This notebook performs a simple NCBI BLASTn (Altschul et al. 1990) against two miRNA databases to attempt to identify miRNA in A.pulchra sRNAseq:

miRBase
- Utilizes a modified version, which includes cnidarian miRNA culled from literature by Jill Ahsley.
- - cnidarian-mirbase-mature-v22.1.fasta
MirGeneDB

Relies on the following software:

fastx_toolkit
- fastx_collapser: Collapses duplicate sequences in FastA/Q into single sequence.

1 Create a Bash variables file

This allows usage of Bash variables across R Markdown chunks.

{
echo "#### Assign Variables ####"
echo ""

echo "# Trimmed FastQ naming pattern"
echo "export trimmed_fastqs_pattern='*fastp-adapters-polyG-31bp-merged.fq.gz'"

echo "# Data directories"
echo 'export deep_dive_dir=/home/shared/8TB_HDD_01/sam/gitrepos/deep-dive'
echo 'export deep_dive_data_dir="${deep_dive_dir}/data"'
echo 'export output_dir_top=${deep_dive_dir}/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase'
echo 'export trimmed_fastqs_dir="${deep_dive_dir}/D-Apul/output/08.2-Apul-sRNAseq-trimming-31bp-fastp-merged/trimmed-reads"'
echo 'export blast_dbs_dir="${deep_dive_dir}/data/blast_dbs"'
echo ""

echo "# Input/Output files"
echo 'export collapsed_reads_fasta="collapsed-reads-all.fasta"'
echo 'export concatenated_trimmed_reads_fastq="concatenated-trimmed-reads-all.fastq.gz"'
echo 'export mirbase_mature_fasta_name="cnidarian-mirbase-mature-v22.1.fasta"'
echo 'export mirbase_mature_fasta_no_U="cnidarian-mirbase-mature-v22.1-no_U.fa"'
echo 'export mirgene_mature_fasta_name="mirgene-mature-all-v2.1.fa"'
echo 'export mirgene_mature_fasta_no_U="mirgene-mature-all-v2.1-no_U.fa"'
echo ""

echo "# External data URLs"
echo 'export mirgenedb_fasta_url="https://www.mirgenedb.org/fasta/ALL?mat=1"'
echo ""

echo "# Paths to programs"
echo 'export ncbi_blast_dir="/home/shared/ncbi-blast-2.15.0+/bin/"'
echo 'export ncbi_blastn="${ncbi_blast_dir}/blastn"'
echo 'export ncbi_makeblast_db="${ncbi_blast_dir}/makeblastdb"'
echo 'export fastx_collapser="/home/shared/fastx_toolkit_0.0.13_binaries_Linux_2.6_amd64/bin/fastx_collapser"'

echo "# Set number of CPUs to use"
echo 'export threads=40'
echo ""

echo "# Initialize arrays"
echo 'export trimmed_fastqs_array=()'


} > .bashvars

cat .bashvars

#### Assign Variables ####

# Trimmed FastQ naming pattern
export trimmed_fastqs_pattern='*fastp-adapters-polyG-31bp-merged.fq.gz'
# Data directories
export deep_dive_dir=/home/shared/8TB_HDD_01/sam/gitrepos/deep-dive
export deep_dive_data_dir="${deep_dive_dir}/data"
export output_dir_top=${deep_dive_dir}/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase
export trimmed_fastqs_dir="${deep_dive_dir}/D-Apul/output/08.2-Apul-sRNAseq-trimming-31bp-fastp-merged/trimmed-reads"
export blast_dbs_dir="${deep_dive_dir}/data/blast_dbs"

# Input/Output files
export collapsed_reads_fasta="collapsed-reads-all.fasta"
export concatenated_trimmed_reads_fastq="concatenated-trimmed-reads-all.fastq.gz"
export mirbase_mature_fasta_name="cnidarian-mirbase-mature-v22.1.fasta"
export mirbase_mature_fasta_no_U="cnidarian-mirbase-mature-v22.1-no_U.fa"
export mirgene_mature_fasta_name="mirgene-mature-all-v2.1.fa"
export mirgene_mature_fasta_no_U="mirgene-mature-all-v2.1-no_U.fa"

# External data URLs
export mirgenedb_fasta_url="https://www.mirgenedb.org/fasta/ALL?mat=1"

# Paths to programs
export ncbi_blast_dir="/home/shared/ncbi-blast-2.15.0+/bin/"
export ncbi_blastn="${ncbi_blast_dir}/blastn"
export ncbi_makeblast_db="${ncbi_blast_dir}/makeblastdb"
export fastx_collapser="/home/shared/fastx_toolkit_0.0.13_binaries_Linux_2.6_amd64/bin/fastx_collapser"
# Set number of CPUs to use
export threads=40

