KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
| Bacterial and Virus Strains | ||
| Agrobacterium tumefaciens strains EHA105 and GV3101 | Widely distributed | EHA105, GV3101 |
| Biological Samples | ||
| SWC wt × SWC rto-like M. guttatus F2 seed | This Paper | |
| LRD wt × LRD rto-like M. guttatus F2 seed | This Paper | |
| rto × SL9 M. lewisii F2 seed | This Paper | |
| Chemicals, Peptides, and Recombinant Proteins | ||
| 1-napthalene acetic acid | PhytoTechnology Labs | Cat#N5999 |
| 2-N-morpholino ethane sulfonic acid | Sigma-Aldrich | Cat#M2933 |
| Acetosyringone (4′-Hydroxy-3′,5′-dimethoxyacetophenone) |
Sigma-Aldrich | Cat#D134406 |
| Calcium-D-gluconate | Acros Organics | Cat#299-28-5 |
| Cefotaxime | GoldBio | Cat#C-104–25 |
| Cetyl Trimethyl Ammonium Bromide (CTAB) | Sigma-Aldrich | Cat#H5882 |
| Gelrite (Gellan gum) | Scotts Labs | Cat#4900–1890 |
| Kanamycin | GoldBio | Cat#K-120–10 |
| Meta-topolin | PhytoTechnology Labs | Cat#T-841 |
| Murashige and Skoog salts with Gamborg B5 vitamins | PhytoTechnology Labs | Cat#M404 |
| N-(2-Chloro-4-pyridyl)-N′-phenylurea (CPPU) | PhytoTechnologyLabs | Cat#C279 |
| Phosphinothricin | GoldBio | Cat#P-165–250 |
| Rifampicin | GoldBio | Cat#R-120–1 |
| Timentin™ Ticarcillin / Clavulanate | GoldBio | Cat#T-104–250 |
| S. pyogenes Cas9 nuclease | New England Biolabs | Cat#M0386S |
| Critical Commercial Assays | ||
| AMPure XP beads | Beckman Coulter | Cat#A63880 |
| Quant-iT dsDNA Broad Range Assay Kit | ThermoFisher | Cat#Q33130 |
| NEBNext Ultra II DNA Library Prep Kit for Illumina | New England Biolabs | Cat#E7645S |
| Multiplex Oligos for Illumina Kit | New England Biolabs | Cat#E7335S |
| Phusion High Fidelity PCR Kit | New England Biolabs | Cat#E0553S |
| Spectrum Plant Total RNA Kit | Sigma-Aldrich | Cat#STRN50–1KT |
| Zymo Direct-Zol RNA kit | Zymo Research | Cat#R2055 |
| SuperScript III First-Strand Synthesis System | Invitrogen | Cat#18080051 |
| DNase I | Invitrogen | Cat#18068015 |
| DNase I Set | Zymo Research | Cat#E1010 |
| GoScript Reverse Transcriptase System | Promega | Cat#A5000 |
| PowerUp SYBR Green Master Mix | Applied Biosystems | Cat#A25741 |
| iQ SYBR Green Supermix | Bio-Rad | Cat#1708880 |
| Deposited Data | ||
| Genomic sequence reads generated for bulked segregant analyses | NCBI Sequence Read Archive | SRA: BioProject PRJNA481753 |
| Mimulus guttatus v2.0 reference genome | Phytozome | https://phytozome.jgi.doe.gov/pz/portal.html#!info?alias=Org_Mguttatus |
| Experimental Models: Organisms/Strains | ||
| Mimulus lewisii | Inbred line LF10 | LF10 |
| Mimulus guttatus | Sweet Creek Road (OR), Littlerock Dam (CA), Trinity Alps Road (CA), Quarry Valley Road (CA), Mackville Road (CA) | SWC, LRD, TAR, QVR, MAC |
| Nicotiana benthamiana | Widely distributed | N/A |
| Bombus impatiens | Koppert Biological Systems | Naturpol |
| Oligonucleotides | ||
| Primers for cloning, sequencing, genotyping, qPCR | This paper (Tables S4–S6) | N/A |
| Recombinant DNA | ||
| pFGC5941 | Arabidopsis Biological Resource Center | CD3–447 |
| pMCS:GW | Arabidopsis Biological Resource Center | CD3–1933 |
| ER-cb | Arabidopsis Biological Resource Center | CD3–954 |
| pMCS:YFP-GW | Arabidopsis Biological Resource Center | CD3–1934 |
| pEarleyGate 100 | Arabidopsis Biological Resource Center | CD3–724 |
| pEarleyGate 104 | Arabidopsis Biological Resource Center | CD3–686 |
| pUC119-gRNA | Addgene | 52255 |
| pFGC-pcoCas9 | Addgene | 52256 |
| pUBC-nYFP-Dest | [71] | N/A |
| pUBC-cYFP-Dest | [71] | N/A |
| pENTR/D-TOPO | Invitrogen | Cat#K240020 |
| AS-121P-CR3 | Alex Schultink, Innovative Genomics Institute | N/A |
| MlRTO RNAi | This paper | N/A |
| MgRTO RNAi | This paper | N/A |
| MlNEGAN RNAi | This paper | N/A |
| MgNEGAN RNAi | This paper | N/A |
| RTO sgRNA+ Cas9 | This paper | N/A |
| RTOpro:CFP-ER | This paper | N/A |
| RTOpro:YFP-RTO | This paper | N/A |
| 35S:MlRTO | This paper | N/A |
| 35S:YFP-MlRTO | This paper | N/A |
| pUBC-cYFP-ANbHLH1 | This paper | N/A |
| pUBC-nYFP-RTO | This paper | N/A |
| pUBC-nYFP-rto | This paper | N/A |
| Software and Algorithms | ||
| bwa | [75] | https://github.com/lh3/bwa |
| samtools | [76] | https://github.com/samtools |
| ImageJ | [77] | https://imagej.nih.gov/ij/ |
| VCell | [48] | http://vcell.org |
| RDsimJ.jar | [12] | Part of the Supporting Online Material |
| JMP Pro 13 | SAS Institute Inc. | https://www.jmp.com/ |
| E-CRISP | [78] | http://www.e-crisp.org/E-CRISP/ |
| Custom Scripts | This Paper | https://github.com/BlackmanLabUCB/Bulked-Segregant-Analysis |