Fenozol

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	RNA from total mice lungs were isolated using Fenozol (A&A Biotechnology). The quantity of ribosomal RNA and DNA contamination was examined using electrophoresis in 1% denaturing formaldehyde gel.

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RNA from total mice lungs were isolated using Fenozol (A&A Biotechnology). The quantity of ribosomal RNA and DNA contamination was examined using electrophoresis in 1% denaturing formaldehyde gel.

Publication protocol

Total RNA from cultured cells and tissues was isolated using the Universal RNA Purification Kit (EURx). RNA from total mice lungs were isolated using Fenozol (A&A Biotechnology). The quantity of ribosomal RNA and DNA contamination was examined using electrophoresis in 1% denaturing formaldehyde gel. Concentration of total RNA was assessed using NanoDrop 1000 Spectrophotometer (Thermo Fisher Scientific). Reverse transcription was performed using 1 μg of total RNA, oligo(dT) 15 primer (Promega) and M-MLV reverse transcriptase (Promega). Real-time PCR was carried out using SybrGreen Master Mix (A&A Biotechnology) and Eco Real-Time PCR System (Illumina). For the examination of mice lung metastasis, specific probes for human GAPDH and mouse GAPDH (Life Technologies) were used with Taq PCR Master Mix (EURx). Gene expression was normalized to elongation factor-2 (EF2). mRNA level in each sample was analyzed in duplicates. The relative level of transcripts was quantified by the ΔΔCt method. Sequences of primers (Genomed) and annealing temperatures are listed in Table S1 in Supplementary Material.

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