{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,25]],"date-time":"2026-08-25T02:31:41Z","timestamp":1787625101242,"version":"build-2736575974"},"reference-count":50,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2025,4,29]],"date-time":"2025-04-29T00:00:00Z","timestamp":1745884800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003975","name":"Israel Cancer Association","doi-asserted-by":"publisher","award":["20150081"],"award-info":[{"award-number":["20150081"]}],"id":[{"id":"10.13039\/501100003975","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003975","name":"Israel Cancer Association","doi-asserted-by":"publisher","award":["670535"],"award-info":[{"award-number":["670535"]}],"id":[{"id":"10.13039\/501100003975","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003975","name":"Israel Cancer Association","doi-asserted-by":"publisher","award":["874606"],"award-info":[{"award-number":["874606"]}],"id":[{"id":"10.13039\/501100003975","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100001698","name":"Israel Cancer Research Foundation","doi-asserted-by":"publisher","award":["20150081"],"award-info":[{"award-number":["20150081"]}],"id":[{"id":"10.13039\/100001698","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100001698","name":"Israel Cancer Research Foundation","doi-asserted-by":"publisher","award":["670535"],"award-info":[{"award-number":["670535"]}],"id":[{"id":"10.13039\/100001698","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100001698","name":"Israel Cancer Research Foundation","doi-asserted-by":"publisher","award":["874606"],"award-info":[{"award-number":["874606"]}],"id":[{"id":"10.13039\/100001698","id-type":"DOI","asserted-by":"publisher"}]},{"name":"EU-H2020-Health","award":["20150081"],"award-info":[{"award-number":["20150081"]}]},{"name":"EU-H2020-Health","award":["670535"],"award-info":[{"award-number":["670535"]}]},{"name":"EU-H2020-Health","award":["874606"],"award-info":[{"award-number":["874606"]}]},{"name":"Kenneth and Sally Leafman Appelbaum Discovery Fund","award":["20150081"],"award-info":[{"award-number":["20150081"]}]},{"name":"Kenneth and Sally Leafman Appelbaum Discovery Fund","award":["670535"],"award-info":[{"award-number":["670535"]}]},{"name":"Kenneth and Sally Leafman Appelbaum Discovery Fund","award":["874606"],"award-info":[{"award-number":["874606"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Acute myeloid leukemia (AML) is associated with unfavorable patient outcomes primarily related to disease relapse. Since specific types of leukemic hematopoietic stem and progenitor cells (HSPCs) are suggested to contribute to AML propagation, this study aimed to identify and explore relapse-initiating HSPC subpopulations present at diagnosis, using single-cell analysis (SCA). We developed unique high-resolution techniques capable of tracking single-HSPC-derived subclones during AML evolution. Each subclone was evaluated for chemo-resistance, in vivo leukemogenic potential, mutational profile, and the cell of origin. In BM samples of 15 AML patients, GMP-like and MLP-like HSPC subpopulations were identified as prevalent at relapse, exhibiting chemo-resistance to commonly used chemotherapy agents cytosine arabinoside (Ara-C) and daunorubicin. Reconstruction of phylogenetic lineage trees combined with genetic analysis of single HSPCs and single-HSPC-derived subclones demonstrated two distinct clusters, originating from MLP-like or GMP-like subpopulations, observed both at diagnosis and relapse. These subpopulations induced leukemia development ex vivo and in vivo. Genetic SCA showed that these relapse-related subpopulations harbored mutated EZH2 and TP53, detected already at diagnosis. This study, using combined molecular, functional, and genomic analyses at the level of single cells, identified patient-specific chemo-resistant HSPC subpopulations at the time of diagnosis, promoting AML relapse.