# Initialize arrays
export trimmed_fastqs_array=()

2 Download MirGeneDB Fasta

# Load bash variables into memory
source .bashvars

# Download MirGeneDB, if it doesn't exist
if [ ! -f "${deep_dive_data_dir}/${mirgene_mature_fasta_name}" ]; then

  wget \
  --no-check-certificate \
  --continue \
  --no-host-directories \
  --no-directories \
  --no-parent \
  --quiet \
  --execute robots=off \
  --output-document ${deep_dive_data_dir}/${mirgene_mature_fasta_name} \
   ${mirgenedb_fasta_url}
 
 fi
 
 ls -lh ${deep_dive_data_dir}

total 20M
drwxr-xr-x 2 sam sam 4.0K Apr  2 07:01 blast_dbs
-rw-rw-r-- 1 sam sam 3.8M Apr  2 06:51 cnidarian-mirbase-mature-v22.1.fasta
-rw-r--r-- 1 sam sam 3.8M Apr  2 07:00 cnidarian-mirbase-mature-v22.1-no_U.fa
-rw-r--r-- 1 sam sam  44K Apr  2 06:49 cnidarian_miRNAs.fasta
-rw-r--r-- 1 sam sam 3.7M Feb 16 12:36 mirbase-mature-v22.1.fa
-rw-r--r-- 1 sam sam 3.7M Dec  4 11:05 mirbase-mature-v22.1-no_spaces.fa
-rw-r--r-- 1 sam sam 3.7M Nov 17 07:59 mirbase-mature-v22.1-no_U.fa
-rw-r--r-- 1 sam sam 726K Nov  7 14:36 mirgene-mature-all-v2.1.fa
-rw-r--r-- 1 sam sam 726K Nov 17 07:59 mirgene-mature-all-v2.1-no_U.fa

2.1 Inspect miRNA FastAs

# Load bash variables into memory
source .bashvars

head "${deep_dive_data_dir}"/[cm][ni]*.fa*

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/cnidarian-mirbase-mature-v22.1.fasta <==
>cel-let-7-5p MIMAT0000001 Caenorhabditis elegans let-7-5p
UGAGGUAGUAGGUUGUAUAGUU
>cel-let-7-3p MIMAT0015091 Caenorhabditis elegans let-7-3p
CUAUGCAAUUUUCUACCUUACC
>cel-lin-4-5p MIMAT0000002 Caenorhabditis elegans lin-4-5p
UCCCUGAGACCUCAAGUGUGA
>cel-lin-4-3p MIMAT0015092 Caenorhabditis elegans lin-4-3p
ACACCUGGGCUCUCCGGGUACC
>cel-miR-1-5p MIMAT0020301 Caenorhabditis elegans miR-1-5p
CAUACUUCCUUACAUGCCCAUA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/cnidarian-mirbase-mature-v22.1-no_U.fa <==
>cel-let-7-5p MIMAT0000001 Caenorhabditis elegans let-7-5p
TGAGGTAGTAGGTTGTATAGTT
>cel-let-7-3p MIMAT0015091 Caenorhabditis elegans let-7-3p
CTATGCAATTTTCTACCTTACC
>cel-lin-4-5p MIMAT0000002 Caenorhabditis elegans lin-4-5p
TCCCTGAGACCTCAAGTGTGA
>cel-lin-4-3p MIMAT0015092 Caenorhabditis elegans lin-4-3p
ACACCTGGGCTCTCCGGGTACC
>cel-miR-1-5p MIMAT0020301 Caenorhabditis elegans miR-1-5p
CATACTTCCTTACATGCCCATA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/cnidarian_miRNAs.fasta <==
>spi-mir-temp-1_Stylophora_pistillata_Liew_et_al._2014_Matches_miR-100_family.
ACCCGUAGAUCCGAACUUGUGG
>spi-mir-temp-2_Stylophora_pistillata_Liew_et_al._2014_NA
UAUCGAAUCCGUCAAAAAGAGA
>spi-mir-temp-3_Stylophora_pistillata_Liew_et_al._2014_NA
UCAGGGAUUGUGGUGAGUUAGUU
>spi-mir-temp-4_Stylophora_pistillata_Liew_et_al._2014_Exact_match_of_nve-miR-2023.
AAAGAAGUACAAGUGGUAGGG
>spi-mir-temp-5_Stylophora_pistillata_Liew_et_al._2014_NA
GAGGUCCGGAUGGUUGA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirbase-mature-v22.1.fa <==
>cel-let-7-5p MIMAT0000001 Caenorhabditis elegans let-7-5p
UGAGGUAGUAGGUUGUAUAGUU
>cel-let-7-3p MIMAT0015091 Caenorhabditis elegans let-7-3p
CUAUGCAAUUUUCUACCUUACC
>cel-lin-4-5p MIMAT0000002 Caenorhabditis elegans lin-4-5p
UCCCUGAGACCUCAAGUGUGA
>cel-lin-4-3p MIMAT0015092 Caenorhabditis elegans lin-4-3p
ACACCUGGGCUCUCCGGGUACC
>cel-miR-1-5p MIMAT0020301 Caenorhabditis elegans miR-1-5p
CAUACUUCCUUACAUGCCCAUA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirbase-mature-v22.1-no_spaces.fa <==
>cel-let-7-5p_MIMAT0000001_Caenorhabditis_elegans_let-7-5p
UGAGGUAGUAGGUUGUAUAGUU
>cel-let-7-3p_MIMAT0015091_Caenorhabditis_elegans_let-7-3p
CUAUGCAAUUUUCUACCUUACC
>cel-lin-4-5p_MIMAT0000002_Caenorhabditis_elegans_lin-4-5p
UCCCUGAGACCUCAAGUGUGA
>cel-lin-4-3p_MIMAT0015092_Caenorhabditis_elegans_lin-4-3p
ACACCUGGGCUCUCCGGGUACC
>cel-miR-1-5p_MIMAT0020301_Caenorhabditis_elegans_miR-1-5p
CAUACUUCCUUACAUGCCCAUA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirbase-mature-v22.1-no_U.fa <==
>cel-let-7-5p MIMAT0000001 Caenorhabditis elegans let-7-5p
TGAGGTAGTAGGTTGTATAGTT
>cel-let-7-3p MIMAT0015091 Caenorhabditis elegans let-7-3p
CTATGCAATTTTCTACCTTACC
>cel-lin-4-5p MIMAT0000002 Caenorhabditis elegans lin-4-5p
TCCCTGAGACCTCAAGTGTGA
>cel-lin-4-3p MIMAT0015092 Caenorhabditis elegans lin-4-3p
ACACCTGGGCTCTCCGGGTACC
>cel-miR-1-5p MIMAT0020301 Caenorhabditis elegans miR-1-5p
CATACTTCCTTACATGCCCATA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirgene-mature-all-v2.1.fa <==
>Aae-Bantam_3p
UGAGAUCAUUUUGAAAGCUGAUU
>Bge-Bantam_3p
UGAGAUCAUUGUGAAAGCUGAUU
>Bpl-Bantam_3p
UGAGAUCAUUGUGAAAACUGAU
>Cgi-Bantam_3p
UGAGAUCAUUGUGAAAACUGAUU
>Cte-Bantam_3p
UGAGAUCAUUGUGAAAACUAAUC

