Total RNA was purified from cells using the
RNeasy Universal Mini Kit (Qiagen) following manufacturer’s specifications. Purified RNA was then transcribed to cDNA using the
SuperScript III Reverse Transcriptase system (Thermo Fisher Scientific), and subjected to qPCR reactions using the
FastStart Universal SYBR Green Master (Rox) system (Roche) with primers targeting mouse
GAPDH, IL-6, MIP-1α and
Arg1. Primer sequences are as follows:
| TREM2, | forward 5’-AGGGCCCATGCCAGCGTGTGGT-3’, |
| Reverse 5’-CCAGAGATCTCCAGCATC-3’; |
| GAPDH, | forward 5’-CCCTTCATTGACCTCAACTA-3’, |
| reverse 5’-CCTTCTCCATGGTGGTGAA-3’; |
| IL-6, | forward 5′-TAGTCCTTCCTACCCCAATTTCC-3′, |
| reverse 5′-TTGGTCCTTAGCCACTCCTTC-3′. |
| MIP-1α, | forward 5′-CCCAGCCAGGTGTCATTTTCC-3′, |
| reverse 5′-GCATTCAGTTCCAGGTCAGTG-3′; |
| Arg1, | forward 5′-CACAGTCTGGCAGTTGGAAGC-3′, |
| reverse 5′-CTTTGGCAGATATGCAGGGAG-3′. |
These experiments were repeated 3 times. Randomization procedures are not applicable to this assay. Data were collected and analyzed in a double-blind fashion. No statistical methods were used for sample size estimation. No data were excluded.
Zhao Y., Wu X., Li X., Jiang L.L., Gui X., Liu Y., Sun Y., Zhu B., Piña-Crespo J.C., Zhang M., Zhang N., Chen X., Bu G., An Z., Huang T.Y, & Xu H. (2018). TREM2 is a receptor for β-amyloid which mediates microglial function. Neuron, 97(5), 1023-1031.e7.
