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AGO set
Tag Antibody Set 

RFA | Baker's yeast replication factor A

AS07 214  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: Saccharomyces cerevisiae

RFA | Baker's yeast replication factor A in the group Antibodies, Bacterial/Fungal at Agrisera AB (Antibodies for research) (AS07 214)

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Datasheet Product citations Add review

Product Information

Immunogen

RPA from Saccharomyces cerevisiae consisting of three subunits RFA1 (70 kDa), RFA2 (30 kDa) and RFA3 (14 kDa); overexpressed in E.coli and purified by chromatography; no affinity tags were added to any of three subunits

Host Rabbit
Clonality Polyclonal
Purity Serum
Format Lyophilized
Quantity 50 µl
Reconstitution For reconstitution add 50 µl of sterile water
Storage Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from lyophilized material adhering to the cap or sides of the tubes.
Tested applications Immunoprecipitation (IP), Chromatin Immunoprecipitation (ChIP), Western blot (WB)
Recommended dilution ChIP, 1 : 20 000 (WB)
Expected | apparent MW

70 + 30 + 14 kDa

Reactivity

Confirmed reactivity Saccharomyces cerevisiae
Predicted reactivity

Saccharomyces cerevisiae

Not reactive in No confirmed exceptions from predicted reactivity are currently known.

Application examples

Application examples

application example

 

TCA precipitated protein extracts from a wild type yeast strain (S. cerevisiae) were separated on 10% gel and transferred to a PVDF membrane. Antibody was used in different dilutions: 1: 5000 (1); 1: 10 000 (2); 1: 20 000 (3);

Besides the bands for RFA1 and RFA2 an unspecific band was detected at ~150 kDa.

 

 

Additional information

Antibody was also successfully used in ChIP application Holstein et al. (2014).

Load of 1 ng of the protein will allow to visualize two subunits of RPA, while load of 5 ng will allow to visualize all three subunits in Western blot technique.

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Background

Background

Saccharomyces cerevisiaereplication protein A (RPA),  also known as replication factor A (RFA) is a single-stranded DNA-binding protein that is required for multiple processes in eukaryotic DNA metabolism. Those processes include DNA replication, DNA repair, and recombination.  Homologues to RPA have been identified in all eukaryotic organisms examined. RPA is heterotrimeric protein composed of subunits of approximately 70, 30, and 14 kDa. Members of this family bind nonspecifically to single-stranded DNA and interact with and/or modify the activities of multiple proteins. Alternative names:  Replication protein A 69 kDa DNA-binding subunit, Single-stranded DNA-binding protein, DNA-binding protein BUF2, replication protein A 36 kDa subunit,    DNA-binding protein BUF1 antibody

Product citations

Selected references Deshpande et al. (2017). Structural Basis of Mec1-Ddc2-RPA Assembly and Activation on Single-Stranded DNA at Sites of Damage. Mol Cell. 2017 Oct 19;68(2):431-445.e5. doi: 10.1016/j.molcel.2017.09.019.
Chen et al. (2017). Dihydrocoumarin, an HDAC Inhibitor, Increases DNA Damage Sensitivity by Inhibiting Rad52. Int J Mol Sci. 2017 Dec 7;18(12). pii: E2655. doi: 10.3390/ijms18122655.
Yeeles et al. (2015). Regulated eukaryotic DNA replication origin firing with purified proteins. Nature. 2015 Mar 4. doi: 10.1038/nature14285.
Holstein et al. (2014). Interplay between Nonsense-Mediated mRNA Decay and DNA Damage Response Pathways Reveals that Stn1 and Ten1 Are the Key CST Telomere-Cap Components. Cell Rep. 2014 May 22;7(4):1259-69. doi: 10.1016/j.celrep.2014.04.017. Epub 2014 May 15. (ChIP)
Deng et al. (2014). RPA antagonizes microhomology-mediated repair of DNA double-strand breaks. Nat Struct Mol Biol. 2014 Mar 9. doi: 10.1038/nsmb.2786. (western blot)
Bentensen et al. (2013). MRX protects fork integrity at protein–DNA barriers, and its absence causes checkpoint activation dependent on chromatin contex. Nucleic Acids Res. 2013 Mar 1;41(5):3173-89. doi: 10.1093/nar/gkt051. (ChIP)