Please use this identifier to cite or link to this item: https://hdl.handle.net/10316/113492
Title: An Efficient Method to Prepare Barcoded cDNA Libraries from Plant Callus for Long-Read Sequencing
Authors: Cordeiro, Daniela 
Camelo, Alexandra 
Pedrosa, Ana Carolina 
Brandão, Inês 
Canhoto, Jorge 
Espirito Santo, Christophe 
Correia, Sandra 
Keywords: amplification-free protocol; direct cDNA sequencing; high-throughput sequencing; MinION; Oxford Nanopore Technologies®; poly(A) RNA; Solanaceae; Solanum betaceum; transcriptome; woody plant
Issue Date: 15-Mar-2023
Publisher: MDPI
Project: SFRH/BD/136925/2018 
CFE-Centre for Functional Ecology-Science for People & the Planet’s Strategic Plan (UIDB/04004/2020) 
CENTRO-01-0145-FEDER-000020 
metadata.degois.publication.title: Methods and Protocols
metadata.degois.publication.volume: 6
metadata.degois.publication.issue: 2
Abstract: Long-read sequencing methods allow a comprehensive analysis of transcriptomes in identifying full-length transcripts. This revolutionary method represents a considerable breakthrough for non-model species since it allows enhanced gene annotation and gene expression studies when compared to former sequencing methods. However, woody plant tissues are challenging to the successful preparation of cDNA libraries, thus, impairing further cutting-edge sequencing analyses. Here, a detailed protocol for preparing cDNA libraries suitable for high throughput RNA sequencing using Oxford Nanopore Technologies® is described. This method was used to prepare eight barcoded cDNA libraries from two Solanum betaceum cell lines: one with compact morphology and embryogenic competency (EC) and another with friable and non-embryogenic (NEC). The libraries were successfully sequenced, and data quality assessment showed high mean quality scores. Using this method, long-read sequencing will allow a comprehensive analysis of plant transcriptomes.
URI: https://hdl.handle.net/10316/113492
ISSN: 2409-9279
DOI: 10.3390/mps6020031
Rights: openAccess
Appears in Collections:FCTUC Ciências da Vida - Artigos em Revistas Internacionais
I&D CFE - Artigos em Revistas Internacionais

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