DinoKnot_data

This folder contains all test data for DinoKnot (https://github.com/HosnaJabbari/DinoKnot)

This README and the corresponding data is organized in sections as presented in the Materials and Methods section of our paper.

qRT-PCR interaction structures

This section contains:

1. the output files containing the dot-bracket interaction structure predicted by DinoKnot within the folder \1_DinoKnot_structures\reverse_primer_RNA_transcript_interaction.

   • The sequence of the interaction structure is located on the 5th line from the bottom of the .txt file (Seq:) (eg. line 401 of 2019-nCoV-N1-R_reference_genome_NC_045512.2.txt)
   • The structure of the interaction structure in dot-bracket form is located on the 3rd line from the bottom of the .txt file (Result_0:) (eg. line 403 of 2019-nCoV-N1-R_reference_genome_NC_045512.2.txt)
   • The Free Energy of the interaction structure is located on the second line from the bottom if the .txt file (Energy_0:) (eg. line 404 of 2019-nCoV-N1-R_reference_genome_NC_045512.2.txt)

2. the VARNA visualizations of the dot-bracket interaction structures within the folder \2_VARNA_Structure_Visualization.

   • The interaction structures are visualized in linear view and the primer sequence is highlighted in red and the expected binding region is highlighted in green.
   • The sequence (Seq:) and dot-bracket structure (Result_0:) from the DinoKnot output is input into VARNA to get the structure visualizations.

3. the sequences of the forward and reverse primers of the primer-probe sets investigated in the file 3_SARS-CoV-2_qRT-PCR_primer_sequences.txt.

4. the free energy of the primer structure predicted by Simfold in the file 4_primer_energy.txt.

5. the sequences in FASTA format of the SARS-CoV-2 (NC_045512.2) gene transcript areas investigated in the file 5_SARS-CoV-2_RNA_transcript_sequences.txt.

6. the structures of the gene transcript areas prior to interaction predicted by Iterative HFold in dot-bracket format within the folder \6_SARS-CoV-2_Transcript_Structures.

7. the WHO qRT-PCR assays containing the sequence of the primer-probe sets investigated in the file 7_whoinhouseassays.pdf.

Comparing DinoKnot to RNAcofold

The folder \2_Comparing_DinoKnot_to_RNAcofold\RNA_cofold_ViennaRNA-2.1.9h contains:

1. an Excel Spread Sheet containing the reverse primer along with the sequence input into RNAcofold, the output from RNAcofold of the interaction structure in dot-bracket form, and the MFE in the file 1_RNAcofold_input_output.xlsx.

2. the terminal output from RNAcofold (the raw terminal log used to generate the data within the excel file from RNAcofold) in the file 2_RNAcofold_terminal_output.txt.

3. the RNAcofold dot-bracket interaction structures visualized with VARNA in the folder \3_RNAcofold_terminal_output.

   • Within this folder the interaction structures are visualized in both linear and nucleic acid view. The primer sequence is highlighted in red and the expected binding region is highlighted in green.

The folder \2_Comparing_DinoKnot_to_RNAcofold\Performance_Measures contains:

4. The output structures of the 109 sRNA and 52 snoRNA interactions are located in 2_Comparing_DinoKnot_to_RNAcofold/Performance_Measures/structure_outputs

5. The scripts used to convert dot-bracket output to bpseq format and to calculate performance measures of DinoKnot and RNAcofold are located in 2_Comparing_DinoKnot_to_RNAcofold/Performance_Measures/scripts

6. The performance measure results are located in 2_Comparing_DinoKnot_to_RNAcofold/Performance_Measures/results

Mutations

1. Primer/probe binding region

The folder \1_Primer_probe_binding_region contains:

1. The output interaction structures of the corresponding primer/probe and the mutated SARS-CoV-2 gene area predicted by DinoKnot in dot-bracket form within the folder \1_DinoKnot_Mutations_in_primer_binding_region.

2. The interaction structure visualizations of the corresponding primer/probe and the mutated SARS-CoV-2 gene area within the folder \2_VARNA_Visualization_Mutation_in_primer_binding_region_Structures.

Note: Both folders are organized with sub-folders labelled with the affected primer/probe.

