2RA (CD25)	Interleukin 2 receptor subunit alpha (located on chromosome 10) 1. Mutations in this gene are associated with interleukin 2 receptor alpha deficiency. 2. Serum IL-2R levels are found to be elevated in patients with different types of carcinomas.	F: 5′-GAACACAACGAAACAAGTGACAC-3′ R: 5′-GGCTGCATTGGACTTTGCATT-3′ Amplicon length: 81 bp
IL2RB (CD122)	Interleukin 2 receptor subunit beta (located on chromosome 22) 1. It is involved in receptor-mediated endocytosis and transduction of mitogenic signals from IL2.	F: 5′-CAGCGGTGAATGGCACTTC-3′ R: 5′-GGCATGGACTTGGCAGGAA-3′ Amplicon length: 113 bp
TGFβ1	Transforming growth factor ß (located on chromosome 19) 1. It encodes a secreted ligand of the TGFß superfamily of proteins. The encoded protein regulates cell proliferation, differentiation, and growth. 2. It is frequently upregulated in tumor cells.	F: 5′-CAATTCCTGGCGATACCTCAG-3′ R: 5′-GCACAACTCCGGTGACATCAA-3′ Amplicon length: 86 bp
SMAD2	SMAD2 family member 2 (located on chromosome 18) 1. The protein encoded mediates the signal of the transforming growth factor (TGF)-beta, thus regulating multiple cellular processes (i.e., cell proliferation, apoptosis, and differentiation).	F: 5′-CCGACACACCGAGATCCTAAC-3′ R: 5′-GAGGTGGCGTTTCTGGAATATAA-3′ Amplicon length: 125 bp
SMAD4	SMAD4 family member 4 (Located on chromosome 18) 1. The encoded protein forms homomeric complexes and heteromeric complexes with other activated Smad proteins that accumulate in the nucleus and regulate the transcription of target genes. 2. The encoded protein acts as a tumor suppressor and inhibits epithelial cell proliferation.	F: 5′-CCACCAAGTAATCGTGCATCG-3′ R: 5′-TGGTAGCATTAGACTCAGATGGG-3′ Amplicon length: 76 bp
CD44 (CSPG8)	CD44 antigen (located on chromosome 11) 1. It is a cell-surface glycoprotein involved in cell–cell interactions, cell adhesion, and migration. 2. The encoded protein participates in several cellular functions (e.g., lymphocyte activation, recirculation and homing, hematopoiesis, and tumor metastasis).	F: 5′-CTGCCGCTTTGCAGGTGTA-3′ R: 5′-CATTGTGGGCAAGGTGCTATT-3′ Amplicon length: 109 bp
CD133 (PROM1)	CD133 (located on chromosome 4) 1. It encodes a pentaspan transmembrane glycoprotein. 2. The encoded protein is often expressed on adult stem cells, where it has been suggested to maintain stem cell properties by suppressing differentiation. 3. It is the marker most commonly used for the isolation of cancer stem cell population from different tumors.	F: 5′-GGCCCAGTACAACACTACCAA-3′ R: 5′-ATTCCGCCTCCTAGCACTGAA-3′ Amplicon length: 75 bp
HES1	HES family basic helix-loop-helix (bHLH) transcription factor 1 (chromosome 3) 1. It is a transcriptional repressor of genes that require a bHLH protein for their transcription. 2. It plays an important role in the Notch signaling pathway. 3. The absence of Hes1 in the developing intestine promotes the increase of Math1 (the production of intestinal cell types).	F: 5′-ACGTGCGAGGGCGTTAATAC-3′ R: 5′-GGGGTAGGTCATGGCATTGA-3′ Amplicon length: 90 bp
NOTCH1	NOTCH receptor 1 (located on chromosome 9) 1. The Notch signaling pathway regulates interactions between physically adjacent cells through the binding of Notch family receptors to their cognate ligands. 2. It plays a role in the development of numerous cell and tissue types. 3. Mutations in NOTCH1 are associated with syndromes, hematological and solid tumors.	F: 5′-TGGACCAGATTGGGGAGTTC-3′ R: 5′-GCACACTCGTCTGTGTTGAC-3′ Amplicon length: 82 bp
LEF1 (TCF10)	Lymphoid enhancer-binding factor 1 (located on chromosome 4) 1. It encodes a transcription factor belonging to a family of proteins that share homology with the high mobility group protein-1. 2. It binds to a functionally important site in the T-cell receptor-alpha (TCRA) enhancer, thus conferring maximal enhancer activity. 3. It is involved in the Wnt signaling pathway, and mutations in this gene have been found in some tumors.	F: 5′-ATGTCAACTCCAAACAAGGCA-3′ R: 5′-CCCGGAGACAAGGGATAAAAAGT-3′ Amplicon length: 76 bp
MTA1	Metastasis-associated 1 (located on chromosome 14) 1. MTA1 expression has been correlated with the metastatic potential of some carcinomas, but it is expressed also in many normal tissues. 2. The profile and activity of the encoded protein suggest that it is involved in regulating transcription and that this may be accomplished by chromatin remodeling.	F: 5′-ACGCAACCCTGTCAGTCTG-3′ R: 5′-GGGCAGGTCCACCATTTCC-3′ Amplicon length: 104 bp
