Description of the data and file structure

The dataset includes chromatogram trace files obtained from Sanger sequencing. The Genotyping analysis was carried out as described below:

Genomic DNA extraction was performed 3-5 days post nucleofection utilizing QuickExtract™ DNA Extraction Solution. Over 10,000 cells were collected and rinsed with 1X PBS. The cell pellet was then resuspended in approximately 20-50 µl of QuickExtract™ solution and subjected to sequential incubation at 68°C and 98°C. The extracted DNA was directly processed for amplification of the target using the appropriate primers. Following the confirmation of bands through gel electrophoresis, the amplified product was purified using the Macherey-Nagel™ NucleoSpin™ PCR Clean-up Kit. For Sanger sequencing, the purified PCR amplicon was processed with BigDye™ Terminator v3.1 Cycle Sequencing. The resulting chromatogram traces are provided in this dataset.

Files and variables

File: Chandraprabha_P_et_al.zip

Description: The zip file contains folders corresponding to figure numbers as in the article, i.e, 1A, 2B etc. Each folder contains another folder within it named Sanger trace files, in order to describe the content within each sub-folder. And every such folder contains the Sanger sequencing trace files of samples of respective figures in the article, and each sample ab1 file is labelled similar to the label given in the data figure (X-axis). Each folder also contains a ‘Control’ ab1 file, which can be used as the unedited reference for analysing the results. Detailed description of each folder is as follows, described in the order as found within the zip file:

Figure 2F -

Subfolders

1_HSPCs: Control and sample (HSPCs) ab1 files for HSPCs plot on Figure 2F
2_Colonies: Control and CFU colonies sample (C1-C26) ab1 files for Colonies plot on Figure 2F

Figure 2I -

Control and sample (Labelled similar to figure 2I labels described in coloured dots on left) ab1 files for plot Figure 2I; 175, 175 158, B4, 158.

Each duplicate per sample is numbered 1, 2 etc.

 Figure 3B –

Control (1 & 2) and sample (Labelled similar to figure 3B labels described in X-axis) ab1 files for plot Figure 3B; DMSO, RS-1, RUS, Alt-R, Q-VD-OPH, Cyclosporin-H, Eltrombopag, SB-203580, Ursolic acid, Zeatin, Zeatin riboside, XL-413, Aphidicolin, Nicotinamide, Hydroxyurea, SCR-7, NU-7441, Trichostatin-A, Valproic acid, Entinostat, Panobinostat, Nutlin-3a, Pfithrin-µ, Pfithrin-α, UNC-2170.

Each duplicate per sample is numbered 1, 2 etc.

 Figure 3C –

Subfolders - Experiment 1 & Experiment 2 (Experimental duplicates)

Both folders contain Control and sample (Labelled similar to figure 3C labels described in X-axis) ab1 files for plot Figure 3C; DMSO, NU-7441, AZD-7648, DMNB, LTURM 34, NU-7026.

Each duplicate per sample is numbered 1, 2 etc.

 Figure 3E –

Subfolders - Experiment 1 & Experiment 2 (Experimental duplicates)

Both folders contain Control and sample (Labelled similar to figure 3E labels described in X-axis) ab1 files for plot Figure 3E; 0 µM, 7 µM, and 20 µM for both Vehicle and RUS samples.

Each duplicate per sample is numbered 1, 2 etc.

Figure 4F –

Subfolders –

Input (Refers to input genotype of infused cells) &

In-vivo (Refers to genotype of transplanted cells at 16th week)  

Both folders contain respective Control and sample (Labelled similar to figure 4F labels described in X-axis) ab1 files for plot Figure 4F;

Input; B4, B4-AZD-7648

In-vivo; B4, B4-AZD-7648 for mouse numbers 1-3 and 2-4, respectively.

Figure 5B& 5C –

Folder contains Control and sample (Labelled similar to figure 5B labels described in X-axis) ab1 files for plot Figure 5B; Control and β-thal model.

Figure 5D –

Folder contains respective Control (Labelled as Wild type) and sample (Labelled similar to figure 5D labels described in X-axis) ab1 files for plot Figure 5D; Wild type, β-thal model_Control, β-thal model_158 C to T, β-thal model_B4

Figure 6B –

Subfolders - Experiment 1 & Experiment 2 (Experimental duplicates)

Both folders contain Control and sample (Labelled similar to figure 6B labels described in X-axis) ab1 files for plot Figure 6B; SC, BM-MSC, WJ-MSC, SC+AZD-7648, BM-MSC+AZD-7648, WJ-MSC+AZD-7648

Each duplicated sample is numbered 1, 2,3 etc.

Figure 7A –

Subfolders (different targets) –

-115 HBG promoter

-175 & -158

BCL11A enhancer

All three folders contain Control and sample (labelled as per sample details in figure 7A and the folder name) ab1 files for plot 7A; 115HBG, 175158, BCL11Aenh. Respective controls are provided separately in each folder.

Each duplicated sample is numbered 1, 2,3 etc.

Supplementary Figure 1B –

Control and sample (labelled as per sample details in figure supplementary 1B) ab1 files for plot supplementary 1B; B2, B4. 3 controls are provided corresponding to multiple donor cells used in duplicates 1,2,3

Each duplicated sample is numbered 1, 2,3 etc.

Supplementary Figure 2C –

Control and sample (labelled as per sample details in supplementary figure 2C) ab1 files for plot supplementary 2C; RNP, DMSO, RUS. 3 controls are provided corresponding to multiple donor cells used in duplicates 1,2,3

Each duplicated sample is numbered 1, 2,3 etc.

Supplementary Figure 2D –

Subfolders (different targets) –

Donor_1

Donor_2

Donor_3

All three folders contain Control and sample (labelled as per sample details in supplementary figure 2D) ab1 files for plot supplementary 2D; CD34+ cells, CD34+CD90+ cells, CD34+CD90- cells (for donor 2 CD34+CD90- cells didn’t generate quality reads for analysis, so excluded). Respective controls are provided separately in each folder.

Each duplicated sample is numbered 1, 2,3 etc.

Supplementary Figure 2F –

Control and sample (labelled as per sample details in supplementary figure 2F) ab1 files for plot supplementary 2F;

3 Controls

(T1-T7: Genotype at different hours as in figure)
T1- 0.5 hours
T2- 1 hour
T3- 2 hours
T4- 3 hours
T5- 6 hours
T6- 12 hours
T7- 48 hours

Supplementary Figure 2G –

Control and sample (labelled as per sample details in supplementary figure 2G) ab1 files for plot supplementary 2G;

Control

(T1-T8: Genotype at different hours as in figure)

T1- 0.5 hours
T2- 1 hour
T3- 2 hours
T4- 3 hours
T5- 6 hours
T6- 12 hours
T7- 24 hours
T7- 48 hours

Supplementary Figure 3B –

Control and sample (labelled as per sample details in figure supplementary 3B) ab1 files for plot supplementary 3B; DMSO, 6 hours, 12 hours, 18 hours, 24 hours

Each duplicated sample is numbered 1, 2 etc.

Code/software

·         ab1 trace files can be viewed using Snapgene sequence viewer

·         Editing efficiency for samples was analysed using Synthego ICE analysis : Inference of CRISPR edits (https://ice.synthego.com/#/)

·         Base editing efficiency was analysed using EditR software (http://baseeditr.com/)

·         Graphs were plotted using GraphPad Prism v.10.2

·         Statistical analyses were performed using GraphPad Prism v.10.2