• For example, the folder \1_DinoKnot_Mutations_in_primer_binding_region\2019-nCoV-N1-P contains the DinoKnot output for the interaction structure of the 2019-nCoV-N1-P probe with the reference genome within the 2019-nCoV-N1-P_reference_genome_NC_045512.2_DNA/DNA.txt file and the mutated genome within the file 2019-nCoV-N1-P_C28,311T_mutation_DNA/DNA.txt
• For example, the folder 2_VARNA_Visualization_Mutation_in_primer_binding_region_Structures\2019-nCoV-N1-P contains the VARNA interaction structure visualation to the reference genome in the file 2019_nCoV_N1-P.jpg and the visualization of the mutated genome in the file 2019_nCoV-N1-P_28311C>T.jpg.
• The reference genome data is present in each folder to be able to compare the mutation to the reference genome.

2. Variants of Concern

The folder \2_Variants_of_Concern contains:

1. The interaction structure of the reverse primer and the gene area transcript of the variant of concern with mutations in this area predicted by DinoKnot in dot-bracket form the folder DinoKnot_Structures. This folder contains subfolders for each of the variants of concern studied.

3. Clinical report of variant causing N gene detection issues

The folder \3_Clinical_report_of_variant_causing_N gene_detection_issues contains:

1. the DinoKnot predicted interaction structure in dot-bracket form of the CCDC-N-F primer with the variant strain (accession ID: EPI_ISL_1061414) in the file CCDC-N-F_EPI_ISL_1061414_DinoKnot_output.txt

2. the VARNA interaction structure visualization of the CCDC-N-F primer with the variant strain (accession ID: EPI_ISL_1061414) in the file CCDC-N-F_EPI_ISL_1061414_VARNA_Structure.png.

3. the DinoKnot predicted interaction structure in dot-bracket form of the CCDC-N-P probe with the variant strain (accession ID: EPI_ISL_1061414) in the file CCDC-N-P_EPI_ISL_1061414_DinoKnot_output(PROBE).txt

4. the DinoKnot predicted interaction structure in dot-bracket form of the CCDC-N-R primer with the variant strain (accession ID: EPI_ISL_1061414) in the file CCDC-N-R_EPI_ISL_1061414_DinoKnot_output(REVERSE).txt

5. the same data as #1-4 above but for the corresponding primer and probe with the SARS-CoV-2 reference genome (NC_045512.2) in the folder \Reference_Genome_Comparison to be able to compare the variant strain to the reference genome.

4. Pseudoknotted Structures

The folder \4_Pseudoknotted_Structures contains:

1. The dot-bracket output structure of the CCR5 mRNA prior to interaction predicted by Iterative HFold in the file CCR5_Iterative_HFold.txt

2. The dot-bracket output structure of the CCR5 mRNA prior to interaction predicted by RNAfold in the file CCR5_RNAfold.txt

3. the DinoKnot predicted interaction structure of the CCR5 mRNA and miR-1224 in dot-bracket form in the file CCR5_miR1224-5p_DinoKnot_output.txt

4. the RNAcofold predicted interaction structure of the CCR5 mRNA and miR-1224 in dot-bracket form in the file CCR5_miR1224-5p_RNAcofold_output.txt

5. The energy computed by HotKnots v2.0 with the DP09 parameters of the pseudoknotted structure predicted by Dinoknot prior to interaction in the file HotKnots_DP09_compute_energy_DinoKnot_pseudoknot_Without_Interaction.txt

6. The energy computed by HotKnots v2.0 with the DP09 parameters of the pseudoknotted structure proposed by Belew et al (2014) in the file HotKnots_DP09_compute_energy_pseudoknot_Belew_et_al_2014

Conference Paper

This folder contains data presented in the IEEE ICHI 2021 conference preceeding paper that are not presented in the journal article: https://ieeexplore.ieee.org/document/9565741 (The conference paper contains preliminary results to our study).

The folder \GISAID_mutation_Screenshot_hCoV-19_app contains screenshots of the GISAID hCoV-19-app on the frequency of mutations in the areas surrounding the primer binding regions of the reverse primers, along with the entire genome from the dates July 23, 2020 and March 28 2021.