Open AccessValidation of CRISPR targeting for proliferation and cytarabine resistance control genes in the acute myeloid leukemia cell line MOLM-13
Author(s) -
Subhash Prajapati,
Nicholas Dunham,
Hao Fan,
Francine E. Garrett-Bakelman
Publication year - 2022
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/btn-2021-0089
Subject(s) - myeloid leukemia , gene knockdown , crispr , cytarabine , leukemia , biology , myeloid , cancer research , rna interference , cell culture , gene , immunology , genetics , rna
Acute myeloid leukemia patients with FMS-like tyrosine kinase 3–internal tandem duplications and mixed lineage leukemia–protein AF9 fusion proteins suffer from poor clinical outcomes. The MOLM-13 acute myeloid leukemia cell line harbors both of these abnormalities and is used in CRISPR experiments to identify disease drivers. However, experimental observations may be biased or inconclusive in the absence of experimentally validated positive control genes. We validated sgRNAs for knockdown of TP53 for cell proliferation and for DCK knockdown and CDA upregulation for cytarabine resistance control genes in MOLM-13 cells. We have provided a detailed CRISPR protocol applicable to both gene knockdown or activation experiments and downstream leukemic phenotype analyses. Inclusion of these controls in CRISPR experiments will enhance the capacity to identify novel myeloid leukemia drivers in MOLM-13 cells.