A comparison of methods for successful triggering of gene silencing in Coprinus cinereus.
Mol Biotechnol
; 35(3): 283-96, 2007 Mar.
Article
en En
| MEDLINE
| ID: mdl-17652792
ABSTRACT
Post-transcriptional gene-silencing methods (PTGS), including RNAi, are becoming increasingly pervasive in functional genomics. To advance analysis of the recently sequenced Coprinus cinereus genome, a high throughput gene silencing method is essential. We have exploited the GFP reporter gene to evaluate and quantify efficacy of three different silencing strategies. Modular constructs that encompassed antisense, untranslatable sense, and RNAi-mediating hairpin sequences, were transformed into a GFP-expressing host strain. Transformants exhibiting strong downregulation and partial suppression of GFP were recovered with all three constructs. Analyses of protein and transcriptional nucleic acids revealed that the antisense and hairpin sequences yielded similar levels of GFP suppression, and were both more efficient than untranslatable sense sequences. Our antisense vectors will expedite functional characterisation of C. cinereus and the modular nature of the constructs should permit exploitation of directional cDNA libraries for high throughput screening.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Coprinus
/
Silenciador del Gen
Idioma:
En
Revista:
Mol Biotechnol
Asunto de la revista:
BIOLOGIA MOLECULAR
/
BIOTECNOLOGIA
Año:
2007
Tipo del documento:
Article
País de afiliación:
Reino Unido