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirgene-mature-all-v2.1-no_U.fa <==
>Aae-Bantam_3p
TGAGATCATTTTGAAAGCTGATT
>Bge-Bantam_3p
TGAGATCATTGTGAAAGCTGATT
>Bpl-Bantam_3p
TGAGATCATTGTGAAAACTGAT
>Cgi-Bantam_3p
TGAGATCATTGTGAAAACTGATT
>Cte-Bantam_3p
TGAGATCATTGTGAAAACTAATC

3 Convert `U` to `T` in miRNA FastAs

This is needed because the sRNAseq sequences do not have uracils (U) - they have thymines (T).

# Load bash variables into memory
source .bashvars

# Convert miRBase FastA
if [ ! -f "${deep_dive_data_dir}/${mirbase_mature_fasta_no_U}" ]; then
  sed '/^[^>]/s/U/T/g' "${deep_dive_data_dir}/${mirbase_mature_fasta_name}" \
  > "${deep_dive_data_dir}/${mirbase_mature_fasta_no_U}"
fi


# Convert MirGene FastA
if [ ! -f "${deep_dive_data_dir}/${mirgene_mature_fasta_no_U}" ]; then
  sed '/^[^>]/s/U/T/g' "${deep_dive_data_dir}/${mirgene_mature_fasta_name}" \
  > "${deep_dive_data_dir}/${mirgene_mature_fasta_no_U}"
fi


head "${deep_dive_data_dir}"/[cm][ni]*U.fa

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/cnidarian-mirbase-mature-v22.1-no_U.fa <==
>cel-let-7-5p MIMAT0000001 Caenorhabditis elegans let-7-5p
TGAGGTAGTAGGTTGTATAGTT
>cel-let-7-3p MIMAT0015091 Caenorhabditis elegans let-7-3p
CTATGCAATTTTCTACCTTACC
>cel-lin-4-5p MIMAT0000002 Caenorhabditis elegans lin-4-5p
TCCCTGAGACCTCAAGTGTGA
>cel-lin-4-3p MIMAT0015092 Caenorhabditis elegans lin-4-3p
ACACCTGGGCTCTCCGGGTACC
>cel-miR-1-5p MIMAT0020301 Caenorhabditis elegans miR-1-5p
CATACTTCCTTACATGCCCATA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirbase-mature-v22.1-no_U.fa <==
>cel-let-7-5p MIMAT0000001 Caenorhabditis elegans let-7-5p
TGAGGTAGTAGGTTGTATAGTT
>cel-let-7-3p MIMAT0015091 Caenorhabditis elegans let-7-3p
CTATGCAATTTTCTACCTTACC
>cel-lin-4-5p MIMAT0000002 Caenorhabditis elegans lin-4-5p
TCCCTGAGACCTCAAGTGTGA
>cel-lin-4-3p MIMAT0015092 Caenorhabditis elegans lin-4-3p
ACACCTGGGCTCTCCGGGTACC
>cel-miR-1-5p MIMAT0020301 Caenorhabditis elegans miR-1-5p
CATACTTCCTTACATGCCCATA

