Journal: Nucleic Acids Research

Article Title: Determination of local chromatin interactions using a combined CRISPR and peroxidase APEX2 system

doi: 10.1093/nar/gkz134

Figure Lengend Snippet: Combining CRISPR system with peroxidase APEX2 to identify local chromatin interactions. ( A ) Schematic of CAPLOCUS system. ( B ) The three components of the CAPLOCUS system: TO-NLS_dCas9_NLS, the RNA binding partner fused with peroxidase APEX2_NLS, and target-specific sgRNA.

Article Snippet: To create the MS2-APEX2_NLS fusion protein expression vector, APEX2 was amplified by polymerase chain reaction (PCR) from pcDNA3 Connexin43-GFP-APEX2 (Addgene plasmid: 49385) and cloned into the pHAGE-EFS-MCP-3XBFPnls vector (Addgene plasmid: 75384) with BamHI and XhoI.

Techniques: CRISPR, RNA Binding Assay

Journal: Molecular cell

Article Title: Proximity RNA labeling by APEX-Seq Reveals the Organization of Translation Initiation Complexes and Repressive RNA Granules

doi: 10.1016/j.molcel.2019.07.030

Figure Lengend Snippet: (A) Diagram of the 43S preinitiation complex. Blue sticks signify biotin adducts formed from the proximity labeling reaction catalyzed by APEX2.

Article Snippet: Method Details DNA construction Plasmids were constructed by Gibson assembly in pCDNA5 or pET28a vectors, as indicated, using PCR amplicons generated using the primers listed below. pCDNA5-APEX2-GFP: APEX2 was amplified from pcDNA3 APEX2-NES (Addgene # 49386).

Techniques: Labeling

Journal: Molecular cell

Article Title: Proximity RNA labeling by APEX-Seq Reveals the Organization of Translation Initiation Complexes and Repressive RNA Granules

doi: 10.1016/j.molcel.2019.07.030

Figure Lengend Snippet: (A) Enrichment of translation initiation factors (orange) in APEX2-eIF4A1 proximity labeling relative to APEX2-GFP. Points in orange signify translation initiation factors. Dashed vertical line signifies proximity to APEX-eIF4A (to the right of the line are proteins in close proximity, and to the left of the line are less proximal proteins). Horizontal dashed line signifies FDR corrected significance cutoff.

Article Snippet: Method Details DNA construction Plasmids were constructed by Gibson assembly in pCDNA5 or pET28a vectors, as indicated, using PCR amplicons generated using the primers listed below. pCDNA5-APEX2-GFP: APEX2 was amplified from pcDNA3 APEX2-NES (Addgene # 49386).

Techniques: Labeling

Journal: Molecular cell

Article Title: Proximity RNA labeling by APEX-Seq Reveals the Organization of Translation Initiation Complexes and Repressive RNA Granules

doi: 10.1016/j.molcel.2019.07.030

Figure Lengend Snippet: (A) Immunofluorescence for flag tagged APEX2-eIF4A1 in unstressed cells.

Article Snippet: Method Details DNA construction Plasmids were constructed by Gibson assembly in pCDNA5 or pET28a vectors, as indicated, using PCR amplicons generated using the primers listed below. pCDNA5-APEX2-GFP: APEX2 was amplified from pcDNA3 APEX2-NES (Addgene # 49386).

Techniques: Immunofluorescence

Journal: Molecular cell

Article Title: Proximity RNA labeling by APEX-Seq Reveals the Organization of Translation Initiation Complexes and Repressive RNA Granules

doi: 10.1016/j.molcel.2019.07.030

Figure Lengend Snippet: (A) Diagram of APEX2-eIF4A1 heat shock time course.

Article Snippet: Method Details DNA construction Plasmids were constructed by Gibson assembly in pCDNA5 or pET28a vectors, as indicated, using PCR amplicons generated using the primers listed below. pCDNA5-APEX2-GFP: APEX2 was amplified from pcDNA3 APEX2-NES (Addgene # 49386).

Techniques: