Additional file 1 of CasKAS: direct profiling of genome-wide dCas9 and Cas9 specificity using ssDNA mapping
Creators: name: Marinov, Georgi K.
name: Kim, Samuel H.
name: Bagdatli, S. Tansu
name: Higashino, Soon Il
name: Trevino, Alexandro E.
name: Tycko, Josh
name: Wu, Tong
name: Bintu, Lacramioara
name: Bassik, Michael C.
name: He, Chuan
name: Kundaje, Anshul
name: Greenleaf, William J.
Publisher: name: figshare
Year: 2023
Subjects: subject: Biophysics
subject: Biochemistry
subject: Space Science
subject: Microbiology
subject: FOS: Biological sciences
subject: Genetics
subject: Molecular Biology
subject: Pharmacology
subject: Chemical Sciences not elsewhere classified
subject: Biological Sciences not elsewhere classified
subject: Cancer
subject: Infectious Diseases
subject: FOS: Health sciences
subject: Computational Biology

description: Additional file 1: Supplementary Figure 1. In vitro dCas9 and Cas9 CasKAS profiles around the mouse Nanog locus using the “Nanog-sg2” and “Nanog-sg3” sgRNAs. Supplementary Figure 2. CasKAS signal in vitro is specific to the activity of the dCas9/Cas9 protein combined with its sgRNA. Supplementary Figure 3. CasKAS signal in vitro around the VEGFA gene with the VEGFA sgRNA. Supplementary Figure 4. CasKAS signal in vivo around the VEGFA gene with the VEGFA sgRNA. Supplementary Figure 5. Time course of in vivo CasKAS signal around the EMX1 gene with the EMX1 sgRNA using dCas9. Supplementary Figure 6. Time course of in vivo CasKAS signal around the VEGFA gene with the VEGFA sgRNA using dCas9. Supplementary Figure 7. Time course of in vivo CasKAS signal around the EMX1 gene with the EMX1 sgRNA using active Cas9. Supplementary Figure 8. Time course of in vivo CasKAS signal around the VEGFA gene with the VEGFA sgRNA using active Cas9. Supplementary Figure 9. CasKAS signal in vitro around the CD2 gene with two different sgRNA targeting the gene. Supplementary Figure 10. CasKAS signal in vivo (HEK293 cells, harvested at 48 hours) around the CD2 gene with two different sgRNA targeting the gene. Supplementary Figure 11. CasKAS signal in vitro around the CD90/THY1 gene with two different sgRNA targeting the gene. Supplementary Figure 12. CasKAS signal in vivo (HEK293 cells, harvested at 48 hours) around the CD90/THY1 gene with two different sgRNA targeting the gene. Supplementary Figure 13. CasKAS signal in vitro around the CD45/PTPRC gene with two different sgRNA targeting the gene. Supplementary Figure 14. CasKAS signal in vivo (HEK293 cells, harvested at 48 hours) around the CD45/PTPRC gene with two different sgRNA targeting the gene. Supplementary Figure 15. CasKAS signal in vitro around the CD298/ATP1B3 gene with two different sgRNA targeting the gene. Supplementary Figure 16. CasKAS signal in vivo (HEK293 cells, harvested at 48 hours) around the CD298/ATP1B3 gene with two different sgRNA targeting the gene. Supplementary Figure 17. CasKAS identifies proper off-target sites that are missed by sgRNA prediction algorithms. Supplementary Figure 18. In vitro dCas9 and Cas9 CasKAS profiles for the “Nanog-sg2” sgRNA. Supplementary Figure 19. In vitro dCas9 and Cas9 CasKAS profiles for the “Nanog-sg3” sgRNA. Supplementary Figure 20. In vitro dCas9 and Cas9 CasKAS profiles for the “EMX1 Tsai” sgRNA. Supplementary Figure 21. In vitro dCas9 and Cas9 CasKAS profiles for the “VEGFA-site1” sgRNA. Supplementary Figure 22. In vitro dCas9 and Cas9 CasKAS profiles for the “CD2-1” sgRNA. Supplementary Figure 22. In vitro dCas9 and Cas9 CasKAS profiles for the “CD2-1” sgRNA. Supplementary Figure 23. In vitro dCas9 and Cas9 CasKAS profiles for the “CD2-2” sgRNA. Supplementary Figure 24. In vitro dCas9 and Cas9 CasKAS profiles for the “CD45-1” sgRNA. Supplementary Figure 25. In vitro dCas9 and Cas9 CasKAS profiles for the “CD45-2” sgRNA. Supplementary Figure 26. In vitro Cas9 CasKAS profiles for the “CD90-1” sgRNA. Supplementary Figure 27. In vitro dCas9 and Cas9 CasKAS profiles for the “CD90-2” sgRNA. Supplementary Figure 28. In vitro dCas9 and Cas9 CasKAS profiles for the “CD298-1” sgRNA. Supplementary Figure 29. In vitro dCas9 and Cas9 CasKAS profiles for the “CD298-2” sgRNA. Supplementary Figure 30. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “Nanog-sg2” sgRNA. Supplementary Figure 31. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “Nanog-sg3” sgRNA. Supplementary Figure 32. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “EMX1 Tsai” sgRNA. Supplementary Figure 33. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “VEGFA-site1” sgRNA. Supplementary Figure 34. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD2-1” sgRNA. Supplementary Figure 35. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD-2” sgRNA. Supplementary Figure 36. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD45-1” sgRNA. Supplementary Figure 37. Multiple sequence alignment of off-target sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD45-2” sgRNA. Supplementary Figure 38. Multiple sequence alignment of off-target sites identified by in vitro Cas9 CasKAS for the “CD90-1” sgRNA. Supplementary Figure 39. Multiple sequence alignment of offtarget sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD90-2” sgRNA.. Supplementary Figure 40. Multiple sequence alignment of off-target sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD298-1” sgRNA. Supplementary Figure 41. Multiple sequence alignment of off-target sites identified by in vitro dCas9 and Cas9 CasKAS for the “CD298-2” sgRNA. Supplementary Figure 42. Multiple sequence alignment of off-target sites identified by in vitro dCas9 and Cas9 CasKAS for the “sgRNA #1” sgRNA outside the list of predicted off-targets by Cass-OFFinder. Supplementary Figure 43. Cutting profiles around on- and off-target sites for the VEGFA sgRNA. Supplementary Figure 44. Cutting profiles around the top 100 on- and off-target sites for the “CD2-1” sgRNA. Supplementary Figure 45. Cutting profiles around the top 100 on- and off-target sites for the “CD2-2” sgRNA. Supplementary Figure 46. Cutting profiles around the top 100 on- and off-target sites for the “CD45-1” sgRNA. Supplementary Figure 47. Cutting profiles around the top 100 on- and off-target sites for the “CD45-2” sgRNA. Supplementary Figure 48. Cutting profiles around the top 100 on- and off-target sites for the “CD90-1” sgRNA. Supplementary Figure 49. Cutting profiles around the top 100 on- and off-target sites for the “CD90-2” sgRNA. Supplementary Figure 50. Cutting profiles around the top 100 on- and off-target sites for the “CD298-1” sgRNA. Supplementary Figure 51. Cutting profiles around the top 100 on- and off-target sites for the “CD298-2” sgRNA. Supplementary Figure 52. Amplicon sequencing of DNA edits with the EMX1 sgRNA. Supplementary Figure 53. Amplicon sequencing of DNA edits with the VEGFA sgRNA. Supplementary Figure 54. Comparing in vitro dCas9 results to using ChIP-seq and CHANGE-seq for off-target profiling. Supplementary Figure 55. Comparing in vitro dCas9 results to using DISCOVER-seq for off-target profiling. Supplementary Figure 56. Comparing in vitro dCas9 results to using DISCOVER-seq for off-target profiling. Supplementary Figure 57. Comparing in vitro dCas9 results to using GUIDE-seq for off-target profiling. Supplementary Figure 58. Comparing in vitro dCas9 results to using Digenome-seq for off-target profiling. Supplementary Figure 59. Most sgRNAs in the human genome contain multiple G nucleotides and are thus subject to labeling by N3-kethoxal. Supplementary Figure 60. Absence of strong correlation between the number of G nucleotides in a sgRNA off-target site and CasKAS signal. Supplementary Figure 61. CasKAS can be performed on pre-sheared DNA.