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/data/mirgene-mature-all-v2.1-no_U.fa <==
>Aae-Bantam_3p
TGAGATCATTTTGAAAGCTGATT
>Bge-Bantam_3p
TGAGATCATTGTGAAAGCTGATT
>Bpl-Bantam_3p
TGAGATCATTGTGAAAACTGAT
>Cgi-Bantam_3p
TGAGATCATTGTGAAAACTGATT
>Cte-Bantam_3p
TGAGATCATTGTGAAAACTAATC

4 Create BLAST Databases

# Load bash variables into memory
source .bashvars



# miRBase BLAST DB
## Make sure output directory exists
if [ ! -d "${blast_dbs_dir}" ]; then
  mkdir --parents "${blast_dbs_dir}"
fi

## Check for pre-exising database
if [ ! -f "${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*}.blastdb.log" ]; then
  ${ncbi_makeblast_db} \
  -in ${deep_dive_data_dir}/${mirgene_mature_fasta_no_U} \
  -title ${mirgene_mature_fasta_no_U%.*} \
  -dbtype nucl \
  -out ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
  -logfile ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*}.blastdb.log
fi

# miRBase BLAST DB
## Make sure output directory exists
if [ ! -d "${blast_dbs_dir}" ]; then
  mkdir --parents "${blast_dbs_dir}"
fi

## Check for pre-exising database
if [ ! -f "${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*}.blastdb.log" ]; then
  ${ncbi_makeblast_db} \
  -in ${deep_dive_data_dir}/${mirbase_mature_fasta_no_U} \
  -title ${mirbase_mature_fasta_no_U%.*} \
  -dbtype nucl \
  -out ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
  -logfile ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*}.blastdb.log
fi

5 Prepare reads for BLASTing

5.1 Concatenate all trimmed reads

# Load bash variables into memory
source .bashvars

# Make output directory, if it doens't exist
if [ ! -d "${output_dir_top}" ]; then
  mkdir --parents "${output_dir_top}"
fi

# Check for existence of concatenated FastA before running
if [ ! -f "${output_dir_top}/${concatenated_trimmed_reads_fastq}" ]; then
  cat ${trimmed_fastqs_dir}/${trimmed_fastqs_pattern} \
  > "${output_dir_top}/${concatenated_trimmed_reads_fastq}"
fi

ls -lh "${output_dir_top}/${concatenated_trimmed_reads_fastq}"

-rw-r--r-- 1 sam sam 1.6G Apr  1 14:03 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/concatenated-trimmed-reads-all.fastq.gz

5.2 Collapse reads to FastA

Uses fastx_collapser to collapse to unique reads.

Requires undocumented quality setting. Have selected 30 as cuttoff: -Q30.

# Load bash variables into memory
source .bashvars

# Check for existence of collapsed FastA before running
time \
if [ ! -f "${output_dir_top}/${collapsed_reads_fasta}" ]; then
  zcat ${output_dir_top}/${concatenated_trimmed_reads_fastq} \
  | ${fastx_collapser} \
  -Q30 \
  -o "${output_dir_top}/${collapsed_reads_fasta}"
fi

head "${output_dir_top}/${collapsed_reads_fasta}"

wc -l "${output_dir_top}/${collapsed_reads_fasta}"


real    0m0.000s
user    0m0.000s
sys 0m0.000s
>1-1843196
GCACTGGTGGTTCAGTGGTAGAATTCTCGCC
>2-1083086
TGAAAATCTTTCTTCTGAGGCGGAA
>3-967720
GCACTGGTGGTTCAGTGGTAGAATTCTCGCT
>4-832824
TCCAAGACCTGAGTTACCACTGACAAGC
>5-750080
GCACTGTGGTTCAGTGGTAGAATTCTCGCC
22165834 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/collapsed-reads-all.fasta

6 Run BLASTn Default E-value

1000 for blastn-short

Runs BLASTn using the blastn-short task for sequences < 30bp.

Look for top match (-max_hsps 1 & -max_target_seqs 1) for each query.

Suppress subsequent warning Examining 5 or more matches is recommended by redirecting stdout: 2> /dev/null

6.1 miRBase BLASTn Default e-value

# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/cnidarian-miRBase-BLASTn-eval_1000.outfmt6 \
-task blastn-short \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null

6.2 MirGene BLASTn Default e-value

# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/MirGene-BLASTn-eval_1000.outfmt6 \
-task blastn-short \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null

7 BLASTn E-value = 10

Running this for simple comparison to the default blastn-short value of 1000.

