RNA was isolated using the QIAGEN RNeasy Micro Kit (QIAGEN) or the ZR-96 Quick RNA (Zymo Research, Irvine, CA) following manufacturer′s guidelines and converted to complementary DNA by reverse transcription (RT) with random hexamers and Multiscribe RT (TaqMan Reverse Transcription Reagents; Thermo Fisher Scientific). For mRNA gene expression assays, probes were obtained from Thermo Fisher Scientific, and the reactions were set up following the manufacturer's guidelines and run on a StepOnePlus Real-Time PCR System (Thermo Fisher Scientific). Values are represented as the difference in cycle threshold (Ct) values normalized to β2-microglobulin for each sample as per the following formula: Relative RNA expression = (2—ΔCt) × 1000. Gene expression data were arranged in heat maps in Fig. 1 to identify virus-specific cytokine expression patterns. Colors are assigned using linear conditional formatting based on relative expression values of each gene for each donor and virus infection as compared to the expression value of that gene for all other donors and virus infections. Red denotes high expression and blue low expression.

For mRNA expression assays, the following probes were used: ATF1 Hs00909673_m1, ATF2 Hs01095345_m1, ATF3 Hs00231069_m1, ATF4 Hs00909569_g1, ATF5 Hs01119208_m1, ATF6 Hs00232586_m1, ATF7 Hs00232499_m1, B2M Hs00187842_m1, BST2 Hs00171632_m1, CCL20 Hs00171125_m1, CD40 Hs01002913_g1, CD74 Hs00269961_m1, CFOS Hs04194186_s1, CXCL10 Hs00171042_m1, CXCL8 Hs99999034_m1, EBI3 Hs01057148_m1, EIF2AK2 Hs00169345_m1, FOS Hs04194186_s1, FosB Hs00171851_m1, FOSL1 Hs04187685_m1, FOSL2 Hs01050117_m1, FOXP3 Hs01085834_m1, GATA3 Hs00231122_m1, GZMB Hs01554355_m1, ICOSLG Hs00323621_m1, IFITM1 Hs00705137_s1, IFITM2 Hs00829485_sH, IFNA1 Hs00855471_g1, IFNA2 Hs00265051_s1, IFNB1 Hs01077958_s1, IFNG Hs00989291_m1, IL10 Hs001174086_m1, IL12B Hs01011519_m1, IL1B Hs99999029_m1, IL17A Hs00174383_m1, IL21 Hs00222327_m1, IL23A Hs00372324_m1, IL27 Hs00377366_m1, IL4 Hs00174122_m1, IL6 Hs00174131_m1, IRF1 Hs009171965_m1, IRF3 Hs01547283_m1, IRF5 Hs00158114_m1, IRF7 Hs00185375_m1, IRF9 Hs00196051_m1, ISG15 Hs00192713_m1, JDP2 Hs00185689_m1, JUN Hs01103582_s1, JUNB Hs00357891_s1, JUND Hs04187679_s1, MOV10 Hs00253093_m1, MX2 Hs01550808_m1, RORC Hs01076122_m1, TBX21 Hs00203436_m1, TGFB1 Hs00998133_m1, TLR1 Hs00413978_m1, TLR2 Hs01014511_m1, TLR3 Hs01551078_m1, TLR4 Hs00152939_m1, TLR5 Hs00152825_m1, TLR6 Hs00271977_m1, TLR7 Hs00152971_m1, TLR8 Hs00152972_m1, TLR9 Hs00152973_m1, TLR10 Hs01675179_m1, TNF Hs00174128_m1, TREX1 Hs03989617_s1, TRIM5 Hs01552559_m1.

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



Q&A
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.