Quantitative mapping of the cellular small RNA landscape with AQRNA-seq.
Nat Biotechnol
; 39(8): 978-988, 2021 08.
Article
en En
| MEDLINE
| ID: mdl-33859402
ABSTRACT
Current next-generation RNA-sequencing (RNA-seq) methods do not provide accurate quantification of small RNAs within a sample, due to sequence-dependent biases in capture, ligation and amplification during library preparation. We present a method, absolute quantification RNA-sequencing (AQRNA-seq), that minimizes biases and provides a direct, linear correlation between sequencing read count and copy number for all small RNAs in a sample. Library preparation and data processing were optimized and validated using a 963-member microRNA reference library, oligonucleotide standards of varying length, and RNA blots. Application of AQRNA-seq to a panel of human cancer cells revealed >800 detectable miRNAs that varied during cancer progression, while application to bacterial transfer RNA pools, with the challenges of secondary structure and abundant modifications, revealed 80-fold variation in tRNA isoacceptor levels, stress-induced site-specific tRNA fragmentation, quantitative modification maps, and evidence for stress-induced, tRNA-driven, codon-biased translation. AQRNA-seq thus provides a versatile means to quantitatively map the small RNA landscape in cells.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Alineación de Secuencia
/
Análisis de Secuencia de ARN
/
MicroARNs
Límite:
Humans
Idioma:
En
Revista:
Nat Biotechnol
Asunto de la revista:
BIOTECNOLOGIA
Año:
2021
Tipo del documento:
Article
País de afiliación:
Estados Unidos