EGFR (ErBb-1)	Epidermal growth factor receptor (located on chromosome 7) 1. It encodes a transmembrane glycoprotein that is a member of the protein kinase (PK) superfamily. 2. EGFR binds to EGF, thus inducing receptor dimerization and tyrosine autophosphorylation, leading to cell proliferation. 3. Mutations in EGFR are associated with lung cancer.	F: 5′-TGCGTCTCTTGCCGGAAT-3′ R: 5′-GGCTCACCCTCCAGAAGGTT-3′ Amplicon length: 71 bp
MAP2K1 (MEK1)	Mitogen-activated protein kinase kinase 1 (located on chromosome 15) 1. The encoded protein is a member of the dual specificity PK family that acts as a mitogen-activated protein (MAP) kinase kinase. 2. The encoded protein stimulates the enzymatic activity of MAP kinases upon a wide variety of extra- and intracellular signals. 3. As a component of the MAP kinase signal transduction pathway, the encoded protein is involved in many cellular processes (e.g., proliferation, differentiation, and transcription regulation).	F: 5′-CAATGGCGGTGTGGTGTTC-3′ R: 5′-GATTGCGGGTTTGATCTCCAG-3′ Amplicon length: 91 bp
FOLH1 (PSMA)	Folate hydrolase 1 (located on chromosome 11) 1. It encodes a type II transmembrane glycoprotein belonging to the M28 peptidase family. 2. Also known as prostate-specific membrane antigen (PSMA), it is expressed in many tissues, including the prostate. 3. In the prostate, the FOLH1/PSMA protein is upregulated in cancer cells and is used as an effective diagnostic and prognostic indicator of prostate cancer.	F: 5′-AGAGGGCGATCTAGTGTATGTT-3′ R: 5′-TGATTTTCATGTCCCGTTCCAAT-3′ Amplicon length: 74 bp
BUB1	BUB1 mitotic checkpoint serine/threonine kinase (located on chromosome 2) 1. It encodes a protein that plays a central role in mitosis. 2. This protein may also function in the DNA damage response. 3. Mutations in this gene have been associated with aneuploidy and several forms of cancer.	F: 5′-AGCCCAGACAGTAACAGACTC-3′ R: 5′-GTTGGCAACCTTATGTGTTTCAC-3′ Amplicon length: 136 bp
RHOA	Ras homolog family member A (located on chromosome 3) 1. It encodes a member of the Rho family of small GTPases that function as molecular switches in signal transduction cascades. 2. Overexpression of this gene is associated with tumor cell proliferation and metastasis.	F: 5′-GGAAAGCAGGTAGAGTTGGCT-3′ R: 5′-GGCTGTCGATGGAAAAACACAT-3′ Amplicon length: 118 bp
PCSK7	Proprotein convertase subtilisin/kexin type 7 (located on chromosome 11) 1. It encodes a type 1 membrane-bound protease that is expressed in many tissues, including the neuroendocrine, liver, gut, and brain. 2. It has been implicated in the transcriptional regulation of housekeeping genes. 3. A chromosomal translocation associated with B-cell lymphoma occurs between this gene and its inverted counterpart.	F: 5′-GCAGCGTCCACTTCAACGA-3′ R: 5′-GCCCAGTCACATTGCGTTC-3′ Amplicon length: 117 bp
PITX2 (ARP1)	Paired-like homeodomain 2 (located on chromosome 4) 1. It encodes a member of the RIEG/PITX homeobox family, which is in the bicoid class of homeodomain proteins. 3. The encoded protein acts as a transcription factor and regulates procollagen lysyl hydroxylase gene expression.	F: 5′-GCCAAGGGCCTTACATCCG-3′ R: 5′-GGTGGGGAAAACATGCTCTG-3′ Amplicon length: 101 bp
ANXA3 (annexin) A3	Annexin A3 (located on chromosome 4) 1. It encodes a member of the annexin family. 1. Members of this calcium-dependent phospholipid-binding protein family play a role in the regulation of cellular growth and in signal transduction pathways. 2. The encoded protein functions in the inhibition of phospholipase A2 and cleavage of inositol 1,2-cyclic phosphate to form inositol 1-phosphate.	F: 5′-TTAGCCCATCAGTGGATGCTG-3′ R: 5′-CTGTGCATTTGACCTCTCAGT-3′ Amplicon length: 104 bp
B2M	β2-microglobulin (located on chromosome 15) – Housekeeping gene	F: 5′-TGCTGTCTCCATGTTTGATGTATCT-3′ R: 5′-TCTCTGCTCCCCACCTCTAAGT-3′ Amplicon length: 86 bp
ACTB	β-actin (located on chromosome 7) – Housekeeping gene	F: 5′- GCCGGGACCTGACTGACTAC-3′ R: 5′- TTCTCCTTAATGTCACGCACGAT-3′ Amplicon length: 100 bp

* Information available at https://www.ncbi.nlm.nih.gov/ (accessed on 10 February 2021).