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Snrp70

Manufactured by Abcam

SNRP70 is a protein that plays a role in the spliceosome, a complex of small nuclear ribonucleoproteins (snRNPs) and other proteins that catalyze the removal of introns from precursor messenger RNA. SNRP70 is a component of the U1 snRNP, which is involved in the early stages of spliceosome assembly.

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3 protocols using snrp70

1

Subcellular Fractionation and Molecular Analyses

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Western blot and quantitative RT-qPCR (RT-qPCR) of marker proteins/transcripts for HEK293-subcellular compartments were used to verify the subcellular fractionation results. Western blots were performed with antibodies for three proteins – GAPDH (Abcam), SNRP70 (Abcam) and histone H3 (Abcam). Samples of subcellular fractions were boiled at 95°C for 10 minutes, then spun at 14000g for 3 minutes at room temperature to minimize the influence of sticky DNA (especially in the chromatin samples) on Western blots. For every Western blot, 1% of the sample volume was used. For RT-qPCR, the same percentage (1%) of RNA samples was used. Then the ratio of each marker gene (GAPDH, U1, ACTIN [intron]) was calculated in the respective chromatin, nucleoplasmic and cytoplasmic fractions.
For mES cells, similar western-blot experiments were carried out using antibodies against actin (Abcam), histone H3 (Abcam) and SNRP70 (Abcam). We confirmed NAI-N3 treatment didn’t affect fractionation. Protein was quantified using the Pierce BCA protein assay kit (Thermo Fischer Scientific). We loaded the protein from each compartment in proportion to the amount obtained (roughly 1:1:2 for chromatin:nucleoplasm:cytoplasm).
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2

Subcellular Fractionation and Molecular Analyses

Check if the same lab product or an alternative is used in the 5 most similar protocols
Western blot and quantitative RT-qPCR (RT-qPCR) of marker proteins/transcripts for HEK293-subcellular compartments were used to verify the subcellular fractionation results. Western blots were performed with antibodies for three proteins – GAPDH (Abcam), SNRP70 (Abcam) and histone H3 (Abcam). Samples of subcellular fractions were boiled at 95°C for 10 minutes, then spun at 14000g for 3 minutes at room temperature to minimize the influence of sticky DNA (especially in the chromatin samples) on Western blots. For every Western blot, 1% of the sample volume was used. For RT-qPCR, the same percentage (1%) of RNA samples was used. Then the ratio of each marker gene (GAPDH, U1, ACTIN [intron]) was calculated in the respective chromatin, nucleoplasmic and cytoplasmic fractions.
For mES cells, similar western-blot experiments were carried out using antibodies against actin (Abcam), histone H3 (Abcam) and SNRP70 (Abcam). We confirmed NAI-N3 treatment didn’t affect fractionation. Protein was quantified using the Pierce BCA protein assay kit (Thermo Fischer Scientific). We loaded the protein from each compartment in proportion to the amount obtained (roughly 1:1:2 for chromatin:nucleoplasm:cytoplasm).
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3

Antibody Panel for Cell Signaling Analysis

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The antibodies used were as follows: NF-κB p65 (Abcam; cat. no. ab7970), Pol II (Santa Cruz; cat. no. SC-899 and SC-900, mixed in a 1:4 ratio), β-tubulin (Abcam; cat. no.ab6046), SNRP70 (Abcam; cat. no ab51266), c-Fos (H-125, Santa Cruz; cat. no. sc-7202), c-Jun (H-79, Santa Cruz; cat. no. sc-1694), and NF-κB p50 (Abcam; cat. no. ab7971).
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