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@ARTICLE{Kranz:834728,
author = {Kranz, Angela and Vogel, Alexander and Degner, Ursula and
Kiefler, Ines and Bott, Michael and Usadel, Björn and
Polen, Tino},
title = {{H}igh precision genome sequencing of engineered
{G}luconobacter oxydans 621{H} by combining long nanopore
and short accurate {I}llumina reads},
journal = {Journal of biotechnology},
volume = {258},
issn = {0168-1656},
address = {Amsterdam [u.a.]},
publisher = {Elsevier Science},
reportid = {FZJ-2017-04627},
pages = {197-205},
year = {2017},
note = {Biotechnologie1},
abstract = {State of the art and novel high-throughput DNA sequencing
technologies enable fascinating opportunities and
applications in the life sciences including microbial
genomics. Short high-quality read data already enable not
only microbial genome sequencing, yet can be inadequately to
solve problems in genome assemblies and for the analysis of
structural variants, especially in engineered microbial cell
factories. Single-molecule real-time sequencing technologies
generating long reads promise to solve such assembly
problems. In our study, we wanted to increase the average
read length of long nanopore reads with R9 chemistry and
conducted a hybrid approach for the analysis of structural
variants to check the genome stability of a recombinant
Gluconobacter oxydans 621H strain (IK003.1) engineered for
improved growth. Therefore we combined accurate Illumina
sequencing technology and low-cost single-molecule nanopore
sequencing using the MinION® device from Oxford Nanopore.
In our hybrid approach with a modified library protocol we
could increase the average size of nanopore 2D reads to
about 18.9 kb. Combining the long MinION nanopore reads with
the high quality short Illumina reads enabled the assembly
of the engineered chromosome into a single contig and
comprehensive detection and clarification of 7 structural
variants including all three known genetically engineered
modifications. We found the genome of IK003.1 was stable
over 70 generations of strain handling including 28 h of
process time in a bioreactor. The long read data revealed a
novel 1420 bp transposon-flanked and ORF-containing sequence
which was hitherto unknown in the G. oxydans 621H reference.
Further analysis and genome sequencing showed that this
region is already present in G. oxydans 621H wild-type
strains. Our data of G. oxydans 621H wild-type DNA from
different resources also revealed in 73 annotated coding
sequences about 91 uniform nucleotide differences including
InDels. Together, our results contribute to an improved high
quality genome reference for G. oxydans 621H which is
available via ENA accession PRJEB18739.},
cin = {IBG-1 / IBG-2},
ddc = {540},
cid = {I:(DE-Juel1)IBG-1-20101118 / I:(DE-Juel1)IBG-2-20101118},
pnm = {581 - Biotechnology (POF3-581)},
pid = {G:(DE-HGF)POF3-581},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:28433722},
UT = {WOS:000412611100024},
doi = {10.1016/j.jbiotec.2017.04.016},
url = {https://juser.fz-juelich.de/record/834728},
}