Key Resource Table.
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Anti-GFP | Protein Tech | 50430-2-AP, RRID:AB_11042881 |
| Anti-Rabbit HRP | Thermo | 31460, RRID:AB_2532095 |
| Anti-Rpb1 Ser2 | Abcam | Ab5095 RRID:AB_304749 |
| Anti-Drosophila Rpb3 | Lab stock (Andrulis et al 2000) 65 | N/A |
| Goat Anti-Rabbit 800 | LiCor | 926-32211 |
| Bacterial and virus strains | ||
| DH5 Alpha Supercompetent Cells | NEB | C2987 |
| Biological samples | ||
| Chemicals, peptides, and recombinant proteins | ||
| Grace’s Insect Medium | Sigma | 45-G8142-500ML |
| Janelia Fluor 650x | Gift from Luke Lavis Janelia Research Farms | Available Now from Promega CS315104 |
| Janelia Fluor 549 | Gift from Luke Lavis Janelia Research Farms | Available Now from Promega GA1110 |
| BbsI Restriction Enzyme | NEB | R0539L |
| T4 DNA Ligase | NEB | M0202L |
| HiFi Assembly Master Mix | NEB | E2621L |
| RNase Cocktail | Thermo | AM2286 |
| Proteinase K | Thermo | AM2548 |
| Protease Inhibitor Cocktail | Thermo | A32963 |
| MNase | NEB | M0247S |
| Sybr Green | Thermo | S7567 |
| Phusion HF Buffer | NEB | B0518S |
| PVDF Membrane | Millipore | 0000167543 |
| Enhanced Chemiluminescence (ECL) | Bio-Rad | 170-5060 |
| CL-XPosure Film | Thermo | 34090 |
| NVP-2 | MedChemExpress | HY-122114A |
| THZ531 | Caymen Chemical | 26386-1 |
| Critical commercial assays | ||
| ENZA Endo-Free Plasmid Midi Kit | Omega | D6915-03 |
| ENZA Plasmid Mini Kit | Omega | D6942-02 |
| Deposited data | ||
| Imaging Data used for figures, Western Blots, and sequences for CRISPR HDR plasmids | This Paper | Mendeley Data: https://data.mendeley.com/preview/4rxj5g76z8?a=6e697080-4aff-4e3e-abeb-c8485503459c DOI: 10.17632/4rxj5g76z8.1 |
| Imaging Data Fig 1: Pol II and DSIF | This Paper |
https://data.mendeley.com/preview/4vhdtshgx7?a=6179b8da-ea08-4e9c-a298-f4f9ed32be73 DOI: 10.17632/4vhdtshgx7.1 |
| Imaging Data Fig 1: P-TEFb and PAF1 | This Paper |
https://data.mendeley.com/preview/sc442hnfj5?a=67646232-428c-4150-b4dc-64a0d36a9be3 DOI: 10.17632/sc442hnfj5.1 |
| Imaging Data Fig 1: Spt6, excel analysis and Prism Graph Pad Plots and fits | This Paper |
https://data.mendeley.com/preview/2xfxmb9zkh?a=c57fbc98-d5ad-4a81-a717-e6f2566579b0 DOI: 10.17632/2xfxmb9zkh.1 |
| Imaging Data Supplemental Figure 2 Raw Data | This Paper |
https://data.mendeley.com/preview/crw4c8nb3y?a=b1654f18-50e8-4dbc-9410-07b3a3b0ee84 DOI: 10.17632/crw4c8nb3y.1 |
| Imaging Data Supplemental Figure 3 Dataset | This Paper |
https://data.mendeley.com/preview/tbb6r7cfzc?a=7e030c1d-c7d9-47e5-94d9-3852093237fd DOI: 10.17632/tbb6r7cfzc.1 |
| Imaging Data Figure 2 and Supplemental Figure 4 Raw Data, excel analysis, Prism graphs and fits | This Paper |
https://data.mendeley.com/preview/gy58myshy9?a=ceea7834-75db-4d62-bd9b-74d236d7e7a7 DOI: 10.17632/gy58myshy9.1 |
| Imaging Data Figure 3 and Supplemental Figure 5 Raw Data, excel analysis and Prism plots and T-test | This Paper |
https://data.mendeley.com/preview/yvbcw7m3dp?a=2c97b57c-e002-4ec3-a13f-85b69d703cc1 DOI: 10.17632/yvbcw7m3dp.1 |
| Imaging Data Fig 4. Pol II, PAF1, Ctr9, Spt6 | This Paper |
https://data.mendeley.com/preview/9t8wddtwsn?a=5e9628f3-5b8d-4a73-a3aa-1ce1ca9b1eda DOI: 10.17632/9t8wddtwsn.1 |
| Imaging Data Figure 4 NELF-A and Supplemental Figure 6 Raw data, excel analysis and Prism plots and T-tests | This Paper |
https://data.mendeley.com/preview/45j7d538cj?a=d9fbb3c7-61c3-4647-ac0a-606070b112c9 DOI: 10.17632/45j7d538cj.1 |
| Imaging Data Figure 5 Cdc73 RNAi Pol II PA-GFP decay. | This Paper |
https://data.mendeley.com/preview/ypyy9k5bfj?a=86fe214f-a54c-411d-9171-be13dbe982ce DOI: 10.17632/ypyy9k5bfj.1 |
