WormBase Tree Display for Antibody: WBAntibody00000657
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| WBAntibody00000657 | Summary | Polyclonal rabbit antibody against recombinant protein. | ||
|---|---|---|---|---|
| Public_name | [cgc6010]::anti-RAD-51 | |||
| Gene | WBGene00004297 | |||
| Isolation | Original_publication | WBPaper00006010 | ||
| Location | TG | |||
| Clonality | Polyclonal | |||
| Antigen | Protein | A polymerase chain reaction (PCR) fragment encoding the N-terminal 103 amino acids of C. elegans RAD-51 was cloned in frame as a glutathione S-transferase (GST) fusion into pGEX-4T-1 (Amersham) and as a maltose binding protein (MBP) fusion into pMal-c2 (New England BioLabs). Recombinant GST and MPB fusions were expressed in Escherichia coli BL21 CodonPlus (Stratagene), purified under native conditions by affinity chromatography using Glutathione-Sepharose 4B (Amersham) and by amylose-resin affinity chromatography (New England BioLabs). The purified GST protein fusion was dialyzed against phosphate-buffered saline (PBS) and used for immunization of rabbits (Eurogentech). For anti-RAD-51 antibody affinity purification MBP-RAD-51 fusion protein was used. The MBP fusion was dialyzed against 1PBS and 2 mg of total protein was covalently bound to an Affi-Gel 15 matrix (BioRad). Crude serum was diluted 1:1 in 20 mM TRIS, pH 7.5 and pre-cleaned by centrifugation and applied to the column to allow for the binding of the antibody. After washing with 10 mM TRIS-HCl, pH 7.5 and 10 mM TRIS-HCl, pH 7.5, 0.5 mM NaCl, antibodies were eluted with 100 mM glycine, pH 2.5 and the eluates were fractionated and immediately neutralized with 1.5 M TRIS, pH 8.0. | ||
| Animal | Rabbit | |||
| Expr_pattern | Expr2647 | |||
| Reference (17) |
