Abstract
Single-nucleotide variants (SNVs) and copy number variations (CNVs) are the most common genomic variations that cause phenotypic diversity and genetic disorders. MALDI–TOF–MS is a rapid and cost-effective technique for multi-variant genotyping, but it is challenging to efficiently detect CNVs and clustered SNVs, especially to simultaneously detect CNVs and SNVs in one reaction. Herein, a novel strategy termed Target-Allele-Specific Probe Single-Base Extension (TASP-SBE) was devised to efficiently detect CNVs and clustered SNVs with MALDI–TOF–MS. By comprehensive use of traditional SBE and TASP-SBE strategies, a MALDI–TOF–MS assay was also developed to simultaneously detect 28 α-/β-thalassemia mutations in a single reaction system, including 4 α-thalassemia deletions, 3 HBA and 21 HBB SNVs. The results showed that all 28 mutations were sensitively identified, and the CNVs of HBA/HBB genes were also accurately analyzed based on the ratio of peak height (RPH) between the target allele and reference gene. The double-blind evaluation results of 989 thalassemia carrier samples showed a 100% concordance of this assay with other methods. In conclusion, a one-tube MALDI–TOF–MS assay was developed to simultaneously genotype 28 thalassemia mutations. This novel TASP-SBE was also verified a practicable strategy for the detection of CNVs and clustered SNVs, providing a feasible approach for multi-variants analysis with MALDI–TOF–MS technique.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
Original data of the present study are available from the corresponding author upon reasonable request.
References
Abbasi W, French CE, Rockowitz S, Kenna MA, Eliot SA (2022) Evaluation of copy number variants for genetic hearing loss: a review of current approaches and recent findings. Hum Genet 141:387–400
Brosens E, Peters N, van Weelden KS, Bendixen C, Brouwer R, Sleutels F, Bruggenwirth HT, van Ijcken W, Veenma D, Otter S, Wijnen R, Eggink AJ, van Dooren MF, Reutter HM, Rottier RJ, Schnater JM, Tibboel D, de Klein A (2021) Unraveling the genetics of congenital diaphragmatic hernia: an ongoing challenge. Front Pediatr 9:800915
Cho YT, Su H, Wu WJ, Wu DC, Hou MF, Kuo CH, Shiea J (2015) Biomarker characterization by MALDI-TOF/MS. Adv Clin Chem 69:209–254
Dai W, Yang T, Wang Y, Zhao Q, Zhan Y, Ye J, Han L, Qiu W, Zhang H, Liang L, Gu X, Yu Y (2021) Rapid detection of twenty-nine common Chinese glucose-6-phosphate dehydrogenase variants using a matrix-assisted laser desorption/ionization-time of flight mass spectrometry assay on dried blood spots. Clin Biochem 94:27–34
Garg P, Jadhav B, Lee W, Rodriguez OL, Martin-Trujillo A, Sharp AJ (2022) A phenome-wide association study identifies effects of copy-number variation of VNTRs and multicopy genes on multiple human traits. Am J Hum Genet 109:1065–1076
Jabato FM, Seoane P, Perkins JR, Rojano E, Garcia MA, Chagoyen M, Pazos F, Ranea J (2021) Systematic identification of genetic systems associated with phenotypes in patients with rare genomic copy number variations. Hum Genet 140:457–475
Jin W, Yang Z, Tang X, Wang X, Huang Y, Hui C, Yao J, Luan J, Tang S, Wu S, Jin S, Ding C (2022) Simultaneous quantification of SMN1 and SMN2 copy numbers by MALDI-TOF mass spectrometry for spinal muscular atrophy genetic testing. Clin Chim Acta 532:45–52
Kariuki SN, Williams TN (2020) Human genetics and malaria resistance. Hum Genet 139:801–811
Kattamis A, Kwiatkowski JL, Aydinok Y (2022) Thalassaemia Lancet 399:2310–2324
Lai Y, Tao F, Zou Y, Huang M, Lin K, Li Y, Huang W, Zhou W (2022) Molecular spectrum of thalassemia in tropical Hainan Island of southern China: high allele frequency with low health burden. J Genet Genomics 49:1162–1164
Li S, Shi Y, Han X, Chen Y, Shen Y, Hu W, Zhao X, Wang Y (2022) Prenatal diagnosis of chromosomal mosaicism in over 18,000 pregnancies: a five-year single-tertiary-center retrospective analysis. Front Genet 13:876887
Liao HK, Su YN, Kao HY, Hung CC, Wang HT, Chen YJ (2005) Parallel minisequencing followed by multiplex matrix-assisted laser desorption/ionization mass spectrometry assay for beta-thalassemia mutations. J Hum Genet 50:139–150
Looi M, Sivalingam M, Husin ND, Radin FZM, Isa RM, Zakaria SZS, Hussin NH, Alias H, Latiff ZA, Ibrahim H, Jamal R (2011) Multiplexed genotyping of beta globin mutations with MALDI-TOF mass spectrometry. Clin Chim Acta 412:999–1002
Mathew MT, Antoniou A, Ramesh N, Hu M, Gaither J, Mouhlas D, Hashimoto S, Humphrey M, Matthews T, Hunter JM, Reshmi S, Schultz M, Lee K, Pfau R, Cottrell C, McBride KL, Navin NE, Chaudhari BP, Leung ML (2022) A decade’s experience in pediatric chromosomal microarray reveals distinct characteristics across ordering specialties. J Mol Diagn 24:1031–1040