7.1 miRBase BLASTn e-value = 10

# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/cindarian-miRBase-BLASTn-eval_10.outfmt6 \
-task blastn-short \
-evalue 10 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null

7.2 MirGene BLASTn e-value = 10

# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/MirGene-BLASTn-eval_10.outfmt6 \
-task blastn-short \
-evalue 10 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null

8 BLASTn E-value = 1

Running this for simple comparison to the default blastn-short value of 1000.

8.1 miRBase BLASTn e-value = 1

# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/cindarian-miRBase-BLASTn-eval_1.outfmt6 \
-task blastn-short \
-evalue 1 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null

8.2 MirGene BLASTn e-value = 1

# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/MirGene-BLASTn-eval_1.outfmt6 \
-task blastn-short \
-evalue 1 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null

9 Results

9.1 Check BLASTn Default e-value results

# Load bash variables into memory
source .bashvars

head ${output_dir_top}/*eval_1000.outfmt6

echo ""
echo "-----------------------------------------"
echo ""

wc -l ${output_dir_top}/*eval_1000.outfmt6

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/cnidarian-miRBase-BLASTn-eval_1000.outfmt6 <==
1-1843196   ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
2-1083086   ppy-miR-1255a   100.000 12  0   0   3   14  22  11  0.37    24.3
3-967720    ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
4-832824    dme-miR-4971-5p 100.000 12  0   0   17  28  18  7   0.45    24.3
5-750080    ppc-miR-8214-5p 100.000 13  0   0   11  23  18  6   0.13    26.3
6-691325    ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
7-659957    ami-miR-2184-5p 100.000 11  0   0   4   14  12  22  2.0 22.3
8-606529    cpo-miR-509c-3p 100.000 12  0   0   13  24  8   19  0.37    24.3
9-571297    adi-nve-F-miR-100_Acropora_digitifera__Praher_et_al._2021_NA    100.000 20  0   0   1   20  1   20  3.98e-06    40.1
10-528956   egr-miR-10227d-5p   100.000 11  0   0   14  24  20  10  1.7 22.3

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/MirGene-BLASTn-eval_1000.outfmt6 <==
1-1843196   Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
2-1083086   Hsa-Mir-3611_3p 100.000 9   0   0   7   15  13  5   8.5 18.3
3-967720    Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
4-832824    Mml-Mir-3200_3p 100.000 10  0   0   5   14  21  12  2.6 20.3
5-750080    Pma-Mir-96-P3o2_5p  100.000 11  0   0   15  25  9   19  0.74    22.3
6-691325    Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.66    22.3
7-659957    Cpi-Mir-2184_5p 100.000 11  0   0   4   14  12  22  0.74    22.3
8-606529    Cpo-Mir-506-o3_3p   100.000 12  0   0   13  24  8   19  0.14    24.3
9-571297    Csc-Mir-10-P2r2_5p  100.000 19  0   0   2   20  3   21  7.12e-06    38.2
10-528956   Xla-Novel-4-P2_5p   100.000 10  0   0   10  19  10  1   2.5 20.3

-----------------------------------------

  11082917 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/cnidarian-miRBase-BLASTn-eval_1000.outfmt6
  11082917 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/MirGene-BLASTn-eval_1000.outfmt6
  22165834 total

9.2 Check BLASTn e-value = 10 results

# Load bash variables into memory
source .bashvars

head ${output_dir_top}/*eval_10.outfmt6

echo ""
echo "-----------------------------------------"
echo ""

wc -l ${output_dir_top}/*eval_10.outfmt6

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/cindarian-miRBase-BLASTn-eval_10.outfmt6 <==
1-1843196   ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
2-1083086   ppy-miR-1255a   100.000 12  0   0   3   14  22  11  0.37    24.3
3-967720    ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
4-832824    dme-miR-4971-5p 100.000 12  0   0   17  28  18  7   0.45    24.3
5-750080    ppc-miR-8214-5p 100.000 13  0   0   11  23  18  6   0.13    26.3
6-691325    ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
7-659957    ami-miR-2184-5p 100.000 11  0   0   4   14  12  22  2.0 22.3
8-606529    cpo-miR-509c-3p 100.000 12  0   0   13  24  8   19  0.37    24.3
9-571297    adi-nve-F-miR-100_Acropora_digitifera__Praher_et_al._2021_NA    100.000 20  0   0   1   20  1   20  3.98e-06    40.1
10-528956   egr-miR-10227d-5p   100.000 11  0   0   14  24  20  10  1.7 22.3

