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Exploring the genes of yerba mate (Ilex paraguariensis A. St.-Hil.) by NGS and de novo transcriptome assembly.

Debat HJ, Grabiele M, Aguilera PM, Bubillo RE, Otegui MB, Ducasse DA, Zapata PD, Marti DA - PLoS ONE (2014)

Bottom Line: We have also pinpointed several members of the gene silencing pathway, and characterized the silencing effector Argonaute1.We present here the first draft of the transcribed genomes of the yerba mate chloroplast and mitochondrion.Moreover, we provide a collection of over 10,800 SSR accessible to the scientific community interested in yerba mate genetic improvement.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Patología Vegetal, Centro de Investigaciones Agropecuarias, Instituto Nacional de Tecnología Agropecuaria (IPAVE-CIAP-INTA), Córdoba, Argentina.

ABSTRACT
Yerba mate (Ilex paraguariensis A. St.-Hil.) is an important subtropical tree crop cultivated on 326,000 ha in Argentina, Brazil and Paraguay, with a total yield production of more than 1,000,000 t. Yerba mate presents a strong limitation regarding sequence information. The NCBI GenBank lacks an EST database of yerba mate and depicts only 80 DNA sequences, mostly uncharacterized. In this scenario, in order to elucidate the yerba mate gene landscape by means of NGS, we explored and discovered a vast collection of I. paraguariensis transcripts. Total RNA from I. paraguariensis was sequenced by Illumina HiSeq-2000 obtaining 72,031,388 pair-end 100 bp sequences. High quality reads were de novo assembled into 44,907 transcripts encompassing 40 million bases with an estimated coverage of 180X. Multiple sequence analysis allowed us to predict that yerba mate contains ∼ 32,355 genes and 12,551 gene variants or isoforms. We identified and categorized members of more than 100 metabolic pathways. Overall, we have identified ∼ 1,000 putative transcription factors, genes involved in heat and oxidative stress, pathogen response, as well as disease resistance and hormone response. We have also identified, based in sequence homology searches, novel transcripts related to osmotic, drought, salinity and cold stress, senescence and early flowering. We have also pinpointed several members of the gene silencing pathway, and characterized the silencing effector Argonaute1. We predicted a diverse supply of putative microRNA precursors involved in developmental processes. We present here the first draft of the transcribed genomes of the yerba mate chloroplast and mitochondrion. The putative sequence and predicted structure of the caffeine synthase of yerba mate is presented. Moreover, we provide a collection of over 10,800 SSR accessible to the scientific community interested in yerba mate genetic improvement. This contribution broadly expands the limited knowledge of yerba mate genes, and is presented as the first genomic resource of this important crop.

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MiR156 gene family in yerba mate.(a) Several mature miRNAs were predicted in yerba mate based in sequence homology to Mirbase. In the particular case of miR156, nine isoform variants were predicted with high sequence homology and minor mismatches. An insertion of a “A” at position 10 in miR156b and c forms, slightly affected the precursors secondary structure at the miRNA/miRNA* coordinates that can be observed as a bulge in (b). While the homology at the mature miR156 is high, the diversity among precursors of the miRNA gene family is extensive (d). A library generated of predicted SPL mRNAs of yerba mate was evaluated as a target of Ipa-miR156. A strong interaction with a high expectation score was in silico predicted for SPL9, SPL6 and SPL4 with Ipa-miR156 (c). These SPL genes significantly differ in their nucleotide sequence, however a strong conservation of the miR156 target can be observed in the 3 genes (green triangle, e).
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pone-0109835-g006: MiR156 gene family in yerba mate.(a) Several mature miRNAs were predicted in yerba mate based in sequence homology to Mirbase. In the particular case of miR156, nine isoform variants were predicted with high sequence homology and minor mismatches. An insertion of a “A” at position 10 in miR156b and c forms, slightly affected the precursors secondary structure at the miRNA/miRNA* coordinates that can be observed as a bulge in (b). While the homology at the mature miR156 is high, the diversity among precursors of the miRNA gene family is extensive (d). A library generated of predicted SPL mRNAs of yerba mate was evaluated as a target of Ipa-miR156. A strong interaction with a high expectation score was in silico predicted for SPL9, SPL6 and SPL4 with Ipa-miR156 (c). These SPL genes significantly differ in their nucleotide sequence, however a strong conservation of the miR156 target can be observed in the 3 genes (green triangle, e).