Munkongdee T, Chen P, Winichagoon P, Fucharoen S, Paiboonsukwong K (2020) Update in laboratory diagnosis of thalassemia. Front Mol Biosci 7:74
Thongnoppakhun W, Jiemsup S, Yongkiettrakul S, Kanjanakorn C, Limwongse C, Wilairat P, Vanasant A, Rungroj N, Yenchitsomanus PT (2009) Simple, efficient, and cost-effective multiplex genotyping with matrix assisted laser desorption/ionization time-of-flight mass spectrometry of hemoglobin beta gene mutations. J Mol Diagn 11:334–346
Unger MS, Blank M, Enzlein T, Hopf C (2021) Label-free cell assays to determine compound uptake or drug action using MALDI-TOF mass spectrometry. Nat Protoc 16:5533–5558
Visscher PM, Yengo L, Cox NJ, Wray NR (2021) Discovery and implications of polygenicity of common diseases. Science 373:1468–1473
Wang T, Antonacci-Fulton L, Howe K, Lawson HA, Lucas JK, Phillippy AM, Popejoy AB, Asri M, Carson C, Chaisson M, Chang X, Cook-Deegan R, Felsenfeld AL, Fulton RS, Garrison EP, Garrison NA, Graves-Lindsay TA, Ji H, Kenny EE, Koenig BA, Li D, Marschall T, McMichael JF, Novak AM, Purushotham D, Schneider VA, Schultz BI, Smith MW, Sofia HJ, Weissman T, Flicek P, Li H, Miga KH, Paten B, Jarvis ED, Hall IM, Eichler EE, Haussler D (2022) The human pangenome project: a global resource to map genomic diversity. Nature 604:437–446
Williams GR, Cook L, Lewis LD, Tsongalis GJ, Nerenz RD (2020) Clinical validation of a 106-SNV MALDI-ToF MS pharmacogenomic panel. J Appl Lab Med 5:454–466
Wu LR, Dai P, Wang MX, Chen SX, Cohen EN, Jayachandran G, Zhang JX, Serrano AV, Xie NG, Ueno NT, Reuben JM, Barcenas CH, Zhang DY (2022) Ensemble of nucleic acid absolute quantitation modules for copy number variation detection and RNA profiling. Nat Commun 13:1791
Xiong F, Sun M, Zhang X, Cai R, Zhou Y, Lou J, Zeng L, Sun Q, Xiao Q, Shang X, Wei X, Zhang T, Chen P, Xu X (2010) Molecular epidemiological survey of haemoglobinopathies in the Guangxi zhuang autonomous region of southern China. Clin Genet 78:139–148
Xu XM, Zhou YQ, Luo GX, Liao C, Zhou M, Chen PY, Lu JP, Jia SQ, Xiao GF, Shen X, Li J, Chen HP, Xia YY, Wen YX, Mo QH, Li WD, Li YY, Zhuo LW, Wang ZQ, Chen YJ, Qin CH, Zhong M (2004) The prevalence and spectrum of alpha and beta thalassaemia in Guangdong Province: implications for the future health burden and population screening. J Clin Pathol 57:517–522
Xu ML, Qin JC, Chen BY, Yang XX, Liu HP, Yuan WX, Zhong JM, Huang LM, Zhou WJ (2020) Characterization of a Novel 71.8 kb alpha(0)-thalassemia deletion and subsequent summary of a practical procedure for thalassemia molecular diagnosis. Hemoglobin 44:259–263
Yamamoto T (2021) Genomic aberrations associated with the pathophysiological mechanisms of neurodevelopmental disorders. Cells 10:2317
Zambonin C, Aresta A (2022) MALDI-TOF/MS analysis of non-invasive human urine and saliva samples for the identification of new cancer biomarkers. Molecules 27:1925
Zhao F, Zhang J, Wang X, Liu L, Gong J, Zhai Z, He L, Meng F, Xiao D (2021) A multisite SNP genotyping and macrolide susceptibility gene method for Mycoplasma pneumoniae based on MALDI-TOF MS. iScience 24:102447
Zhou K, Liu Y, Yuan Q, Lai D, Guo S, Wang Z, Su L, Zhang H, Wang X, Guo W, Ji X, Gu X, Huang Q, Guo X, Xing J (2022) Next-generation sequencing-based analysis of urine cell-free mtDNA reveals aberrant fragmentation and mutation profile in cancer patients. Clin Chem 68:561–573
Funding
This study was supported by the National Natural Science Foundation of China (grant no. 81972008), Basic and Applied Basic Research Foundation of Guangdong Province (grant no. 2021A1515012585), and Guangming Soft Science Research Project of Shenzhen (grant no. 2021R01098).
Author information
Authors and Affiliations
Contributions
LJ, XY, and WZ supervised the project and revised the article. QC, WH, YL, and FT analyzed the data and wrote the original draft. ZL, ZH, and XZ provided samples and interpreted the data. MX, and XY performed the experiments and acquired the data.
Corresponding authors
Ethics declarations
Conflict of interest
The authors have no conflict to declare.
Ethical approval
Ethics approval was obtained from the Ethics Committee of Zhujiang Hospital (approval number 2021-SJ-001-01)
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Chen, Q., Yang, X., Huang, W. et al. Target-allele-specific probe single-base extension (TASP-SBE): a novel MALDI–TOF–MS strategy for multi-variants analysis and its application in simultaneous detection of α-/β-thalassemia mutations. Hum Genet 142, 445–456 (2023). https://doi.org/10.1007/s00439-023-02520-w
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00439-023-02520-w