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/MirGene-BLASTn-eval_10.outfmt6 <==
1-1843196   Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
2-1083086   Hsa-Mir-3611_3p 100.000 9   0   0   7   15  13  5   8.5 18.3
3-967720    Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
4-832824    Mml-Mir-3200_3p 100.000 10  0   0   5   14  21  12  2.6 20.3
5-750080    Pma-Mir-96-P3o2_5p  100.000 11  0   0   15  25  9   19  0.74    22.3
6-691325    Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.66    22.3
7-659957    Cpi-Mir-2184_5p 100.000 11  0   0   4   14  12  22  0.74    22.3
8-606529    Cpo-Mir-506-o3_3p   100.000 12  0   0   13  24  8   19  0.14    24.3
9-571297    Csc-Mir-10-P2r2_5p  100.000 19  0   0   2   20  3   21  7.12e-06    38.2
10-528956   Xla-Novel-4-P2_5p   100.000 10  0   0   10  19  10  1   2.5 20.3

-----------------------------------------

  11059697 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/cindarian-miRBase-BLASTn-eval_10.outfmt6
  10189571 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/MirGene-BLASTn-eval_10.outfmt6
  21249268 total

9.3 Check BLASTn e-value = 1 results

# Load bash variables into memory
source .bashvars

head ${output_dir_top}/*eval_1.outfmt6

echo ""
echo "-----------------------------------------"
echo ""

wc -l ${output_dir_top}/*eval_1.outfmt6

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/cindarian-miRBase-BLASTn-eval_1.outfmt6 <==
1-1843196   ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
2-1083086   ppy-miR-1255a   100.000 12  0   0   3   14  22  11  0.37    24.3
3-967720    ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
4-832824    dme-miR-4971-5p 100.000 12  0   0   17  28  18  7   0.45    24.3
5-750080    ppc-miR-8214-5p 100.000 13  0   0   11  23  18  6   0.13    26.3
6-691325    ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3
8-606529    cpo-miR-509c-3p 100.000 12  0   0   13  24  8   19  0.37    24.3
9-571297    adi-nve-F-miR-100_Acropora_digitifera__Praher_et_al._2021_NA    100.000 20  0   0   1   20  1   20  3.98e-06    40.1
11-456706   dme-miR-4971-5p 100.000 12  0   0   17  28  18  7   0.45    24.3
12-455987   ppc-miR-8214-5p 100.000 13  0   0   12  24  18  6   0.11    26.3

==> /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/MirGene-BLASTn-eval_1.outfmt6 <==
1-1843196   Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
3-967720    Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
5-750080    Pma-Mir-96-P3o2_5p  100.000 11  0   0   15  25  9   19  0.74    22.3
6-691325    Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.66    22.3
7-659957    Cpi-Mir-2184_5p 100.000 11  0   0   4   14  12  22  0.74    22.3
8-606529    Cpo-Mir-506-o3_3p   100.000 12  0   0   13  24  8   19  0.14    24.3
9-571297    Csc-Mir-10-P2r2_5p  100.000 19  0   0   2   20  3   21  7.12e-06    38.2
12-455987   Pma-Mir-96-P3o2_5p  100.000 11  0   0   16  26  9   19  0.78    22.3
13-415294   Pma-Mir-96-P3o2_5p  100.000 11  0   0   15  25  9   19  0.74    22.3
14-383875   Nve-Mir-10_5p   100.000 21  0   0   2   22  2   22  5.47e-07    42.1

-----------------------------------------

  5108736 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/cindarian-miRBase-BLASTn-eval_1.outfmt6
  4196511 /home/shared/8TB_HDD_01/sam/gitrepos/deep-dive/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase/MirGene-BLASTn-eval_1.outfmt6
  9305247 total

The default e-value results in alignments for ~50% of sequences.

Decreasing the e-value to 10 resulted in fewer query alignments, as we’d expect, but only minimally.

Decreasing the e-value to 1 resulted in ~25% of query alignments.

For further analysis, we should probably discuss a reasonable e-value to use for filtering, as it might be worthwhile to test even lower e-value thresholds.

Citations

Altschul, Stephen F., Warren Gish, Webb Miller, Eugene W. Myers, and David J. Lipman. 1990. “Basic Local Alignment Search Tool.” Journal of Molecular Biology 215 (3): 403–10. https://doi.org/10.1016/s0022-2836(05)80360-2.

---
title: "10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase"
author: "Sam White"
date: "2024-03-29"
output: 
  bookdown::html_document2:
    theme: cosmo
    toc: true
    toc_float: true
    number_sections: true
    code_folding: show
    code_download: true
  github_document:
    toc: true
    number_sections: true
  html_document:
    theme: cosmo
    toc: true
    toc_float: true
    number_sections: true
    code_folding: show
    code_download: true
bibliography: references.bib
link-citations: true
---

```{r setup, include=FALSE}
library(knitr)
library(kableExtra)
library(dplyr)
library(reticulate)
knitr::opts_chunk$set(
  echo = TRUE,         # Display code chunks
  eval = FALSE,        # Evaluate code chunks
  warning = FALSE,     # Hide warnings
  message = FALSE,     # Hide messages
  comment = "",        # Prevents appending '##' to beginning of lines in code output
  width = 1000         # adds scroll bar
)
```


This notebook performs a simple [NCBI BLASTn](https://www.ncbi.nlm.nih.gov/books/NBK279690/) [@altschul1990] against two miRNA databases to attempt to identify miRNA in _A.pulchra_ sRNAseq:

- [miRBase](https://mirbase.org/download/)

  - Utilizes a modified version, which includes cnidarian miRNA culled from literature by Jill Ahsley.
  