<\/jats:p>","DOI":"10.3390\/ijms26094224","type":"journal-article","created":{"date-parts":[[2025,4,29]],"date-time":"2025-04-29T07:49:36Z","timestamp":1745912976000},"page":"4224","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["GMP-like and MLP-like Subpopulations of Hematopoietic Stem and Progenitor Cells Harboring Mutated EZH2 and TP53 at Diagnosis Promote Acute Myeloid Leukemia Relapse: Data of Combined Molecular, Functional, and Genomic Single-Stem-Cell Analyses"],"prefix":"10.3390","volume":"26","author":[{"given":"Tal","family":"Shahar Gabay","sequence":"first","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"},{"name":"The Ruth and Bruce Rappaport Faculty of Medicine, Technion, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nofar","family":"Stolero","sequence":"additional","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-1053-8199","authenticated-orcid":false,"given":"Niv","family":"Rabhun","sequence":"additional","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rawan","family":"Sabah","sequence":"additional","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"},{"name":"The Ruth and Bruce Rappaport Faculty of Medicine, Technion, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ofir","family":"Raz","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8221-4050","authenticated-orcid":false,"given":"Yaara","family":"Neumeier","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"},{"name":"Department of Immunology, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zipora","family":"Marx","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liming","family":"Tao","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tamir","family":"Biezuner","sequence":"additional","affiliation":[{"name":"Department of Immunology, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shiran","family":"Amir","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rivka","family":"Adar","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ron","family":"Levy","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noa","family":"Chapal-Ilani","sequence":"additional","affiliation":[{"name":"Department of Immunology, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4950-3691","authenticated-orcid":false,"given":"Natalia","family":"Evtiugina","sequence":"additional","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liran I.","family":"Shlush","sequence":"additional","affiliation":[{"name":"Department of Immunology, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ehud","family":"Shapiro","sequence":"additional","affiliation":[{"name":"Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 761001, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0608-8130","authenticated-orcid":false,"given":"Shlomit","family":"Yehudai-Resheff","sequence":"additional","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"},{"name":"The Ruth and Bruce Rappaport Faculty of Medicine, Technion, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6204-977X","authenticated-orcid":false,"given":"Tsila","family":"Zuckerman","sequence":"additional","affiliation":[{"name":"Hematology Research Center, Clinical Research Institute at Rambam, Rambam Health Care Campus, Haifa 3109601, Israel"},{"name":"The Ruth and Bruce Rappaport Faculty of Medicine, Technion, Haifa 3109601, Israel"},{"name":"Department of Hematology and Bone Marrow Transplantation, Rambam Health Care Campus, Haifa 3109601, Israel"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,4,29]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"2209","DOI":"10.1056\/NEJMoa1516192","article-title":"Genomic Classification and Prognosis in Acute Myeloid Leukemia","volume":"374","author":"Papaemmanuil","year":"2016","journal-title":"N. Engl. J. Med."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"371","DOI":"10.1182\/blood.2019002697","article-title":"Impact of NPM1\/FLT3-ITD genotypes defined by the 2017 European LeukemiaNet in patients with acute myeloid leukemia","volume":"135","author":"Dohner","year":"2020","journal-title":"Blood"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1038\/s41586-021-04206-7","article-title":"Non-genetic determinants of malignant clonal fitness at single-cell resolution","volume":"601","author":"Fennell","year":"2022","journal-title":"Nature"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"5327","DOI":"10.1038\/s41467-020-19119-8","article-title":"Clonal evolution of acute myeloid leukemia revealed by high-throughput single-cell genomics","volume":"11","author":"Morita","year":"2020","journal-title":"Nat. Commun."