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Supplemental Material for Sunitha et al., 2019
dataset
posted on 2019-01-14, 19:00 authored by Sukumaran Sunitha, Rodrigo Loyola, José Antonio Alcalde, Patricio Arce-Johnson, José Tomás Matus, Christopher D. RockSupplementary Figure S1. Size distributions of adapter-trimmed and pre-filtered reads in the 28 UV experimental sRNA libraries.
Supplementary Figure S2. Functional validation of differentially expressed miRNA/tasiRNA activities under high-fluence solar UV-B in the field by qRT-PCR of target genes. A) Schematic alignment of vvi-TAS4a, -TAS4b and -TAS4c primary transcript sequences flanking the miR828 binding and TAS4 3’ D4(-) phasi-RNA positions, and location of locus-specific primers used for qRT-PCR. B) Expression profile of miR828* (left panel; only sRNA reads mapping to vvi-MIR828) and pre-MIR828 transcript abundance. Panels C-E: Expression profiles of TAS4a, TAS4b and TAS4c siRNAs (left panels) and their cognate primary undiced transcript abundances. C) TAS4a 3’ D1(+) trans acting siRNA expression profile and the expression profile of TAS4a primary transcript. D) TAS4b 3’ D4(-) trans acting siRNA expression profile and the expression profile of TAS4b primary transcript. E) TAS4c 3’ D4(-) trans acting siRNA expression profile and the expression profile of TAS4c primary transcript. F) TAS4-3'D4(-) target MYBA6 phasi-RNA-3'D6(+) expression profile and the expression profile of MYBA6 mRNA. G) TAS4-3'D4(-) target MYBA7 phasi-RNA-3'D6(+) expression profile and the expression profile of MYBA7 mRNA. Error bars are s.d., except panel B: s.e.m. Asterisks (*) denote significant differences based on analysis of variance (n=4 biological replicates) and comparisons using the Tukey-Kramer honestly significance test (HSD; p < 0.05). Insets, B-G: Target:miRNA/tasi-RNA binding positions (mRNA nucleotide coordinates) and primer binding sites (arrows) are depicted to scale. Target:miRNA binding depicted in Red; Target:tasiRNA binding depicted in Green; Target:phasiRNA binding depicted in Pink; miR828* species is depicted in Peach.
Supplementary Figure S3. miR403f mature expression profile and the expression profile of MIR403f primary transcript, showing concordant inductions during berry development. Error bars are s.d. Asterisks (*) denote significant differences based on analysis of variance (n=4 biological replicates) and comparisons using the Tukey-Kramer honestly significance test (HSD; p < 0.05). Inset: pre-MIR403f:miRNA binding position (mRNA nucleotide coordinates) and primer binding sites (arrows) are depicted to scale. Target:miRNA binding depicted in Red.
Supplementary File S4. CleaveLand-validated miRNA target T plots.
Supplementary File S5. PhaseTank Align and Cascades Output, UV-regulated PHASI loci.
Supplementary File S6. Novel MIRNA loci characterized de novo by ShortStack, drawn from reads of libraries constructed from UV-B treated samples.
Supplementary File S7. List of primers and parameters used in this study.
Supplementary Table 1a and 1b. small RNA library (a) and degradome (b) quality control parameters.
Supplementary Table 2a and 2b. a) List of validated miRNA targets by CleaveLand4.4 analysis of Pantaleo et al. (2010) degradome dataset. b) Meta-analysis of validated miRNA targets for three published datasets for transcriptome changes of berry skins in response to: UV-C (Suzuki et al., 2015), UV-B (Carbonell-Bejerano et al., 2017) and development (Massonnet et al., 2017).
Supplementary Table 3a-d. DESeq2 output and associated ShortStack raw Counts dataset parameters for differentially expressed miRNA comparisons outlined in Table 1 summarized for target functions, and normalized reads per 20M for visualization of compared effects.
Supplementary Table 4. PhaseTank output of degradome TAS loci targets and miRNA/phasi-RNA trigger predictions.
Supplementary Figure S2. Functional validation of differentially expressed miRNA/tasiRNA activities under high-fluence solar UV-B in the field by qRT-PCR of target genes. A) Schematic alignment of vvi-TAS4a, -TAS4b and -TAS4c primary transcript sequences flanking the miR828 binding and TAS4 3’ D4(-) phasi-RNA positions, and location of locus-specific primers used for qRT-PCR. B) Expression profile of miR828* (left panel; only sRNA reads mapping to vvi-MIR828) and pre-MIR828 transcript abundance. Panels C-E: Expression profiles of TAS4a, TAS4b and TAS4c siRNAs (left panels) and their cognate primary undiced transcript abundances. C) TAS4a 3’ D1(+) trans acting siRNA expression profile and the expression profile of TAS4a primary transcript. D) TAS4b 3’ D4(-) trans acting siRNA expression profile and the expression profile of TAS4b primary transcript. E) TAS4c 3’ D4(-) trans acting siRNA expression profile and the expression profile of TAS4c primary transcript. F) TAS4-3'D4(-) target MYBA6 phasi-RNA-3'D6(+) expression profile and the expression profile of MYBA6 mRNA. G) TAS4-3'D4(-) target MYBA7 phasi-RNA-3'D6(+) expression profile and the expression profile of MYBA7 mRNA. Error bars are s.d., except panel B: s.e.m. Asterisks (*) denote significant differences based on analysis of variance (n=4 biological replicates) and comparisons using the Tukey-Kramer honestly significance test (HSD; p < 0.05). Insets, B-G: Target:miRNA/tasi-RNA binding positions (mRNA nucleotide coordinates) and primer binding sites (arrows) are depicted to scale. Target:miRNA binding depicted in Red; Target:tasiRNA binding depicted in Green; Target:phasiRNA binding depicted in Pink; miR828* species is depicted in Peach.
Supplementary Figure S3. miR403f mature expression profile and the expression profile of MIR403f primary transcript, showing concordant inductions during berry development. Error bars are s.d. Asterisks (*) denote significant differences based on analysis of variance (n=4 biological replicates) and comparisons using the Tukey-Kramer honestly significance test (HSD; p < 0.05). Inset: pre-MIR403f:miRNA binding position (mRNA nucleotide coordinates) and primer binding sites (arrows) are depicted to scale. Target:miRNA binding depicted in Red.
Supplementary File S4. CleaveLand-validated miRNA target T plots.
Supplementary File S5. PhaseTank Align and Cascades Output, UV-regulated PHASI loci.
Supplementary File S6. Novel MIRNA loci characterized de novo by ShortStack, drawn from reads of libraries constructed from UV-B treated samples.
Supplementary File S7. List of primers and parameters used in this study.
Supplementary Table 1a and 1b. small RNA library (a) and degradome (b) quality control parameters.
Supplementary Table 2a and 2b. a) List of validated miRNA targets by CleaveLand4.4 analysis of Pantaleo et al. (2010) degradome dataset. b) Meta-analysis of validated miRNA targets for three published datasets for transcriptome changes of berry skins in response to: UV-C (Suzuki et al., 2015), UV-B (Carbonell-Bejerano et al., 2017) and development (Massonnet et al., 2017).
Supplementary Table 3a-d. DESeq2 output and associated ShortStack raw Counts dataset parameters for differentially expressed miRNA comparisons outlined in Table 1 summarized for target functions, and normalized reads per 20M for visualization of compared effects.
Supplementary Table 4. PhaseTank output of degradome TAS loci targets and miRNA/phasi-RNA trigger predictions.
