Preparation of multiplexed control materials for cancer mutation analysis by genome editing in GM12878 cells. Issue 1 (23rd November 2021)

• Record Type: Journal Article
• Authors: Lin, Guigao; Zhang, Kuo; Han, Yanxi; Peng, Rongxue; Li, Jinming
• Is Part Of: Journal of clinical laboratory analysis. Volume 36:Issue 1(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2021-11-23
• Subjects: control materials -- CRISPR/Cas9 -- genome editing -- next‐generation sequencing -- somatic mutation; Diagnosis, Laboratory -- Periodicals; Medical laboratory technology -- Periodicals; 616
• DOI: 10.1002/jcla.24139
• Languages: English
• ISSNs: 0887-8013

Abstract

Abstract: Background: Quality control materials are necessary for assay development, test validation, and proficiency testing in cancer mutation analysis. Most of the existing controls for somatic mutations only harbor a single variant and are derived from unstable cell lines. This study aimed to establish a method to create stable multianalyte controls in a defined background by genome editing in GM12878 cells, which also can be applied for the reference of next‐generation sequencing. Methods: GM12878 cells were electroporated with a donor plasmid containing a mutant DNA sequence and a Cas9/sgRNA expressing vector. The genome‐edited GM12878 cell was validated with Sanger sequencing, amplification refractory mutation system (ARMS), and next‐generation sequencing (NGS). Results: We have successfully generated a mutant GM12878 cell line harboring the defined variants including single‐nucleotide variants (SNVs), small insertions and deletions (indels), and structural variants (SVs). The introduction of intended mutations in GM12878 cell line was confirmed by both ARMS and sequencing methods. Conclusions: We developed a method for the preparation of the multiplexed controls for reference mutations in cancer gene by genome editing in GM12878 cells. This methodology can be used to generate other stable cancer reference materials with an unlimited supply. Abstract : We describe a method for the preparation of stable and multiplexed control materials for cancer mutation testing. The control materials were developed by integration of a mutant DNA sequence harboring SNVs, indels, and SVs in GM12878 cells.