PRIM1 Polyclonal ANTIBODY Catalog Number:10773-1-AP 2 Publications

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PRIM1 Polyclonal ANTIBODY Catalog Number:10773-1-AP 2 Publications

For Research Use Only PRIM1 Polyclonal ANTIBODY www.ptglab.com Catalog Number:10773-1-AP 2 Publications Catalog Number: GenBank Accession Number: Purification Method: Basic Information 10773-1-AP BC005266 Antigen affinity purification Size: GeneID (NCBI): Recommended Dilutions: 150UL , Concentration: 133 μg/ml by 5557 WB 1:500-1:1000 Bradford method using BSA as the Full Name: IP 0.5-4.0 ug for IP and 1:200-1:1000 standard; primase, DNA, polypeptide 1 (49kDa) for WB Source: IHC 1:20-1:200 Calculated MW: IF 1:50-1:500 Rabbit 50 kDa Isotype: Observed MW: IgG 50 kDa Immunogen Catalog Number: AG1124 Applications Tested Applications: Positive Controls: IF, IHC, IP, WB, ELISA WB : K-562 cells, HeLa cells, Sp2/0 cells Cited Applications: IP : HeLa cells, IHC IHC : human lymphoma tissue, Species Specificity: human, mouse, rat IF : HeLa cells, Cited Species: mouse Note-IHC: suggested antigen retrieval with TE buffer pH 9.0; (*) Alternatively, antigen retrieval may be performed with citrate buffer pH 6.0 DNA replication in human cells is initiated by a complex apparatus containing a DNA polymerase-alpha/primase Background Information complex. Primase synthesizes oligoribonucleotides that serve as primers for the initiation of DNA synthesis. It plays a role in both the initiation of DNA replication and the synthesis of Okazaki fragments for lagging strand synthesis. PRIM1 is a subnuit of this complex. Notable Publications Author Pubmed ID Journal Application Pierre Kunz 30986223 PLoS One IHC Liam C Hunt 31365869 Cell Rep Storage: Storage Store at -20°C. Stable for one year after shipment. Storage Buffer: PBS with 0.1% sodium azide and 50% glycerol pH 7.3. Aliquoting is unnecessary for -20ºC storage For technical support and original validation data for this product please contact: This product is exclusively available under Proteintech T: 1 (888) 4PTGLAB (1-888-478-4522) (toll free E: [email protected] Group brand and is not available to purchase from any in USA), or 1(312) 455-8498 (outside USA) W: ptglab.com other manufacturer. Selected Validation Data K-562 cells were subjected to SDS PAGE followed Immunohistochemical analysis of paraffin- IP Result of anti-PRIM1 (IP:10773-1-AP, 4ug; by western blot with 10773-1-AP (PRIM1 antibody) embedded human lymphoma using 10773-1-AP Detection:10773-1-AP 1:300) with HeLa cells lysate at dilution of 1:500 incubated at room temperature (PRIM1 antibody) at dilution of 1:100 (under 10x 2800ug. for 1.5 hours. lens). Immunofluorescent analysis of (4% PFA) fixed HeLa cells using 10773-1-AP (PRIM1 antibody) at dilution of 1:50 and Alexa Fluor 488-conjugated AffiniPure Goat Anti-Rabbit IgG(H+L)..

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Supplementary Text and Figures.Pdf
Supplemental Figure S2. PRIM1 immunoblots (20-75 kDa). (A) Corresponds to Figure 2D, with additional bands seen in P2 at 75 kDa and 25 kDa. (B) Independent experiment with cell lysates from C and P2 in which additional bands at 25 kDa not evident. (C) Corresponds to Figure 2D. No additional band visible at 25 kDa. (D) Validation of monoclonal antibody (8G10) with siRNA to PRIM1 in HeLa cells. siLUC, luciferase negative control. siPRIM1, siRNA targeting PRIM1 transcript. Vinculin used as a loading control. Supplemental Figure S3: Predicted destabilisation of PRIM1 by the C301R substitution. (A) Cysteine 301 lies in a buried hydrophobic region in PRIM1. DNA primase dimer crystal structure (PDB: 4BPU). PRIM1 residues shaded according to solvent accessibility. Cysteine 301, substituted to arginine in P5, lies in a buried hydrophobic region. (B) Cys301 is evolutionarily conserved in vertebrates. Neither arginine or other large or charged amino acids are observed at this position in other species. (C) The C301R substitution observed in P5 is predicted to lead to destabilisation of all available PRIM1 crystal structures. In contrast, substitution with leucine or threonine, as observed in some orthologous proteins, is not predicted to lead to destabilisation. Data points, predicted changes in free-energy (ΔΔG) from FoldX plotted for each of 14 available crystal structures. (D) Immunoblotting of RPE1 cells transfected with dual reporter constructs (as described in Figure 3B and Materials and Methods) confirms production of PRIM1-GFP and FLAG-SR at expected molecular weights and shows reduced protein levels for the C301R variant. n=2 experiments shown. C301R protein levels relative to WT after normalization to actin loading control indicated underneath the blot.
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