Analyzing Plant Gene Targeting Outcomes and Conversion Tracts with Nanopore Sequencing.
amplicon sequencing
bioinformatics
genome engineering
homologous recombination
Journal
International journal of molecular sciences
ISSN: 1422-0067
Titre abrégé: Int J Mol Sci
Pays: Switzerland
ID NLM: 101092791
Informations de publication
Date de publication:
08 Sep 2021
08 Sep 2021
Historique:
received:
30
07
2021
revised:
30
08
2021
accepted:
01
09
2021
entrez:
28
9
2021
pubmed:
29
9
2021
medline:
29
10
2021
Statut:
epublish
Résumé
The high-throughput molecular analysis of gene targeting (GT) events is made technically challenging by the residual presetabce of donor molecules. Large donor molecules restrict primer placement, resulting in long amplicons that cannot be readily analyzed using standard NGS pipelines or qPCR-based approaches such as ddPCR. In plants, removal of excess donor is time and resource intensive, often requiring plant regeneration and weeks to months of effort. Here, we utilized Oxford Nanopore Amplicon Sequencing (ONAS) to bypass the limitations imposed by donor molecules with 1 kb of homology to the target and dissected GT outcomes at three loci in
Identifiants
pubmed: 34575882
pii: ijms22189723
doi: 10.3390/ijms22189723
pmc: PMC8467259
pii:
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : MnDRIVE - a partnership between the University of Minnesota and the State of Minnesota.
ID : None
Références
Plant Biotechnol J. 2021 Jul;19(7):1314-1324
pubmed: 33511745
Nat Plants. 2020 Jun;6(6):620-624
pubmed: 32483329
Cell. 1987 Nov 6;51(3):503-12
pubmed: 2822260
Curr Protoc Plant Biol. 2017 Dec;2(4):350-369
pubmed: 33383981
Curr Opin Plant Biol. 2020 Apr;54:79-84
pubmed: 32143167
Plant Cell. 2017 Jun;29(6):1196-1217
pubmed: 28522548
Nat Biotechnol. 2020 Jan;38(1):84-89
pubmed: 31844292
Nat Cell Biol. 2016 Dec 23;19(1):1-9
pubmed: 28008184
Genome Biol. 2019 Jun 24;20(1):129
pubmed: 31234903
Proc Natl Acad Sci U S A. 2021 Jun 1;118(22):
pubmed: 34050019
Bioinformatics. 2018 Sep 15;34(18):3094-3100
pubmed: 29750242
Proc Natl Acad Sci U S A. 1999 Jul 20;96(15):8321-3
pubmed: 10411868
Curr Genet. 2019 Feb;65(1):1-9
pubmed: 29947969
Genes (Basel). 2020 Dec 31;12(1):
pubmed: 33396568
Cell Rep. 2014 Apr 10;7(1):293-305
pubmed: 24685129
Nature. 2017 Apr 20;544(7650):377-380
pubmed: 28405019
PLoS Genet. 2014 Apr 03;10(4):e1004251
pubmed: 24699519
PLoS Comput Biol. 2018 Nov 20;14(11):e1006583
pubmed: 30458005
Gigascience. 2019 May 1;8(5):
pubmed: 30824940
Plant Cell Rep. 2019 Apr;38(4):443-453
pubmed: 30673818
Plant Cell. 2014 Jan;26(1):151-63
pubmed: 24443519
Nat Methods. 2021 Feb;18(2):165-169
pubmed: 33432244
Nat Commun. 2017 May 26;8:15605
pubmed: 28548094