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The spots in panels C2 and C3 contain 10 ug of Px.
1.5 – 5 ug of Purified DNA (5ug recommended) suspended in TE Buffer in a volume not to exceed 50 ul.
1 – 5 ug of Purified DNA (5ug recommended) suspended in TE Buffer in a volume not to exceed 50 ul.
1 2 ug of total RNA from each sample was used to construct an RNA-seq library using TruSeq RNA sample preparation v2 kit (Illumina).
Briefly, 2 ug of total RNA was reverse transcribed into cDNA using a Revert Aid First Strand cDNA Synthesis Kit (Thermo).
For immunoprecipitation (IP), 500 ug of total lysate was precleared for 1 hour.
We have performed in-solution digestion and analysis of samples with as little as 0.1 ug of protein.
Approximately 2 ug of total RNAs were used for reverse transcription using the Reverse Transcription System (Promega, USA).
10 uL (IDA) and 20 uL (MRM experiment) sample loop volumes provided injection of ~1 ug of digest (25 fM of standard peptides) and ~2 ug of digest (50 fM of standard peptides) for IDA and MRM runs, respectively.
500 ug of membrane protein was used for each IP.
Approximately 10 ug of protein was injected per fraction.
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