| Imaging Data Figure 5. Luciferase RNAi data Pol II photoactivation and decay | This Paper |
https://data.mendeley.com/preview/dy7kjr8jrt?a=44e6db3f-cdfc-4c56-a25c-def729fdeed7 DOI: 0.17632/dy7kjr8jrt.1 |
| Imaging Data Figure 5 MS2 Raw data both RNAi | This Paper |
https://data.mendeley.com/preview/wp82jngz8s?a=d093e582-03d9-445f-9960-0f7fbce4d149 DOI: 10.17632/wp82jngz8s.1 |
| Imaging Data. Supplemental Figure 7C Raw Data | This Paper |
https://data.mendeley.com/preview/jpwfvsv6x4?a=b508bfdb-3afe-4b7b-b57a-0e6af56eeb27 DOI: 10.17632/jpwfvsv6x4.1 |
| Supplemental Figure 7H Raw Data | This Paper |
https://data.mendeley.com/preview/8dkzfjkxr2?a=3db6bb5d-22dc-44cd-a719-17fbc2f78beb DOI: 10.17632/8dkzfjkxr2.1 |
| Experimental models: Cell lines | ||
| Experimental models: Organisms/strains | ||
| Fly: y1w1118 | Bloomington Stock Center | #6598 |
| Fly: PiggyBac Transposase | Bloomington Stock Center | #8285 |
| Fly: FM7A Balancer | Bloomington Stock Center | #785 |
| Fly: Tm3,Sb/Tm6b Tb Balancer | This Paper | N/A |
| Fly: Cyo/Xasta ; Tm6b,Tb/Xasta | Gift from Dan Barbash | N/A |
| Fly: Luciferase RNAi | Bloomington Stock Center | 31603 |
| Fly: Spt6 RNAi | Bloomington Stock Center | 32373 |
| Fly: Cdc73/Hyx RNAi | Bloomington Stock Center | 31722 |
| Fly: Rpb9-Halo | This Paper | N/A |
| Fly: Rpb9-PA-GFP | This Paper | N/A |
| Fly: Rpb9-PA-TagRFP | This Paper | N/A |
| Fly: Halo-NELF-A | This Paper | N/A |
| Fly: Spt5-PA-GFP | This Paper | N/A |
| Fly: Cyclin-T-PA-GFP | This Paper | N/A |
| Fly: PAF1-Halo | This Paper | N/A |
| Fly: PAF1-PA-GFP | This Paper | N/A |
| Fly: Ctr9-Halo | This Paper | N/A |
| Fly: Ctr9-PA-GFP | This Paper | N/A |
| Fly: Spt6-PA-GFP | This Paper | N/A |
| Fly: HSF-mNeonGreen | This Paper | N/A |
| Fly: HSF-Halo | This Paper | N/A |
| Fly: Gal4 | Bloomington Stock Center | 6979 |
| Fly: Gal4 | Bloomington Stock Center | 6983 |
| Fly: Hsp70Transgene, LacI-mCherry ; hsp83-MCP-GFP | Buckley et al. 2014 25 | N/A |
| Oligonucleotides | ||
| See Table S2 | This Paper | N/A |
| Recombinant DNA | ||
| pCFD3-dU6 3gRNA | DGRC | RRID:DGRC_1435 1435 |
| pHD-Scarless-dsRed | DGRC | RRID:DGRC_1364 1364 |
| EasyFusion Halo | Addgene | 112850 |
| pTW-Rpb9-paG | Buckley et al. 2014 25 | N//A |
| pTG603 HSF-mNeonGreen homology donor | This Paper | N/A |
| pTG607 HSF-Halo HDR template | This Paper | N/A |
| pTG610 HSF-sgRNA | This Paper | N/A |
| Rpb9-Halo Homology Donor | This Paper | N/A |
| Rpb9-PA-GFP Homology Donor | This Paper | N/A |
| Rpb9-PA-TagRFP Homology Donor | This Paper | N/A |
| Rpb9 sgRNA | This Paper | N/A |
| Halo-NELF-A Homology Donor | This Paper | N/A |
| NELF-A sgRNA | This Paper | N/A |
| Spt5-PA-GFP Homology Donor | This Paper | N/A |
| Spt5 sgRNA | This Paper | N/A |
| Cyclin-T-PA-GFP Homology Donor | This Paper | N/A |
| Cyclin-T sgRNA | This Paper | N/A |
| PAF1-Halo Homology Donor | This Paper | N/A |
| PAF1-PA-GFP Homology Donor | This Paper | N/A |
| PAF1 sgRNA | This Paper | N/A |
| Ctr9-Halo Homology Donor | This Paper | N/A |
| Ctr9-PA-GFP Homology Donor | This Paper | N/A |
| Ctr9 sgRNA | This Paper | N/A |
| Spt6-PA-GFP Homology Donor | This Paper | N/A |
| Software and algorithms | ||
| FIJI Version 2.14.0 | Schindelin et al. 2012 | https://imagej.net/software/fiji/ |
| PRISM Version 10.1.1 | https://www.graphpad.com/features | N/A |
| Microsoft Excel 16.16.27 | https://www.microsoft.com/en-us/microsoft-365/p/excel-home-and-student/CFQ7TTC0HLKR?activetab=pivot:overviewtab | N/A |
| flyCRISPR Target Finder | Gratz, S.J.*, Ukken, F.P.*, et al. (2014) | http://targetfinder.flycrispr.neuro.brown.edu/ |
| Other | ||
| 35 mm PolyD-Lysine Coated No 1.5 Imaging Dishes | MatTek | P35GC-1.5-14-C |