Mentions: MicroRNAs (miRNAs) are small non-coding RNAs that modulate plant gene expression by means of gene silencing through sequence-specific inhibition of target mRNAs. MiRNAs derive from precise processing of precursor transcripts with stem-loop secondary-structure features that are recognized by a Dicer-like complex. Mature miRNAs are loaded predominantly onto AGO1 and target endogenous RNAs for their degradation or translational arrest [46]. By combining two in silico based approaches we engaged in an attempted characterization of putative miRNA precursors in yerba mate. A yerba mate miRNA sequence prediction report based in UEA small RNA workbench platform and canonical relaxed mapping of conserved precursor miRNAs to the yerba mate transcriptome, indicated the presence of at least 59 pre-miRNAs corresponding to 41 of both young and ancient miRNA families (Table S10). The miR156 gene family has been involved in the regulation of developmental timing, vegetative phase change, flowering and sex identity in plants [47]–[49]. In yerba mate several mature miRNAs were predicted based in sequence homology to miRBase [50] (Figure 6a). In the particular case of miR156, nine isoform variants were predicted with high sequence homology and minor mismatches. An insertion of a “A” at position 10 in miR156b and c forms, slightly affected the precursors secondary structure at the miRNA/miRNA* coordinates that can be observed as a bulge in Figure 6b. While the homology at the mature miR156 was high, the diversity among precursors of the miRNA gene family was extensive (Figure 6d). A library generated of predicted Squamosa Promoter Binding Protein-Like (SPL) mRNAs of yerba mate was evaluated as a target of Ipa-miR156. A strong interaction with a high expectation score was in silico predicted for SPL9, SPL6 and SPL4 with Ipa-miR156 (Figure 6c), which are typically conserved and validated targets of miR156 in plants [51]. These SPL genes significantly differed in their nucleotide sequence, however a strong conservation of the specific miR156 target could be observed in the 3 genes (Figure 6e). The identification of transcripts related to sex identity, such as miR156 and SPL gene families, is of special interest in yerba mate. In this diclino-dioecious crop, plants cannot be recognized as male or female prior to their first blooming, which occurs between 3 and 10 years post seedling emergence [14], delaying considerably the selection of parentals for breeding purposes. So, a cost-effective early sex determination system would be promising for the yerba mate breeding programs. It is tempting to postulate, that perhaps the determination of expression levels of these particular genes during yerba mate plant development, may be employed as a gender predictor at early stages.


Exploring the genes of yerba mate (Ilex paraguariensis A. St.-Hil.) by NGS and de novo transcriptome assembly.

Debat HJ, Grabiele M, Aguilera PM, Bubillo RE, Otegui MB, Ducasse DA, Zapata PD, Marti DA - PLoS ONE (2014)

MiR156 gene family in yerba mate.(a) Several mature miRNAs were predicted in yerba mate based in sequence homology to Mirbase. In the particular case of miR156, nine isoform variants were predicted with high sequence homology and minor mismatches. An insertion of a “A” at position 10 in miR156b and c forms, slightly affected the precursors secondary structure at the miRNA/miRNA* coordinates that can be observed as a bulge in (b). While the homology at the mature miR156 is high, the diversity among precursors of the miRNA gene family is extensive (d). A library generated of predicted SPL mRNAs of yerba mate was evaluated as a target of Ipa-miR156. A strong interaction with a high expectation score was in silico predicted for SPL9, SPL6 and SPL4 with Ipa-miR156 (c). These SPL genes significantly differ in their nucleotide sequence, however a strong conservation of the miR156 target can be observed in the 3 genes (green triangle, e).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4199719&req=5

