Quantitative Gene Expression Analysis

For quantitative gene expression analysis, 40 cycles of real-time PCR were performed on the StepOnePlus real-time detection system (Applied Biosystems). Every PCR reaction was carried out in duplicates with 2.5 ng of cDNA in a final volume of 12.5 μl Power SYBR® Green PCR Master Mix (Applied Biosystem). StepOneTM Software v2.2 was used for data analysis. Ppia was used as housekeeping gene to normalize target gene expression. We used the following primer sequences for detection of Pdpn transcripts (41 (link)): FW 5′-agagaacacgagagtacaacc-3′; RV 5′-caacaatgaagatccctccgac-3′, Arg1 (42 (link)): FW 5′-TTGGGTGGATGCTCACACTG-3′; RV 5′-TTGCCCATGCAGATTCCC-3′; Ppia: FW 5′-AATTCATGTGCCAGGGTGGTG-3′; RV 5′-TGCCTTCTTTCACCTTCCCAA-3′. Additional primer sequences are given in Table S1.