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"934","DOI":"10.1200\/JCO.2016.71.2208","article-title":"Genomics of Acute Myeloid Leukemia Diagnosis and Pathways","volume":"35","author":"Bullinger","year":"2017","journal-title":"J. Clin. Oncol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"264","DOI":"10.1016\/j.cell.2012.06.023","article-title":"The origin and evolution of mutations in acute myeloid leukemia","volume":"150","author":"Welch","year":"2012","journal-title":"Cell"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1016\/j.ccr.2014.01.031","article-title":"Functional heterogeneity of genetically defined subclones in acute myeloid leukemia","volume":"25","author":"Klco","year":"2014","journal-title":"Cancer Cell"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1501","DOI":"10.1038\/s41375-024-02211-z","article-title":"Mutation order in acute myeloid leukemia identifies uncommon patterns of evolution and illuminates phenotypic heterogeneity","volume":"38","author":"Schwede","year":"2024","journal-title":"Leukemia"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"477","DOI":"10.1038\/s41586-020-2864-x","article-title":"Single-cell mutation analysis of clonal evolution in myeloid malignancies","volume":"587","author":"Miles","year":"2020","journal-title":"Nature"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"153","DOI":"10.1007\/978-3-030-14366-4_9","article-title":"Acute Myeloid Leukemia Stem Cell Heterogeneity and Its Clinical Relevance","volume":"1139","author":"Karantanos","year":"2019","journal-title":"Adv. Exp. Med. Biol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"138","DOI":"10.1016\/j.ccr.2010.12.012","article-title":"Coexistence of LMPP-like and GMP-like leukemia stem cells in acute myeloid leukemia","volume":"19","author":"Goardon","year":"2011","journal-title":"Cancer Cell"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"850","DOI":"10.1002\/gcc.22805","article-title":"The cell of origin and the leukemia stem cell in acute myeloid leukemia","volume":"58","author":"Chopra","year":"2019","journal-title":"Genes Chromosomes Cancer"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"418","DOI":"10.1038\/nature25022","article-title":"From haematopoietic stem cells to complex differentiation landscapes","volume":"553","author":"Laurenti","year":"2018","journal-title":"Nature"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"432","DOI":"10.1038\/nature22370","article-title":"Haematopoietic stem and progenitor cells from human pluripotent stem cells","volume":"545","author":"Sugimura","year":"2017","journal-title":"Nature"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"218","DOI":"10.1126\/science.1201219","article-title":"Isolation of single human hematopoietic stem cells capable of long-term multilineage engraftment","volume":"333","author":"Notta","year":"2011","journal-title":"Science"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"839","DOI":"10.1002\/gcc.22806","article-title":"Clonal evolution of acute myeloid leukemia from diagnosis to relapse","volume":"58","author":"Vosberg","year":"2019","journal-title":"Genes Chromosomes Cancer"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1113","DOI":"10.1038\/s41375-018-0319-2","article-title":"Single cell analysis of clonal architecture in acute myeloid leukaemia","volume":"33","author":"Potter","year":"2019","journal-title":"Leukemia"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Hughes, A.E., Magrini, V., Demeter, R., Miller, C.A., Fulton, R., Fulton, L.L., Eades, W.C., Elliott, K., Heath, S., and Westervelt, P. (2014). Clonal architecture of secondary acute myeloid leukemia defined by single-cell sequencing. PLoS Genet., 10.","DOI":"10.1371\/journal.pgen.1004462"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Paguirigan, A.L., Smith, J., Meshinchi, S., Carroll, M., Maley, C., and Radich, J.P. (2015). Single-cell genotyping demonstrates complex clonal diversity in acute myeloid leukemia. Sci. Transl. Med., 7.","DOI":"10.1126\/scitranslmed.aaa0763"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"457","DOI":"10.1016\/j.exphem.2014.01.010","article-title":"Single cell analysis exposes intratumor heterogeneity and suggests that FLT3-ITD is a late event in leukemogenesis","volume":"42","author":"Shouval","year":"2014","journal-title":"Exp. Hematol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"424","DOI":"10.1182\/blood-2016-08-733196","article-title":"Diagnosis and management of AML in adults: 2017 ELN recommendations from