471
west- east direction, with mean annual values ranging from 2500 mm in the west, on
the border with Chile, to 400 mm in the east, in the Patagonian steppe (Pastorino
and Gallo 2002). This species has a great capacity to adapt to the marginal, arid and
semi-arid zones of North Patagonia, and therefore it represents an ideal species for
studies of adaptive variability, constituting also a productive alternative for semiarid temperate regions. An exploratory study using candidate genes from model
species allowed identifying genes potentially involved in the adaptive response to
drought stress. Nucleotide diversity and neutrality tests were applied to search for
evidence of adaptive selection in natural populations of Austrocedrus chilensis
(Pomponio et al. 2013). However, the main limitations of this study included the
lack of genomic resources in the species and the low transferability of genes from
model species.
Currently, there are no studies examining the transcriptome-wide gene and allelic
expression patterns under water stress conditions of A. chilensis. Therefore, RNASeq was performed using Roche/454 technology to achieve a global view of differential expression genes that contribute to the tolerance to water-restrictive
environments (Torales et al. unpublished data). Assembly and annotation is presented in Table 17.1 and Fig. 17.2. Ninety percent of the isotigs and 50% of the
singletons had at least one blast hit, of which approximately 60–80% had at least
one assigned ontology (GO) gene term (Table 17.1). An analysis of potential molecular markers was performed, obtaining SNP and SSR markers (Table 17.1). The
differential expression was estimated using the 1731 isotigs (transcripts), taking as
an expression measure the reading counts for each isotig and each condition, normalizing by the total amount of readings of each library. After this analysis, 354
transcripts showed differential expression between control and stress conditions
with values of −2> log2 (FC)> 2 (stress/control comparison): 11 stress-related gene
categories with upregulation and 126 metabolic and cell wall organization gene
categories with downregulation. Validation of 100 SSRs in 6 individuals from 3
A. chilensis populations is currently under analysis, and until now 10 (ten) microsatellite markers are polymorphic.
Cordia trichotoma (Vell.) Steud. is a species of high timber value with the potential of cultivation in its distribution area. In Argentina, it is present in the Yungas and
in the Alto Paraná rainforests. It is also cultivated as an ornamental plant and is
widely used by traditional communities for medicinal purposes for the treatment of
respiratory and urogenital diseases, such as anti-inflammatory, anti-microbial and
anti-helminthic (Menezes et al. 2004; Matias et al. 2015; Oza and Kulkarni 2017).
Another native forest species of interest is Handroanthus impetiginosus (Vell.)
Mattos also valued for its high-quality wood. Its bark has pharmacological applications, and it is used as ornamental in gardens and public areas (Ettori et al. 1996;
Neto and de Morais 2003; Martins 2012; Silva et al. 2012). Transcriptome sequencing of these two subtropical tree species has been performed on leaf tissue using
Roche/454 technology. The de novo assembly allowed to generate isotigs (around
400 pb) and to identify SSR markers (Table 17.1). The total of SSR loci from
H. impetiginosus for which primers were designed are 7324 and the number of
unique markers was 6178. Of those markers, a total of 50 SSRs were selected and
17 Application of High-Throughput Sequencing Technologies in Native Forest Tree…
west- east direction, with mean annual values ranging from 2500 mm in the west, on
the border with Chile, to 400 mm in the east, in the Patagonian steppe (Pastorino
and Gallo 2002). This species has a great capacity to adapt to the marginal, arid and
semi-arid zones of North Patagonia, and therefore it represents an ideal species for
studies of adaptive variability, constituting also a productive alternative for semiarid temperate regions. An exploratory study using candidate genes from model
species allowed identifying genes potentially involved in the adaptive response to
drought stress. Nucleotide diversity and neutrality tests were applied to search for
evidence of adaptive selection in natural populations of Austrocedrus chilensis
(Pomponio et al. 2013). However, the main limitations of this study included the
lack of genomic resources in the species and the low transferability of genes from
model species.
Currently, there are no studies examining the transcriptome-wide gene and allelic
expression patterns under water stress conditions of A. chilensis. Therefore, RNASeq was performed using Roche/454 technology to achieve a global view of differential expression genes that contribute to the tolerance to water-restrictive
environments (Torales et al. unpublished data). Assembly and annotation is presented in Table 17.1 and Fig. 17.2. Ninety percent of the isotigs and 50% of the
singletons had at least one blast hit, of which approximately 60–80% had at least
one assigned ontology (GO) gene term (Table 17.1). An analysis of potential molecular markers was performed, obtaining SNP and SSR markers (Table 17.1). The
differential expression was estimated using the 1731 isotigs (transcripts), taking as
an expression measure the reading counts for each isotig and each condition, normalizing by the total amount of readings of each library. After this analysis, 354
transcripts showed differential expression between control and stress conditions
with values of −2> log2 (FC)> 2 (stress/control comparison): 11 stress-related gene
categories with upregulation and 126 metabolic and cell wall organization gene
categories with downregulation. Validation of 100 SSRs in 6 individuals from 3
A. chilensis populations is currently under analysis, and until now 10 (ten) microsatellite markers are polymorphic.
Cordia trichotoma (Vell.) Steud. is a species of high timber value with the potential of cultivation in its distribution area. In Argentina, it is present in the Yungas and
in the Alto Paraná rainforests. It is also cultivated as an ornamental plant and is
widely used by traditional communities for medicinal purposes for the treatment of
respiratory and urogenital diseases, such as anti-inflammatory, anti-microbial and
anti-helminthic (Menezes et al. 2004; Matias et al. 2015; Oza and Kulkarni 2017).
Another native forest species of interest is Handroanthus impetiginosus (Vell.)
Mattos also valued for its high-quality wood. Its bark has pharmacological applications, and it is used as ornamental in gardens and public areas (Ettori et al. 1996;
Neto and de Morais 2003; Martins 2012; Silva et al. 2012). Transcriptome sequencing of these two subtropical tree species has been performed on leaf tissue using
Roche/454 technology. The de novo assembly allowed to generate isotigs (around
400 pb) and to identify SSR markers (Table 17.1). The total of SSR loci from
H. impetiginosus for which primers were designed are 7324 and the number of
unique markers was 6178. Of those markers, a total of 50 SSRs were selected and
17 Application of High-Throughput Sequencing Technologies in Native Forest Tree…