Journal: Scientific Reports

Article Title: Evaluating the quantity, quality and size distribution of cell-free DNA by multiplex droplet digital PCR

doi: 10.1038/s41598-020-69432-x

Figure Lengend Snippet: A multiplex ddPCR assay to QC cfDNA samples. Our assay permits discriminating between OR cfDNA fragments (black bars) of different sizes. Annealing sites for more than one forward primer (black arrows) and hydrolysis probe (blue bar: 6-FAM-labelled probe; green bar: HEX-labelled probe) only become available in OR cfDNA fragments exceeding 165 bp. Our assay also includes the co-amplification of a diploid locus ( STAT6 ) to estimate absolute DNA concentration. A visualization of the raw data derived from the analysis of 1 ng of cfDNA can be observed in the right panel.

Article Snippet: A 20 × pre-mix of primers and probes (1:1 primers/probe ratio) for the STAT6 locus was ordered from Bio-Rad (Hercules, CA, USA).

Techniques: Multiplex Assay, Amplification, Concentration Assay, Derivative Assay

Journal: Oncology Letters

Article Title: MicroRNA-135a induces prostate cancer cell apoptosis via inhibition of STAT6

doi: 10.3892/ol.2018.9791

Figure Lengend Snippet: ONCOMINE® prostate cancer database analyses. (A) STAT6 was overexpressed in prostate cancer tissues compared with normal prostate gland tissues in the Wallace study (*P<0.0001). (B) The tumors in the TCGA dataset have an increased STAT6 copy number (*P=0.0002). (C) Increased levels of STAT6 expression were associated with a reduced overall survival rate in the Grasso study (P=0.0009). (D) Increased levels of STAT6 expression were associated with the biochemical relapse rate (P=0.10) following radical prostatectomy in the Lapointe study. (E and F) GSEA analyses of gene ontology terms indicated that there was enriched expression of gene sets involved in (E) cell apoptosis (P=0.001) and (F) cancer survival (P<0.001). (G and H) Microarray data from the GSE40026 and GSE21032 datasets were retrieved and revealed that miR-135a was underexpressed, whereas STAT6 was overexpressed in DU145 and PC3 cells compared with LNCaP cells (P=0.10). STAT6, signal transducer and activator or transcription 6; TCGA, The Cancer Genome Atlas; GSEA, Gene Set Enrichment Analysis; GSEA, Gene Series; miR-135a, microRNA-135a.

Article Snippet: The STAT6 target probe was synthesized with a 3′-biotin modification (Invitrogen; Thermo Fisher Scientific, Inc.) and the sequence was 5′-acg ttg aat TTCTAAGAA aga ggg a-3′, in which the binding region is underlined.

Techniques: Expressing, Microarray

Journal: Oncology Letters

Article Title: MicroRNA-135a induces prostate cancer cell apoptosis via inhibition of STAT6

doi: 10.3892/ol.2018.9791

Figure Lengend Snippet: STAT6 is a direct target of miR-135a in prostate cancer cells. (A) TargetScan and RNAhybrid databases revealed that miR-135a may bind to a target sequence located in the region of nucleotides 1,100-1,107 of the 3′UTR of STAT6. (B) The STAT6 3′UTR was cloned into a luciferase reporter cloning site in the pMIR-report™ Luciferase miRNA Expression Reporter vector and increased miR-135a level was associated with decreased luciferase activity. *P<0.05. STAT6, signal transducer and activator or transcription 6; miR-135a, microRNA-135a; UTR, untranslated region; miRNA, microRNA; NC, negative control.

Article Snippet: The STAT6 target probe was synthesized with a 3′-biotin modification (Invitrogen; Thermo Fisher Scientific, Inc.) and the sequence was 5′-acg ttg aat TTCTAAGAA aga ggg a-3′, in which the binding region is underlined.

Techniques: Sequencing, Clone Assay, Luciferase, Expressing, Plasmid Preparation, Activity Assay, Negative Control

Journal: Oncology Letters

Article Title: MicroRNA-135a induces prostate cancer cell apoptosis via inhibition of STAT6

doi: 10.3892/ol.2018.9791

Figure Lengend Snippet: Protein expression levels and DNA-binding ability of STAT6 and p-STAT6 in prostate cancer cells treated with miR-135a and NC. (A) Endogenous STAT6 and p-STAT6 stimulated with IL-4 were inhibited by transfection with miR-135a in western blot analysis. (B) An electrophoretic mobility shift assay was performed to assess the binding of a consensus STAT6 DNA oligo probe using miR-135a and NC nuclear extracts and indicated a decrease in DNA-binding activity in prostate cancer cells following induced miR-135a expression. STAT6, signal transducer and activator or transcription 6; p-STAT6, phosphorylated signal transducer and activator or transcription 6; miR-135a, microRNA-135a; IL-4, interleukin 4; NC, negative control.

Article Snippet: The STAT6 target probe was synthesized with a 3′-biotin modification (Invitrogen; Thermo Fisher Scientific, Inc.) and the sequence was 5′-acg ttg aat TTCTAAGAA aga ggg a-3′, in which the binding region is underlined.

Techniques: Expressing, Binding Assay, Transfection, Western Blot, Electrophoretic Mobility Shift Assay, Activity Assay, Negative Control