2.10 Quantitative real-time PCR analysis
Total RNA was extracted from cells using TRIzol reagent (Invitrogen),
according to the manufacturer’s instructions. RNA was reverse
transcribed into cDNA using a reverse transcription system (Vazyme,
Nanjing, China). Real-time PCR was performed using a LightCycler® 480
System (Roche) with AceQ Universal SYBR qPCR Master Mix (Vazyme
Biotechnology, Nanjing, China). The 2−ΔΔCT method was
used to calculate the relative mRNA levels. GAPDH was used as an
internal reference. The primer sequences used for quantitative real-time
PCR
were:
TGF-β forward 5′-CCACCTGCAAGACCATCGAC-3′ and reverse
5′-CTGGCGAGCCTTAGTTTGGAC-3′.
Arg-1 forward
5′-CTCCAAGCCAAAGTCCTTAGAG-3′
and reverse 5′-AGGAGCTGTCATTAGGGACATC-3′, GAPDH forward
5′-AACGACCCCTTCATTGAC-3′ and reverse 5′-TCCACGACATACTCAGCAC-3′.