SHAGIN, D.A., M.A. TURCHANINOVA, I.A. SHAGINA, Mikhail SHUGAY, A.R. ZARETSKY, O.I. ZUEVA, D.A. BOLOTIN, S. LUKYANOV a Dmitriy CHUDAKOV. Application of nonsense-mediated primer exclusion (NOPE) for preparation of unique molecular barcoded libraries. BMC Genomics. London: BioMed Central Ltd, roč. 18, JUN, s. 440-450. ISSN 1471-2164. doi:10.1186/s12864-017-3815-2. 2017.
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Základní údaje
Originální název Application of nonsense-mediated primer exclusion (NOPE) for preparation of unique molecular barcoded libraries
Autoři SHAGIN, D.A. (643 Rusko), M.A. TURCHANINOVA (643 Rusko), I.A. SHAGINA (643 Rusko), Mikhail SHUGAY (643 Rusko, domácí), A.R. ZARETSKY (643 Rusko), O.I. ZUEVA (643 Rusko), D.A. BOLOTIN (643 Rusko), S. LUKYANOV (643 Rusko) a Dmitriy CHUDAKOV (643 Rusko, garant, domácí).
Vydání BMC Genomics, London, BioMed Central Ltd, 2017, 1471-2164.
Další údaje
Originální jazyk angličtina
Typ výsledku Článek v odborném periodiku
Obor 10603 Genetics and heredity
Stát vydavatele Velká Británie a Severní Irsko
Utajení není předmětem státního či obchodního tajemství
WWW URL
Impakt faktor Impact factor: 3.730
Kód RIV RIV/00216224:14740/17:00100344
Organizační jednotka Středoevropský technologický institut
Doi http://dx.doi.org/10.1186/s12864-017-3815-2
UT WoS 000404077700008
Klíčová slova anglicky High-throughput sequencing; Unique molecular identifiers; Targeted resequencing; PCR
Štítky OA, rivok
Příznaky Mezinárodní význam, Recenzováno
Změnil Změnila: Mgr. Pavla Foltynová, Ph.D., učo 106624. Změněno: 13. 3. 2018 14:46.
Anotace
Background: Recently we proposed efficient method to exclude undesirable primers at any stage of amplification reaction, here termed NOPE (NOnsense-mediated Primer Exclusion). According to this method, added oligonucleotide overlapping with the 3'-end of unwanted amplification primer (NOPE oligo) simultaneously provides a template for its elongation. This elongation disrupts specificity of unwanted primer, preventing its further participation in PCR. The suggested approach allows to rationally manage the course of PCR reactions in order to facilitate analysis of complex DNA mixtures as well as to perform multistage PCR bypassing intermediate purification steps. Results: Here we apply NOPE method to DNA library preparation for the high-throughput sequencing (HTS) with the PCR-based introduction of unique molecular identifiers (UMI). We show that NOPE oligo efficiently neutralizes UMI-containing oligonucleotides after introduction of UMI into sample DNA molecules, thus allowing to proceed with further amplification steps without purification and associated loss of starting material. At the same time, NOPE oligo does not affect the efficiency of target PCR amplification. Conclusion: We describe a simple, robust and cheap modification of UMI-labeled HTS libraries preparation procedure, that allows to bypass purification step and thus to preserve starting material which may be limited, e.g. circulating tumor DNA, circulating fetal DNA, or small amounts of isolated cells of interest. Furthermore, demonstrated simplicity and robustness of NOPE method should make it popular in various PCR protocols.
Návaznosti
LQ1601, projekt VaVNázev: CEITEC 2020 (Akronym: CEITEC2020)
Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, CEITEC 2020
633592, interní kód MUNázev: APERIM - Advanced bioinformatics platform for PERsonalised cancer IMmunotherapy (Akronym: APERIM)
Investor: Evropská unie, APERIM - Advanced bioinformatics platform for PERsonalised cancer IMmunotherapy, Health, demographic change and wellbeing (Societal Challenges)
VytisknoutZobrazeno: 29. 3. 2024 15:37