an international expert panel","volume":"129","author":"Dohner","year":"2017","journal-title":"Blood"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"953","DOI":"10.1182\/blood.2023022851","article-title":"Identification and surveillance of rare relapse-initiating stem cells during complete remission after transplantation","volume":"143","author":"Dimitriou","year":"2024","journal-title":"Blood"},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Garcia-Garcia, A., Klein, T., Born, G., Hilpert, M., Scherberich, A., Lengerke, C., Skoda, R.C., Bourgine, P.E., and Martin, I. (2021). Culturing patient-derived malignant hematopoietic stem cells in engineered and fully humanized 3D niches. Proc. Natl. Acad. Sci. USA, 118.","DOI":"10.1073\/pnas.2114227118"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"e216","DOI":"10.3324\/haematol.2016.162909","article-title":"Mutational analysis in serial marrow samples during azacitidine treatment in patients with post-transplant relapse of acute myeloid leukemia or myelodysplastic syndromes","volume":"102","author":"Woo","year":"2017","journal-title":"Haematologica"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"680","DOI":"10.1038\/s41375-023-01811-5","article-title":"Immunophenotypic aberrant hematopoietic stem cells in myelodysplastic syndromes: A biomarker for leukemic progression","volume":"37","author":"Hanekamp","year":"2023","journal-title":"Leukemia"},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Lahnemann, D., Koster, J., Szczurek, E., McCarthy, D.J., Hicks, S.C., Robinson, M.D., Vallejos, C.A., Campbell, K.R., Beerenwinkel, N., and Mahfouz, A. (2020). Eleven grand challenges in single-cell data science. Genome Biol., 21.","DOI":"10.1186\/s13059-020-1926-6"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"104","DOI":"10.1038\/nature22993","article-title":"Tracing the origins of relapse in acute myeloid leukaemia to stem cells","volume":"547","author":"Shlush","year":"2017","journal-title":"Nature"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"849","DOI":"10.1016\/j.ccell.2016.11.002","article-title":"Characterization of Rare, Dormant, and Therapy-Resistant Cells in Acute Lymphoblastic Leukemia","volume":"30","author":"Ebinger","year":"2016","journal-title":"Cancer Cell"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"988","DOI":"10.1158\/2159-8290.CD-15-0298","article-title":"Mass Cytometric Functional Profiling of Acute Myeloid Leukemia Defines Cell-Cycle and Immunophenotypic Properties That Correlate with Known Responses to Therapy","volume":"5","author":"Behbehani","year":"2015","journal-title":"Cancer Discov."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"388","DOI":"10.1158\/2159-8290.CD-21-0410","article-title":"Implementing a Functional Precision Medicine Tumor Board for Acute Myeloid Leukemia","volume":"12","author":"Malani","year":"2022","journal-title":"Cancer Discov."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"506","DOI":"10.1038\/nature10738","article-title":"Clonal evolution in relapsed acute myeloid leukaemia revealed by whole-genome sequencing","volume":"481","author":"Ding","year":"2012","journal-title":"Nature"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"328","DOI":"10.1038\/nature13038","article-title":"Identification of pre-leukaemic haematopoietic stem cells in acute leukaemia","volume":"506","author":"Shlush","year":"2014","journal-title":"Nature"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"603","DOI":"10.1182\/blood-2011-10-388629","article-title":"Cell lineage analysis of acute leukemia relapse uncovers the role of replication-rate heterogeneity and microsatellite instability","volume":"120","author":"Shlush","year":"2012","journal-title":"Blood"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"e228","DOI":"10.3324\/haematol.2019.222323","article-title":"EZH2 mutations and impact on clinical outcome: An analysis in 1,604 patients with newly diagnosed acute myeloid leukemia","volume":"105","author":"Stasik","year":"2020","journal-title":"Haematologica"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"69","DOI":"10.1038\/nm.4247","article-title":"Loss of the histone methyltransferase EZH2 induces resistance to multiple drugs in acute myeloid leukemia","volume":"23","author":"Gollner","year":"2017","journal-title":"Nat. Med."