pone-0109835-g006: MiR156 gene family in yerba mate.(a) Several mature miRNAs were predicted in yerba mate based in sequence homology to Mirbase. In the particular case of miR156, nine isoform variants were predicted with high sequence homology and minor mismatches. An insertion of a “A” at position 10 in miR156b and c forms, slightly affected the precursors secondary structure at the miRNA/miRNA* coordinates that can be observed as a bulge in (b). While the homology at the mature miR156 is high, the diversity among precursors of the miRNA gene family is extensive (d). A library generated of predicted SPL mRNAs of yerba mate was evaluated as a target of Ipa-miR156. A strong interaction with a high expectation score was in silico predicted for SPL9, SPL6 and SPL4 with Ipa-miR156 (c). These SPL genes significantly differ in their nucleotide sequence, however a strong conservation of the miR156 target can be observed in the 3 genes (green triangle, e).
Mentions: MicroRNAs (miRNAs) are small non-coding RNAs that modulate plant gene expression by means of gene silencing through sequence-specific inhibition of target mRNAs. MiRNAs derive from precise processing of precursor transcripts with stem-loop secondary-structure features that are recognized by a Dicer-like complex. Mature miRNAs are loaded predominantly onto AGO1 and target endogenous RNAs for their degradation or translational arrest [46]. By combining two in silico based approaches we engaged in an attempted characterization of putative miRNA precursors in yerba mate. A yerba mate miRNA sequence prediction report based in UEA small RNA workbench platform and canonical relaxed mapping of conserved precursor miRNAs to the yerba mate transcriptome, indicated the presence of at least 59 pre-miRNAs corresponding to 41 of both young and ancient miRNA families (Table S10). The miR156 gene family has been involved in the regulation of developmental timing, vegetative phase change, flowering and sex identity in plants [47]–[49]. In yerba mate several mature miRNAs were predicted based in sequence homology to miRBase [50] (Figure 6a). In the particular case of miR156, nine isoform variants were predicted with high sequence homology and minor mismatches. An insertion of a “A” at position 10 in miR156b and c forms, slightly affected the precursors secondary structure at the miRNA/miRNA* coordinates that can be observed as a bulge in Figure 6b. While the homology at the mature miR156 was high, the diversity among precursors of the miRNA gene family was extensive (Figure 6d). A library generated of predicted Squamosa Promoter Binding Protein-Like (SPL) mRNAs of yerba mate was evaluated as a target of Ipa-miR156. A strong interaction with a high expectation score was in silico predicted for SPL9, SPL6 and SPL4 with Ipa-miR156 (Figure 6c), which are typically conserved and validated targets of miR156 in plants [51]. These SPL genes significantly differed in their nucleotide sequence, however a strong conservation of the specific miR156 target could be observed in the 3 genes (Figure 6e). The identification of transcripts related to sex identity, such as miR156 and SPL gene families, is of special interest in yerba mate. In this diclino-dioecious crop, plants cannot be recognized as male or female prior to their first blooming, which occurs between 3 and 10 years post seedling emergence [14], delaying considerably the selection of parentals for breeding purposes. So, a cost-effective early sex determination system would be promising for the yerba mate breeding programs. It is tempting to postulate, that perhaps the determination of expression levels of these particular genes during yerba mate plant development, may be employed as a gender predictor at early stages.

Bottom Line: We have also pinpointed several members of the gene silencing pathway, and characterized the silencing effector Argonaute1.We present here the first draft of the transcribed genomes of the yerba mate chloroplast and mitochondrion.Moreover, we provide a collection of over 10,800 SSR accessible to the scientific community interested in yerba mate genetic improvement.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Patología Vegetal, Centro de Investigaciones Agropecuarias, Instituto Nacional de Tecnología Agropecuaria (IPAVE-CIAP-INTA), Córdoba, Argentina.

ABSTRACT
Yerba mate (Ilex paraguariensis A. St.-Hil.) is an important subtropical tree crop cultivated on 326,000 ha in Argentina, Brazil and Paraguay, with a total yield production of more than 1,000,000 t. Yerba mate presents a strong limitation regarding sequence information. The NCBI GenBank lacks an EST database of yerba mate and depicts only 80 DNA sequences, mostly uncharacterized. In this scenario, in order to elucidate the yerba mate gene landscape by means of NGS, we explored and discovered a vast collection of I. paraguariensis transcripts. Total RNA from I. paraguariensis was sequenced by Illumina HiSeq-2000 obtaining 72,031,388 pair-end 100 bp sequences. High quality reads were de novo assembled into 44,907 transcripts encompassing 40 million bases with an estimated coverage of 180X. Multiple sequence analysis allowed us to predict that yerba mate contains ∼ 32,355 genes and 12,551 gene variants or isoforms. We identified and categorized members of more than 100 metabolic pathways. Overall, we have identified ∼ 1,000 putative transcription factors, genes involved in heat and oxidative stress, pathogen response, as well as disease resistance and hormone response. We have also identified, based in sequence homology searches, novel transcripts related to osmotic, drought, salinity and cold stress, senescence and early flowering. We have also pinpointed several members of the gene silencing pathway, and characterized the silencing effector Argonaute1. We predicted a diverse supply of putative microRNA precursors involved in developmental processes. We present here the first draft of the transcribed genomes of the yerba mate chloroplast and mitochondrion. The putative sequence and predicted structure of the caffeine synthase of yerba mate is presented. Moreover, we provide a collection of over 10,800 SSR accessible to the scientific community interested in yerba mate genetic improvement. This contribution broadly expands the limited knowledge of yerba mate genes, and is presented as the first genomic resource of this important crop.

Show MeSH
Related in: MedlinePlus