Fluorogenic sequencing using halogen-fluorescein-labeled nucleotides.
Chembiochem
; 16(8): 1153-7, 2015 May 26.
Article
em En
| MEDLINE
| ID: mdl-25846104
ABSTRACT
Fluorogenic sequencing is a sequencing-by-synthesis technology that combines the advantages of pyrosequencing and fluorescence detection. With native duplex DNA as the major product, we employ polymerase to incorporate the complement- arily matched terminal phosphate-labeled fluorogenic nucleotides into the DNA template and release halogen-fluorescein as the reporter. This red-emitting fluorophore successfully avoids spectral overlap with the autofluorescence background of the flow chip. We fully characterized the enzymatic reaction kinetics of the new substrates, and performed a 35-base sequencing experiment with 60 reaction cycles. Our achievement expands the substrate repertoire for fluorogenic sequencing, and extends the spectral range to obtain better signal-to-background performance.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Análise de Sequência de DNA
/
Fluoresceína
/
Corantes Fluorescentes
/
Halogênios
/
Nucleotídeos
Idioma:
En
Revista:
Chembiochem
Assunto da revista:
BIOQUIMICA
Ano de publicação:
2015
Tipo de documento:
Article