  - - [`cnidarian-mirbase-mature-v22.1.fasta`](../../data/cnidarian-mirbase-mature-v22.1.fasta)

- [MirGeneDB](https://www.mirgenedb.org/download)

Relies on the following software:
  
- [fastx_toolkit](http://hannonlab.cshl.edu/fastx_toolkit/)

  - `fastx_collapser`: Collapses duplicate sequences in FastA/Q into single sequence.

# Create a Bash variables file

This allows usage of Bash variables across R Markdown chunks.

```{r save-bash-variables-to-rvars-file, engine='bash', eval=TRUE}
{
echo "#### Assign Variables ####"
echo ""

echo "# Trimmed FastQ naming pattern"
echo "export trimmed_fastqs_pattern='*fastp-adapters-polyG-31bp-merged.fq.gz'"

echo "# Data directories"
echo 'export deep_dive_dir=/home/shared/8TB_HDD_01/sam/gitrepos/deep-dive'
echo 'export deep_dive_data_dir="${deep_dive_dir}/data"'
echo 'export output_dir_top=${deep_dive_dir}/D-Apul/output/10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase'
echo 'export trimmed_fastqs_dir="${deep_dive_dir}/D-Apul/output/08.2-Apul-sRNAseq-trimming-31bp-fastp-merged/trimmed-reads"'
echo 'export blast_dbs_dir="${deep_dive_dir}/data/blast_dbs"'
echo ""

echo "# Input/Output files"
echo 'export collapsed_reads_fasta="collapsed-reads-all.fasta"'
echo 'export concatenated_trimmed_reads_fastq="concatenated-trimmed-reads-all.fastq.gz"'
echo 'export mirbase_mature_fasta_name="cnidarian-mirbase-mature-v22.1.fasta"'
echo 'export mirbase_mature_fasta_no_U="cnidarian-mirbase-mature-v22.1-no_U.fa"'
echo 'export mirgene_mature_fasta_name="mirgene-mature-all-v2.1.fa"'
echo 'export mirgene_mature_fasta_no_U="mirgene-mature-all-v2.1-no_U.fa"'
echo ""

echo "# External data URLs"
echo 'export mirgenedb_fasta_url="https://www.mirgenedb.org/fasta/ALL?mat=1"'
echo ""

echo "# Paths to programs"
echo 'export ncbi_blast_dir="/home/shared/ncbi-blast-2.15.0+/bin/"'
echo 'export ncbi_blastn="${ncbi_blast_dir}/blastn"'
echo 'export ncbi_makeblast_db="${ncbi_blast_dir}/makeblastdb"'
echo 'export fastx_collapser="/home/shared/fastx_toolkit_0.0.13_binaries_Linux_2.6_amd64/bin/fastx_collapser"'

echo "# Set number of CPUs to use"
echo 'export threads=40'
echo ""

echo "# Initialize arrays"
echo 'export trimmed_fastqs_array=()'


} > .bashvars

cat .bashvars
```


# Download MirGeneDB Fasta
```{r download-mirgene-db-FastA, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

# Download MirGeneDB, if it doesn't exist
if [ ! -f "${deep_dive_data_dir}/${mirgene_mature_fasta_name}" ]; then

  wget \
  --no-check-certificate \
  --continue \
  --no-host-directories \
  --no-directories \
  --no-parent \
  --quiet \
  --execute robots=off \
  --output-document ${deep_dive_data_dir}/${mirgene_mature_fasta_name} \
   ${mirgenedb_fasta_url}
 
 fi
 
 ls -lh ${deep_dive_data_dir}
```

## Inspect miRNA FastAs
```{r inspect-miRNA-FastAs, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

head "${deep_dive_data_dir}"/[cm][ni]*.fa*
```


# Convert `U` to `T` in miRNA FastAs

This is needed because the sRNAseq sequences do _not_ have uracils (`U`) - they have thymines (`T`).

```{r convert-U-to-T, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

# Convert miRBase FastA
if [ ! -f "${deep_dive_data_dir}/${mirbase_mature_fasta_no_U}" ]; then
  sed '/^[^>]/s/U/T/g' "${deep_dive_data_dir}/${mirbase_mature_fasta_name}" \
  > "${deep_dive_data_dir}/${mirbase_mature_fasta_no_U}"
fi


# Convert MirGene FastA
if [ ! -f "${deep_dive_data_dir}/${mirgene_mature_fasta_no_U}" ]; then
  sed '/^[^>]/s/U/T/g' "${deep_dive_data_dir}/${mirgene_mature_fasta_name}" \
  > "${deep_dive_data_dir}/${mirgene_mature_fasta_no_U}"
fi


head "${deep_dive_data_dir}"/[cm][ni]*U.fa
  
