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Increased transcriptome sequencing efficiency with modified Mint-2 digestion-ligation protocol.
Kammonen, Juhana I; Smolander, Olli-Pekka; Sipilä, Timo; Overmyer, Kirk; Auvinen, Petri; Paulin, Lars.
Afiliación
  • Kammonen JI; Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland. Electronic address: juhana.kammonen@helsinki.fi.
  • Smolander OP; Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland.
  • Sipilä T; Department of Biosciences, Faculty of Biological and Environmental Sciences, University of Helsinki, 00014 Helsinki, Finland.
  • Overmyer K; Department of Biosciences, Faculty of Biological and Environmental Sciences, University of Helsinki, 00014 Helsinki, Finland.
  • Auvinen P; Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland.
  • Paulin L; Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland.
Anal Biochem ; 477: 38-40, 2015 May 15.
Article en En | MEDLINE | ID: mdl-25513723
The standard digestion-ligation cloning method enables synthesis of large amounts of complementary DNA (cDNA) from a model organism facilitating study of the transcriptome. Here, we used cDNA amplification of the dimorphic yeast Taphrina betulina as an example of how a library construction protocol can significantly increase sequencing throughput. Two modification steps were introduced to the Evrogen standard Mint-2 protocol to improve its suitability for next-generation sequencing projects. We performed two partial Illumina MiSeq sequencing runs with the modified protocol: one with and one without biotin-purified primers. The results demonstrated that biotinylated libraries increased both accuracy and throughput of the modified protocol. Moreover, our sequencing results indicate that a sequence-specific miscall may affect the output of Illumina's MiSeq platform.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Ascomicetos / ADN Polimerasa Dirigida por ARN / Análisis de Secuencia / Perfilación de la Expresión Génica Idioma: En Revista: Anal Biochem Año: 2015 Tipo del documento: Article Pais de publicación: Estados Unidos
Texto completo
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Imprimir
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Ascomicetos / ADN Polimerasa Dirigida por ARN / Análisis de Secuencia / Perfilación de la Expresión Génica Idioma: En Revista: Anal Biochem Año: 2015 Tipo del documento: Article Pais de publicación: Estados Unidos