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ythdc1  (Cell Signaling Technology Inc)
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after <t>YTHDC1</t> knockdown. (G) Quantification of western blot (n=3).
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after <t>YTHDC1</t> knockdown. (G) Quantification of western blot (n=3).
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after <t>YTHDC1</t> knockdown. (G) Quantification of western blot (n=3).
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after <t>YTHDC1</t> knockdown. (G) Quantification of western blot (n=3).
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after <t>YTHDC1</t> knockdown. (G) Quantification of western blot (n=3).
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anti ythdc1  (Cell Signaling Technology Inc)
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after <t>YTHDC1</t> knockdown. (G) Quantification of western blot (n=3).
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(A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after YTHDC1 knockdown. (G) Quantification of western blot (n=3).

Journal: bioRxiv

Article Title: Regulation and Function of the HPV16 CircE7 RNA

doi: 10.64898/2026.03.09.710444

Figure Lengend Snippet: (A) Western blot from MDA-1483 cells transfected with WT or IRES mut circE7 plasmid. Representative blot (left), quantification (right). ***p<0.001 (B) RNA levels of the IRES mutant cricE7 RNA by RT-qPCR in MDA-1483 cells. Difference not statistically significant by t-test. Twice as much RNA was transfected in to get equivalent levels of RNA between IRES and WT. (C) Representative tracing of circE7-transfected cells after polysome enrichment assay with the monosome (M), light polysome (L), and heavy polysome (H) fractions indicated. Polysomes isolated from SCC-154 HPV+ (red) and MDA1483 HPV- (blue) cells. (D) Endpoint PCR of CircE7 or circHPK3, in light and heavy polysomes from SCC-154 (HPV+) cells, MDA1483 (HPV-) cells, and BJ cells transfected with hTERT circE7. All samples were RNAse R treated. (E) Western blot from HeLa cells co-transfected with a circE7-Flag expression vector and the indicated siRNA. HSP90, loading control. *indicates non-specific band. (F) RT-qPCR for circE7 RNA levels after YTHDC1 knockdown. (G) Quantification of western blot (n=3).

Article Snippet: Membranes were incubated overnight at 4°C with the following primary antibodies: YTHDC1 (Cell Signaling, 77422), YTHDF3 (Cell Signaling, 24206), YTHDC2 (Cell Signaling, 46324), FLAG (Sigma, A8592), GAPDH (Santa Cruz, sc-32233), HSP90 (Cell Signaling, 4877S).

Techniques: Western Blot, Transfection, Plasmid Preparation, Mutagenesis, Quantitative RT-PCR, Isolation, Expressing, Control, Knockdown