Protocol detail

Nanopore Direct RNA Sequencing of Arabidopsis

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Total RNA was isolated from Col-0, fpa-8 and 35S::FPA:YFP seedlings as described above. mRNA was isolated and Nanopore DRS libraries prepared (using the SQK-RNA001 Nanopore DRS Kit; Oxford Nanopore Technologies) as previously described (Parker et al., 2020 (link)). Libraries were loaded onto R9.4 flow cells (Oxford Nanopore Technologies) and sequenced using a 48 hr runtime. Four biological replicates were performed for each genotype.

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Parker M.T., Knop K., Zacharaki V., Sherwood A.V., Tomé D., Yu X., Martin P.G., Beynon J., Michaels S.D., Barton G.J, & Simpson G.G. (2021). Widespread premature transcription termination of Arabidopsis thaliana NLR genes by the spen protein FPA. eLife, 10, e65537.

Publication 2021

SQK-RNA001 Nanopore DRS Kit R9.4 flow cells

Biological Cells Genotype Mrna Seedlings

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Variable analysis

independent variables

Genotype (Col-0, fpa-8, 35S::FPA:YFP)

dependent variables

Total RNA expression
MRNA expression

control variables

RNA isolation method
Nanopore DRS library preparation protocol
Sequencing runtime (48 hr)
Number of biological replicates (4)

controls

Positive control: Not specified
Negative control: Not specified

Annotations

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