Application of different DNA extraction procedures, library preparation protocols and sequencing platforms: impact on sequencing results

F. Pasquali, I. Do Valle, F. Palma, D. Remondini, G. Manfreda, G. Castellani, Rene S. Hendriksen, A. De Cesare*

*Corresponding author for this work

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Abstract

In this study three DNA extraction procedures, two library preparation protocols and two sequencing platforms were applied to analyse six bacterial cultures and their corresponding DNA obtained as part of a proficiency test. The impact of each variable on sequencing results was assessed using the following parameters: reads quality, assembly and alignment statistics; number of single nucleotide polymorphisms (SNPs), detected applying assembly- and alignment-based strategies; antimicrobial resistance genes (ARGs), identified on de novo assemblies of all sequenced genomes. The investigated nucleic acid extraction procedures, library preparation kits and sequencing platforms do not significantly affect de novo assembly statistics and number of SNPs and ARGs. The only exception was observed for two duplicates, which were associated to one PCR-based library preparation kit. Results from this comparative study can support researchers in the choice toward the available pre-sequencing and sequencing options, and might suggest further comparisons to be performed.
Original languageEnglish
Article numbere02745
JournalHeliyon
Volume5
Issue number10
Number of pages9
ISSN2405-8440
DOIs
Publication statusPublished - 2019

Cite this

Pasquali, F., Do Valle, I., Palma, F., Remondini, D., Manfreda, G., Castellani, G., ... De Cesare, A. (2019). Application of different DNA extraction procedures, library preparation protocols and sequencing platforms: impact on sequencing results. Heliyon, 5(10), [e02745]. https://doi.org/10.1016/j.heliyon.2019.e02745
Pasquali, F. ; Do Valle, I. ; Palma, F. ; Remondini, D. ; Manfreda, G. ; Castellani, G. ; Hendriksen, Rene S. ; De Cesare, A. / Application of different DNA extraction procedures, library preparation protocols and sequencing platforms: impact on sequencing results. In: Heliyon. 2019 ; Vol. 5, No. 10.
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abstract = "In this study three DNA extraction procedures, two library preparation protocols and two sequencing platforms were applied to analyse six bacterial cultures and their corresponding DNA obtained as part of a proficiency test. The impact of each variable on sequencing results was assessed using the following parameters: reads quality, assembly and alignment statistics; number of single nucleotide polymorphisms (SNPs), detected applying assembly- and alignment-based strategies; antimicrobial resistance genes (ARGs), identified on de novo assemblies of all sequenced genomes. The investigated nucleic acid extraction procedures, library preparation kits and sequencing platforms do not significantly affect de novo assembly statistics and number of SNPs and ARGs. The only exception was observed for two duplicates, which were associated to one PCR-based library preparation kit. Results from this comparative study can support researchers in the choice toward the available pre-sequencing and sequencing options, and might suggest further comparisons to be performed.",
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Application of different DNA extraction procedures, library preparation protocols and sequencing platforms: impact on sequencing results. / Pasquali, F.; Do Valle, I.; Palma, F.; Remondini, D.; Manfreda, G.; Castellani, G.; Hendriksen, Rene S.; De Cesare, A.

In: Heliyon, Vol. 5, No. 10, e02745, 2019.

Research output: Contribution to journalJournal articleResearchpeer-review

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T1 - Application of different DNA extraction procedures, library preparation protocols and sequencing platforms: impact on sequencing results

AU - Pasquali, F.

AU - Do Valle, I.

AU - Palma, F.

AU - Remondini, D.

AU - Manfreda, G.

AU - Castellani, G.

AU - Hendriksen, Rene S.

AU - De Cesare, A.

PY - 2019

Y1 - 2019

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AB - In this study three DNA extraction procedures, two library preparation protocols and two sequencing platforms were applied to analyse six bacterial cultures and their corresponding DNA obtained as part of a proficiency test. The impact of each variable on sequencing results was assessed using the following parameters: reads quality, assembly and alignment statistics; number of single nucleotide polymorphisms (SNPs), detected applying assembly- and alignment-based strategies; antimicrobial resistance genes (ARGs), identified on de novo assemblies of all sequenced genomes. The investigated nucleic acid extraction procedures, library preparation kits and sequencing platforms do not significantly affect de novo assembly statistics and number of SNPs and ARGs. The only exception was observed for two duplicates, which were associated to one PCR-based library preparation kit. Results from this comparative study can support researchers in the choice toward the available pre-sequencing and sequencing options, and might suggest further comparisons to be performed.

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