Total RNA of cell lines was extracted using TRIzol and used to synthesize
complementary DNA (cDNA) by reverse transcription through M-MLV (an
RNA-dependent DNA polymerase that can be used in cDNA synthesis with long
messenger RNA templates) and cDNA amplification using the Power SYBR Green
Master Mix kit (Takara, Otsu, Japan). In addition, total RNA for miRNAs was
collected using a High Pure miRNA isolation kit (Roche) and real-time polymerase
chain reaction (RT-PCR) using a TaqMan MicroRNA Reverse Transcription kit (Life
Technologies, Carlsbad, CA, USA). A miScript Primer Assay (QIAGEN) was used for
the miR-191-5p and U6. Data were analyzed using the comparative Ct method (2–ΔΔCt). The procedure of
quantification of PD-L1 expression in RT-PCR was conducted as previously described.21 (link) The primers are listed inSupplementary Table S2 .
complementary DNA (cDNA) by reverse transcription through M-MLV (an
RNA-dependent DNA polymerase that can be used in cDNA synthesis with long
messenger RNA templates) and cDNA amplification using the Power SYBR Green
Master Mix kit (Takara, Otsu, Japan). In addition, total RNA for miRNAs was
collected using a High Pure miRNA isolation kit (Roche) and real-time polymerase
chain reaction (RT-PCR) using a TaqMan MicroRNA Reverse Transcription kit (Life
Technologies, Carlsbad, CA, USA). A miScript Primer Assay (QIAGEN) was used for
the miR-191-5p and U6. Data were analyzed using the comparative Ct method (2–ΔΔCt). The procedure of
quantification of PD-L1 expression in RT-PCR was conducted as previously described.21 (link) The primers are listed in
