To isolate total cellular RNA, cells lysed in 1 ml of TRIzol were mixed with 200 μl of chloroform and centrifuged at 12,000g at 4°C for 10 min. The RNA in the aqueous phase was precipitated with isopropanol, pelleted by centrifugation at 16,000g at 4°C for 15 min, and subsequently washed twice with 70% ethanol. One microgram of RNA was reverse-transcribed into complementary DNA (cDNA) with the High-Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) according to the manufacturer’s instructions. Gene expression was quantified by qPCR using Fast SYBR Green Master Mix (Thermo Fisher Scientific) and the following primers: hIFNA1, 5′-TGGAAGCCTGTGTGAT-3′ (forward) and 5′-ATGATTTCTGCTCTGACA-3′ (reverse); hIFNB, 5′-GTCAGAGTGGAAATCCTAAG-3′ (forward) and 5′-CTGTAAGTCTGTTAATGAAG-3′ (reverse); hB2M, 5′-CTCCGTGGCCTTAGCTGTG-3′ (forward) and 5′-TTTGGAGTACGCTGGATAGCCT-3′ (reverse); mGAPDH, 5′-AACTTTGGCATTGTGGAAG-3′ (forward) and 5′-ACACATTGGGGGTAGGAAC-3′ (reverse); mMX1, 5′-GATCCGACTTCACTTCCAGATGG-3′ (forward) and 5′-CATCTCAGTGGTAGTCAACCC-3′ (reverse); mIFIT1, 5′-TGCTGAGATGGACTGTGAGG-3′ (forward) and 5′-CTCCACTTTCAGAGCCTTCG-3′ (reverse); mIFIT2, 5′-GGAGAGCAATCTGCGACAG-3′ (forward) and 5′-GCTGCCTCATTTAGACCTCTG-3′ (reverse); mIFIH1, 5′-ATGGACGCAGATGTTCGTGG-3′ (forward) and 5′-TCCCTTCTCGAAGCAAGTGTC-3′ (reverse); mCXCL10, 5′-CCAAGTGCTGCCGTCATTTT-3′ (forward) and 5′-CTCAACACGTGGGCAGGATA-3′ (reverse); and mOAS1A, 5′-GCCTGATCCCAGAATCTATGC-3′ (forward) and 5′-GAGCAACTCTAGGGCGTACTG-3′ (reverse). Expression values were normalized by the ΔΔCt method.

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.