Nature - USA (2019-07-18)

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nature research | reporting summary

April 2018

Data

Policy information about availability of data

All manuscripts must include a data availability statement. This statement should provide the following information, where applicable:

• Accession codes, unique identifiers, or web links for publicly available datasets


• A list of figures that have associated raw data


• A description of any restrictions on data availability

All the raw data used in this study has been deposited at Gene Expression Omnibus, under the accession number GSE117826

Field-specific reporting

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Life sciences Behavioural & social sciences Ecological, evolutionary & environmental sciences

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Life sciences study design

All studies must disclose on these points even when the disclosure is negative.

Sample size A total of 29,815 cells were included from ET (n = 5) and MF (n = 4) patients. This enabled comparison of mutant and wildtype cells within

each patient sample within the context of cell identity; data from replicate patient samples confirmed the findings within each patient.

Data exclusions Based on the conventional practices of the scRNA-seq field set by the experts, we filtered out poor quality cells in 10x data analysis with

following criteria: mitochondrial RNA fraction (>10%), minimal detected number of genes (<200), maximal detected number of genes (>mean

+3*standard deviation across cells). In the amplicon data analysis, we applied specific thresholds (mismatch ratio <0.2, total number of

duplicates >=2) based our experimental data (species-mixing study). Detailed methods for processing amplicon data are fully described in the

online methods section and pipeline available on GitHub at https://github.com/thinktank-Q/IronThrone-GoT.

Replication To verify the analyzed results, we have included five CALR+ ET and four CALR+ MF patient samples. The main findings of the study were

reproduced in these replicate samples

Randomization Since the analyses are based on comparison between mutant and wildtype cells within each patient samples, randomization was not relevant

for this study.

Blinding Since the analyses are based on comparison between mutant and wildtype cells within each patient samples, blinding was not relevant for this

study.

Reporting for specific materials, systems and methods

Materials & experimental systems

n/a Involved in the study

Unique biological materials

Antibodies

Eukaryotic cell lines

Palaeontology

Animals and other organisms

Human research participants

Methods

n/a Involved in the study

ChIP-seq

Flow cytometry

MRI-based neuroimaging

Antibodies

Antibodies used CD34-PE-Vio770 (clone AC136, lot #5180718070, Miltenyi Biotec) for GoT; CD34-PeCy7 (clone 561, lot #B257238, BioLegend),

CD38-APC (clone HIT2, lot #B247250, BioLegend) and CD10-FITC (clone HI10a, lot#B254556, BioLegend) for ddPCR. The

antibodies were diluted according to manufacturers' recommendations.

Validation Negative control has been performed on every patient sample to properly set-up the gate of CD34, CD38, and CD10 positive cell