Abstract
Internal tandem duplication (ITD) in the fms-like tyrosine kinase 3 (FLT3) gene is one of the most frequent genetic alteration in acute myeloid leukemia (AML), and it is associated with worse clinical outcome. Not only the presence but also the size, localization and the rate of this variant or the presence of multiple ITDs has prognostic information. The traditional PCR based diagnostic methods cannot provide information about all of these parameters in one assay, however the application of next generation sequencing (NGS) technique can be a reliable solution for this diagnostic problem. In order to evaluate the analytical properties of an NGS-based FLT3-ITD detection assay a quality control sample was prepared from DNA of AML patients containing 19 different FLT3-ITD variants identified by NGS. The higher the total read count was in a certain sample of the NGS run, the more ITD variant types could be detected. The maximal sensitivity of FLT3-ITD detection by NGS technique was as low as 0.007% FLT3-ITD/total allele rate, however, below 0.1% rate, the reproducibility of the quantitation was poor (CV > 25%). DNA pools with several FLT3-ITDs can be used efficiently for analytical evaluation of NGS-based FLT3-ITD quantitation testing several different ITD sequences and rates, simultaneously.
| Original language | English |
|---|---|
| Pages (from-to) | 25-29 |
| Number of pages | 5 |
| Journal | Journal of Biotechnology |
| Volume | 303 |
| DOIs | |
| Publication status | Published - szept. 10 2019 |
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ASJC Scopus subject areas
- Biotechnology
- Bioengineering
- Applied Microbiology and Biotechnology
Cite this
A DNA pool of FLT3-ITD positive DNA samples can be used efficiently for analytical evaluation of NGS-based FLT3-ITD quantitation - Testing several different ITD sequences and rates, simultaneously. / Mezei, Zoltán A.; Tornai, Dávid; Földesi, Róza; Madar, László; Sümegi, Andrea; Papp, M.; Antal-Szalmás, P.
In: Journal of Biotechnology, Vol. 303, 10.09.2019, p. 25-29.Research output: Article
}
TY - JOUR
T1 - A DNA pool of FLT3-ITD positive DNA samples can be used efficiently for analytical evaluation of NGS-based FLT3-ITD quantitation - Testing several different ITD sequences and rates, simultaneously
AU - Mezei, Zoltán A.
AU - Tornai, Dávid
AU - Földesi, Róza
AU - Madar, László
AU - Sümegi, Andrea
AU - Papp, M.
AU - Antal-Szalmás, P.
PY - 2019/9/10
Y1 - 2019/9/10
N2 - Internal tandem duplication (ITD) in the fms-like tyrosine kinase 3 (FLT3) gene is one of the most frequent genetic alteration in acute myeloid leukemia (AML), and it is associated with worse clinical outcome. Not only the presence but also the size, localization and the rate of this variant or the presence of multiple ITDs has prognostic information. The traditional PCR based diagnostic methods cannot provide information about all of these parameters in one assay, however the application of next generation sequencing (NGS) technique can be a reliable solution for this diagnostic problem. In order to evaluate the analytical properties of an NGS-based FLT3-ITD detection assay a quality control sample was prepared from DNA of AML patients containing 19 different FLT3-ITD variants identified by NGS. The higher the total read count was in a certain sample of the NGS run, the more ITD variant types could be detected. The maximal sensitivity of FLT3-ITD detection by NGS technique was as low as 0.007% FLT3-ITD/total allele rate, however, below 0.1% rate, the reproducibility of the quantitation was poor (CV > 25%). DNA pools with several FLT3-ITDs can be used efficiently for analytical evaluation of NGS-based FLT3-ITD quantitation testing several different ITD sequences and rates, simultaneously.
AB - Internal tandem duplication (ITD) in the fms-like tyrosine kinase 3 (FLT3) gene is one of the most frequent genetic alteration in acute myeloid leukemia (AML), and it is associated with worse clinical outcome. Not only the presence but also the size, localization and the rate of this variant or the presence of multiple ITDs has prognostic information. The traditional PCR based diagnostic methods cannot provide information about all of these parameters in one assay, however the application of next generation sequencing (NGS) technique can be a reliable solution for this diagnostic problem. In order to evaluate the analytical properties of an NGS-based FLT3-ITD detection assay a quality control sample was prepared from DNA of AML patients containing 19 different FLT3-ITD variants identified by NGS. The higher the total read count was in a certain sample of the NGS run, the more ITD variant types could be detected. The maximal sensitivity of FLT3-ITD detection by NGS technique was as low as 0.007% FLT3-ITD/total allele rate, however, below 0.1% rate, the reproducibility of the quantitation was poor (CV > 25%). DNA pools with several FLT3-ITDs can be used efficiently for analytical evaluation of NGS-based FLT3-ITD quantitation testing several different ITD sequences and rates, simultaneously.
KW - Analytical validation
KW - Deep next generation sequencing (NGS)
KW - Fms-like tyrosine kinase 3 (FLT3)
KW - Internal tandem duplication (ITD)
UR - http://www.scopus.com/inward/record.url?scp=85069723243&partnerID=8YFLogxK
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U2 - 10.1016/j.jbiotec.2019.06.305
DO - 10.1016/j.jbiotec.2019.06.305
M3 - Article
AN - SCOPUS:85069723243
VL - 303
SP - 25
EP - 29
JO - Journal of Biotechnology
JF - Journal of Biotechnology
SN - 0168-1656
ER -