Argonaute CLIP defines a deregulated miR-122 bound transcriptome that correlates with patient survival in human liver cancer - Raw autoradiograms and Western Blots

Published: 23 Jun 2017 | Version 4 | DOI: 10.17632/v4wp2mmztd.4
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Description of this data

JL0109 - Autoradiogram of 32P-labelled RNA crosslinked to IP purified AGO from five normal adjacent (left) or matched HCC tumor from from livers of human HCC patients (right) using the 2A8 anti-Ago antibody, exposed overnight after running on an 8% Novex NuPage gel. Irrelevant antibody control “IgG”, non-crosslinked control "-XL", and high “Hi” RNAse A control are shown. RNA-protein complexes from 110-150kDa were used for subsequent RNA sequencing library preparation. A weak band of ~110kDa is typically observed under non-crosslinked conditions. Protein size markers labeled with boxes at indicated molecular weights. Mass of sample (in milligrams) are indicated beneath each lane.

JL0173 - Autoradiogram of 32P-labelled RNA crosslinked to IP purified AGO from four 122KO (top) or five WT floxed control livers (bottom) using the 2A8 anti-Ago antibody, exposed overnight after running on an 8% Novex NuPage gel. Irrelevant antibody control “IgG”, non-crosslinked control "-XL", and high “Hi” RNAse A control are shown. RNA-protein complexes from 110-150kDa were used for subsequent RNA sequencing library preparation. A weak band of ~110kDa was typically observed under non-crosslinked conditions. Protein size markers labeled with boxes at indicated molecular weights. Mass of sample (in milligrams) and sex of animal are indicated beneath each lane.

Fig 7A - FLAG-tagged human BCL9 expression levels in 293T cells after transfection with 50nM control or miR-122 mimic. WT BCL9 is compared to a mutant where all 6 exonic miR-122 sites were mutated while preserving amino acid sequence. Blots were imaged on a LiCor with ImageStudio Software V5.2. Short exposure displays Flag BCL9 channel (800nm) and GAPDH channel (680nm). Long exposure shows only the flag-BCL9 channel signal (800nm).

Fig 7B - FLAG-tagged human BCL9 expression levels in MHCC-LM3 cells after transfection with 50nM control or miR-122 mimic. WT BCL9 is compared to a mutant where all 6 exonic miR-122 sites were mutated while preserving amino acid sequence. Blots were imaged on a LiCor with ImageStudio Software V5.2. Short exposure used for GAPDH normalization (lower set of bands). Long exposure shows only the flag-BCL9 channel signal in the upper set of bands.

Fig S7 - FLAG-tagged human BCL9 expression charcterization. Immunoblots of H293T cells transfected with empty vector (lane 1), FLAG-BCL9 expression vector (lane 2), or no vector (lane 3) and harvested after 48 hours. Blots were probed with anti-FLAG and anti-tubulin (red channel) and anti-BCL9 (green channel) antibodies. Merged channel is also shown.

Experiment data files

This data is associated with the following publication:

Argonaute CLIP Defines a Deregulated miR-122-Bound Transcriptome that Correlates with Patient Survival in Human Liver Cancer

Published in: Molecular Cell

Latest version

  • Version 4

    2017-06-23

    Published: 2017-06-23

    DOI: 10.17632/v4wp2mmztd.4

    Cite this dataset

    Luna, Joseph (2017), “Argonaute CLIP defines a deregulated miR-122 bound transcriptome that correlates with patient survival in human liver cancer - Raw autoradiograms and Western Blots”, Mendeley Data, v4 http://dx.doi.org/10.17632/v4wp2mmztd.4

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Institutions

Ohio State University, Rockefeller University

Categories

Western Blot, Autoradiography

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