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"13","DOI":"10.1016\/j.exphem.2020.06.003","article-title":"AML chemoresistance: The role of mutant TP53 subclonal expansion and therapy strategy","volume":"87","author":"Yan","year":"2020","journal-title":"Exp. Hematol."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Tao, L., Raz, O., Marx, Z., Ghosh, M.S., Huber, S., Greindl-Junghans, J., Biezuner, T., Amir, S., Milo, L., and Adar, R. (2021). Retrospective cell lineage reconstruction in humans by using short tandem repeats. Cell Rep. Methods, 1.","DOI":"10.1016\/j.crmeth.2021.100054"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2436","DOI":"10.1093\/nar\/gky1318","article-title":"Short tandem repeat stutter model inferred from direct measurement of in vitro stutter noise","volume":"47","author":"Raz","year":"2019","journal-title":"Nucleic Acids Res."},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Price, M.N., Dehal, P.S., and Arkin, A.P. (2010). FastTree 2-approximately maximum-likelihood trees for large alignments. PLoS ONE, 5.","DOI":"10.1371\/journal.pone.0009490"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"452","DOI":"10.1038\/s41586-018-0043-0","article-title":"Renewing Felsenstein\u2019s phylogenetic bootstrap in the era of big data","volume":"556","author":"Lemoine","year":"2018","journal-title":"Nature"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1007\/978-1-0716-4310-5_2","article-title":"Tracking Somatic Mutations for Lineage Reconstruction","volume":"2886","author":"Neumeier","year":"2025","journal-title":"Methods Mol. Biol."},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Biezuner, T., Brilon, Y., Arye, A.B., Oron, B., Kadam, A., Danin, A., Furer, N., Minden, M.D., Hwan Kim, D.D., and Shapira, S. (2022). An improved molecular inversion probe based targeted sequencing approach for low variant allele frequency. NAR Genom. Bioinform., 4.","DOI":"10.1093\/nargab\/lqab125"},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Bushnell, B., Rood, J., and Singer, E. (2017). BBMerge-Accurate paired shotgun read merging via overlap. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0185056"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"10","DOI":"10.14806\/ej.17.1.200","article-title":"Cutadapt Removes Adapter Sequences from High-Throughput Sequencing Reads","volume":"17","author":"Martin","year":"2011","journal-title":"EMBnet J."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Li, W., and Freudenberg, J. (2014). Mappability and read length. Front. Genet., 5.","DOI":"10.3389\/fgene.2014.00381"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"2078","DOI":"10.1093\/bioinformatics\/btp352","article-title":"The Sequence Alignment\/Map format and SAMtools","volume":"25","author":"Li","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1297","DOI":"10.1101\/gr.107524.110","article-title":"The Genome Analysis Toolkit: A MapReduce framework for analyzing next-generation DNA sequencing data","volume":"20","author":"McKenna","year":"2010","journal-title":"Genome Res."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"568","DOI":"10.1101\/gr.129684.111","article-title":"VarScan 2: Somatic mutation and copy number alteration discovery in cancer by exome sequencing","volume":"22","author":"Koboldt","year":"2012","journal-title":"Genome Res."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"912","DOI":"10.1038\/ng.3036","article-title":"Integrating mapping-, assembly- and haplotype-based approaches for calling variants in clinical sequencing applications","volume":"46","author":"Rimmer","year":"2014","journal-title":"Nat. Genet."},{"key":"ref_50","doi-asserted-by":"crossref","unstructured":"Wang, K., Li, M., and Hakonarson, H. (2010). ANNOVAR: Functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res., 38.","DOI":"10.1093\/nar\/gkq603"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/9\/4224\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T17:24:06Z","timestamp":1760030646000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/9\/4224"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,4,29]]},"references-count":50,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2025,5]]}},"alternative-id":["ijms26094224"],"URL":"https:\/\/doi.org\/10.3390\/ijms26094224","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,4,29]]}}}