SPT3 Overexpression Lysate

Western Blot: SPT3 Overexpression Lysate (Adult Normal) [NBL1-16616] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for SPT3.

• Reactivity: Hu
• Applications: WB

SPT3 Overexpression Lysate Summary

• Description:

  SPT3 Transient Overexpression Lysate

  
  Expression Host: HEK293T
  
  Plasmid: RC208269
  
  Accession#: NM_181356
  
  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: SUPT3H

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: 37.2 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 SPT3 Overexpression Lysate

• SPT3-like protein
• SPT3SPT3L
• suppressor of Ty (S.cerevisiae) 3 homolog
• suppressor of Ty 3 homolog (S. cerevisiae)
• transcription initiation protein SPT3 homolog

Background

The transcription of many RNA polymerase II-dependent genes requires Spt3, a member of the S. cerevisiae SAGA complex. Transcription from delta sequences, the long terminal repeats that flank yeast Ty elements, requires the yeast SPT3 gene. Spt3 and Spt20 work together to recruit TATA-box binding protein (TBP) to the core promoter allowing TBP to bind to SAGA-dependent promoters. Null mutations in the Spt3 gene cause defects in sporulation, diploid filamentous growth, and haploid invasive growth, indicating that Spt3 has an important role in both mating and development pathways in yeast. At the promoters of some genes including yeast HO, HIS3 and TRP3 genes, Spt3 inhibits binding of TBP, resulting in reduced transcription. This repressive effect of Spt3 can be overcome by another member of the SAGA complex, GCN5, which promotes the formation of a TBP/TFIIA complex by histone acetylation.

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: SUPT3H