TRIzol Reagent

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	- For low 260/230 readings the best approach is to try washing sample (precipitation) with ethanol to desalt it.

Protocol tips
- For low 260/230 readings the best approach is to try washing sample (precipitation) with ethanol to desalt it.

Publication protocol

Total RNA was isolated from cells using TRIzol reagent (Invitrogen) and RNeasy Plus Mini Kit (Qiagen). Genomic DNA was eliminated with RNase free DNase 1 (Roche), and total RNA was then subjected to another round of purification by phenol solution with pH4.3 ± 0.2 (Sigma-Aldrich).

For Gdf3 RNA was isolated from whole bone marrow mononuclear cells or CD45.2+ / LSK cells using TRIzol reagent (Invitrogen) and RNeasy mini kit (QIAGEN). cDNA was synthesized with Superscript First-Strand Synthesis System according to manufacturer's instructions (Invitrogen). qPCR was performed with SYBR reagent (Bio-Rad). Primers used were: Gdf3_F1 (Forward): 5’- GAA CCT GCT TCA GCT TCT CC-3’; Gdf3_R1 (Reverse): 5’-CCA TGG TCA ACT TTG CCT TT-3’. Gdf3 expression levels were normalized to that of Gapdh: Forward 5’- CCA GCC TCG TCC CGT AGA C-3’; Reverse 5’-CCC TTG ACT GTG CCG TTG-3’.

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Manufacturer protocol

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