mRNA Quantification

Total RNA from wild-type (WT) MEFs and C1qbp-KO MEFs transfected with various C1qbp expression vectors was isolated with an RNeasy Mini Kit (QIAGEN) according to the manufacturer’s instructions. The concentration and purity of total RNA were measured on a NanoDrop spectrophotometer (NanoDrop Technologies). Reverse transcription of 1 μg of total RNA was performed with a PrimeScript RT Reagent Kit (TAKARA). Mouse C1qbp mRNA and mouse 18S ribosomal RNA (control) were detected by quantitative PCR with SYBR Premix Ex Taq II (TAKARA) and a thermal cycler (StepOnePlus, Applied Biosystems). Primers were as follows: 5′-CGCGGTTCTATTTTGTTGGT-3′ (18S rRNA forward), 5′-AGTCGGCATCGTTTATGGTC-3′ (18S rRNA reverse), 5′-GGCCTTCGTTGAATTCTTGA-3′ (C1qbp mRNA forward), and 5′- GCCTCATCTTCGTGTCCAAT-3′ (C1qbp mRNA reverse). An unpaired Student’s t test was used to determine statistical differences between two groups. Values of ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.005 were considered to be statistically significant.