Transcriptome survey of Patagonian southern beech Nothofagus nervosa (= N. Alpina): assembly, annotation and molecular marker discovery
- Equal contributors
1 Instituto de RecursosBiológicos, IRB, Instituto Nacional de Tecnología Agropecuaria (INTA Castelar), CC 25, Castelar, B1712WAA, Argentina
2 Instituto de Biotecnología, CICVyA, Instituto Nacional de Tecnología Agropecuaria (INTA Castelar), CC 25, Castelar, B1712WAA, Argentina
3 EEA Bariloche, Genética Ecológica y Mejoramiento Forestal, Instituto Nacional de Tecnología Agropecuaria (INTA, Bariloche), CC 277, 8400, Bariloche, Argentina
4 Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina
5 CONICET, Buenos Aires, Argentina
BMC Genomics 2012, 13:291 doi:10.1186/1471-2164-13-291Published: 2 July 2012
Nothofagus nervosa is one of the most emblematic native tree species of Patagonian temperate forests. Here, the shotgun RNA-sequencing (RNA-Seq) of the transcriptome of N. nervosa, including de novo assembly, functional annotation, and in silico discovery of potential molecular markers to support population and associations genetic studies, are described.
Pyrosequencing of a young leaf cDNA library generated a total of 111,814 high quality reads, with an average length of 447 bp. De novo assembly using Newbler resulted into 3,005 tentative isotigs (including alternative transcripts). The non-assembled sequences (singletons) were clustered with CD-HIT-454 to identify natural and artificial duplicates from pyrosequencing reads, leading to 21,881 unique singletons. 15,497 out of 24,886 non-redundant sequences or unigenes, were successfully annotated against a plant protein database. A substantial number of simple sequence repeat markers (SSRs) were discovered in the assembled and annotated sequences. More than 40% of the SSR sequences were inside ORF sequences. To confirm the validity of these predicted markers, a subset of 73 SSRs selected through functional annotation evidences were successfully amplified from six seedlings DNA samples, being 14 polymorphic.
This paper is the first report that shows a highly precise representation of the mRNAs diversity present in young leaves of a native South American tree, N. nervosa, as well as its in silico deduced putative functionality. The reported Nothofagus transcriptome sequences represent a unique resource for genetic studies and provide a tool to discover genes of interest and genetic markers that will greatly aid questions involving evolution, ecology, and conservation using genetic and genomic approaches in the genus.