PABP Overexpression Lysate

Western Blot: PABP Overexpression Lysate (Adult Normal) [NBP2-10839] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for PABP.

• Reactivity: Hu
• Applications: WB

PABP Overexpression Lysate Summary

• Description:

  PABP Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC207354
  
  Accession#: NM_002568
  
  Protein Tag: C-MYC/DDK
  
  You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
• Gene: PABPC1

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.
  
  Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.
• Theoretical MW: 70.5 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

• Storage: Store at -80C. Avoid freeze-thaw cycles.
• Buffer: RIPA buffer

Lysate Details for Array

• Type: Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for PABP Overexpression Lysate

• PAB1polyadenylate-binding protein 1
• PABP
• PABP-1
• PABP1poly(A) binding protein, cytoplasmic 2
• PABPC2
• PABPL1
• poly(A) binding protein, cytoplasmic 1
• Poly(A)-binding protein 1
• poly(A)-binding protein, cytoplasmic 2

Background

The poly(A)-binding protein (PABP), which is found complexed to the 3-prime poly(A) tail of eukaryotic mRNA, is required for poly(A) shortening and translation initiation. Grange et al. (1987) isolated a melanoma cell cDNA encoding human PABP. The predicted 633-amino acid protein contains 4 repeats of an approximately 80-amino acid unit in its N-terminal half. The authors found that this repeat region is highly conserved between human and yeast PABP and is sufficient for poly(A) binding. In vitro translation of the human PABP cDNA yielded a protein with an apparent molecular mass of 73 kD by SDS-PAGE. Northern blot analysis indicated that PABP is expressed as a 2.9-kb mRNA in human melanoma cells. Gorlach et al. (1994) noted that each of the 4 repeats of PABP is a ribonucleoprotein (RNP) consensus sequence RNA-binding domain. They determined that PABP has a pI of approximately 10.3 and is a very abundant, stable protein. Immunofluorescence studies of mammalian cells indicated that PABP is located exclusively in the cytoplasm. However, using both indirect immunofluorescence and tagging of PABP1 by fusion to the green fluorescent protein (GFP), Afonina et al. (1998) demonstrated that PABP1 shuttles between the nucleus and cytoplasm. PABP1 accumulated in the nucleus when transcription was inhibited, suggesting that active transcription is required for nuclear export of PABP1.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Bioinformatics

• Gene Symbol: PABPC1