Table 1.
“As is” primer pairs (A to S) and “Mix and match” primer pairs (a to f) along with their location on the SOX2 gene as well as the resulting PCR product sizes. The reference numbers in the last column cite the peer-reviewed articles from which the primer-sequences were taken. The list of primer-pair references is included in S1-Table. (SOX2 SRY-box 2 [Homo sapiens (human)] Gene ID: 6657, updated on 7-Jan-2018. NC_000003.12:181711924–181714436 Homo sapiens chromosome 3, GRCh38.p7 Primary Assembly.).
Fig 1.
Position of SOX2 primers on the gene (Gene ID: 6657).
Upper case letters indicate the primer pairs selected from various publications and each pair used as described or “as is”. The lower case letters denote the primer pairs that are “mixed and matched” to cover the entire gene, i.e. 5’ UTR, exon and the 3’ UTR. (SOX2 SRY-box 2 [Homo sapiens (human)] Gene ID: 6657, updated on 7-Jan-2018, NC_000003.12:181711924–181714436 Homo sapiens chromosome 3, GRCh38.p7 Primary Assembly.).
Fig 2.
Parts of SOX2 gene found associated with exosomes originated from stem cells NSCs, cancer cells GBMs, cancer stem cells CD133+ GBMs and SH-SY5Y.
The parts of exosomal SOX2 confirmed with BLAST analysis are highlighted in gray. Binding sites for some of the miRNAs are shown in red and green triangles in 3’UTR. (Details of the clones in supplementary material: S3 Fig).
Fig 3.
Comparison of SNP in NSC, GBM and SH-SY5Y PCR products.
The original nucleotide FASTA sequences of the clones obtained from exosomal DNA amplified with hSOX2- F-18/R-18 (1885–2398). PCR product cloned into the pCR4-TOPO-TA vector. In the BLAST analysis, NSC clone shows a 100% identity to the SOX2 gene. Clones from GBM exosomal DNA, CD133+ GBM’s exosomal DNA (denoted in the figure as “GBMCS” for GBM cancer stem cells) and SH-SY5Y exosomal DNA show multiple SNPs. The SNPs are presented in bold letters. An NCBI reported SNP present in exosomal clones is underlined. The SNP rs1297749385 (3:181714249 T>C) reported in the NCBI database is found only in the exosomal DNA of CD133+ GBM clones. Some of the SNPs identified are reported in the NCBI database. (A detailed version of this figure is provided in supplementary figure: S5 Fig).
Fig 4.
Comparison of SNP in nucleotide sequences of NSC, GBM and SH-SY5Y PCR products.
The original nucleotide FASTA sequences of the clones obtained from exosomal DNA amplified with hSOX2- F-19/R-19 (2328–2686). PCR product cloned into the pCR4-TOPO-TA vector. In the BLAST analysis, NSC and SH-SY5Y clones showed no SNP, whereas clone from GBM exosomal DNA and CD133+ GBM exosomal DNA (denoted in the figure as “GBMCS” for GBM cancer stem cells) show 1 and 2 SNP respectively. The SNPs are presented in bold letters. (A detailed version of this figure is provided in the supplementary figure: S6 Fig).
Fig 5.
A SOX2 SNP, rs11915160, at chr3:181713783 (A>C) evaluated for susceptibility to breast cancer.
The comparison of original nucleotide FASTA sequences of the clones obtained from exosomal DNA amplified with hSOX2- F-11/R-13 (1440–1963). PCR product cloned into the pCR4-TOPO-TA vector. In the BLAST analysis, NSC, GBM, CD133+GBM and SH-SY5Y exosomal DNA clones (denoted in the figure as “GBMCS” for GBM cancer stem cells) show this particular SNP. The SNP is presented in bold letters. Information about the SNP is available in the NCBI database. (A detailed version of this figure is provided in the supplementary figure: S7 Fig).