```

# Create BLAST Databases
```{r make-blast-dbs, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars



# miRBase BLAST DB
## Make sure output directory exists
if [ ! -d "${blast_dbs_dir}" ]; then
  mkdir --parents "${blast_dbs_dir}"
fi

## Check for pre-exising database
if [ ! -f "${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*}.blastdb.log" ]; then
  ${ncbi_makeblast_db} \
  -in ${deep_dive_data_dir}/${mirgene_mature_fasta_no_U} \
  -title ${mirgene_mature_fasta_no_U%.*} \
  -dbtype nucl \
  -out ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
  -logfile ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*}.blastdb.log
fi

# miRBase BLAST DB
## Make sure output directory exists
if [ ! -d "${blast_dbs_dir}" ]; then
  mkdir --parents "${blast_dbs_dir}"
fi

## Check for pre-exising database
if [ ! -f "${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*}.blastdb.log" ]; then
  ${ncbi_makeblast_db} \
  -in ${deep_dive_data_dir}/${mirbase_mature_fasta_no_U} \
  -title ${mirbase_mature_fasta_no_U%.*} \
  -dbtype nucl \
  -out ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
  -logfile ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*}.blastdb.log
fi
```


# Prepare reads for BLASTing

## Concatenate all trimmed reads
```{r concatenate-trimmed-reads, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

# Make output directory, if it doens't exist
if [ ! -d "${output_dir_top}" ]; then
  mkdir --parents "${output_dir_top}"
fi

# Check for existence of concatenated FastA before running
if [ ! -f "${output_dir_top}/${concatenated_trimmed_reads_fastq}" ]; then
  cat ${trimmed_fastqs_dir}/${trimmed_fastqs_pattern} \
  > "${output_dir_top}/${concatenated_trimmed_reads_fastq}"
fi

ls -lh "${output_dir_top}/${concatenated_trimmed_reads_fastq}"
```

## Collapse reads to FastA

Uses `fastx_collapser` to collapse to unique reads.

Requires undocumented quality setting. Have selected `30` as cuttoff: `-Q30`.
```{r collapse-reads-to-FastA, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

# Check for existence of collapsed FastA before running
time \
if [ ! -f "${output_dir_top}/${collapsed_reads_fasta}" ]; then
  zcat ${output_dir_top}/${concatenated_trimmed_reads_fastq} \
  | ${fastx_collapser} \
  -Q30 \
  -o "${output_dir_top}/${collapsed_reads_fasta}"
fi

head "${output_dir_top}/${collapsed_reads_fasta}"

wc -l "${output_dir_top}/${collapsed_reads_fasta}"
```

# Run BLASTn Default E-value

- 1000 for blastn-short

Runs BLASTn using the `blastn-short` task for sequences < 30bp.

Look for top match (`-max_hsps 1` & `-max_target_seqs 1`) for each query.

  - Suppress subsequent warning `Examining 5 or more matches is recommended` by
  redirecting stdout: `2> /dev/null`

## miRBase BLASTn Default e-value
```{r miRBase-BLASTn-default-eval, engine='bash', cache=TRUE}
# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/cnidarian-miRBase-BLASTn-eval_1000.outfmt6 \
-task blastn-short \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null
```

## MirGene BLASTn Default e-value

```{r MirGene-BLASTn-default-eval, engine='bash', cache=TRUE}
# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/MirGene-BLASTn-eval_1000.outfmt6 \
-task blastn-short \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null
```


# BLASTn E-value = 10

Running this for simple comparison to the default `blastn-short` value of 1000.

## miRBase BLASTn e-value = 10
```{r miRBase-BLASTn-10-eval, engine='bash', cache=TRUE}
# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/cindarian-miRBase-BLASTn-eval_10.outfmt6 \
-task blastn-short \
-evalue 10 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null
```

## MirGene BLASTn e-value = 10

```{r MirGene-BLASTn-10-eval, engine='bash', cache=TRUE}
# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/MirGene-BLASTn-eval_10.outfmt6 \
-task blastn-short \
-evalue 10 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null
```


# BLASTn E-value = 1

Running this for simple comparison to the default `blastn-short` value of 1000.

## miRBase BLASTn e-value = 1
```{r miRBase-BLASTn-1-eval, engine='bash', cache=TRUE}
# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirbase_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/cindarian-miRBase-BLASTn-eval_1.outfmt6 \
-task blastn-short \
-evalue 1 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null
```

## MirGene BLASTn e-value = 1

```{r MirGene-BLASTn-1-eval, engine='bash', cache=TRUE}
# Load bash variables into memory
source .bashvars

time \
${ncbi_blastn} \
-db ${blast_dbs_dir}/${mirgene_mature_fasta_no_U%.*} \
-query ${output_dir_top}/${collapsed_reads_fasta} \
-out ${output_dir_top}/MirGene-BLASTn-eval_1.outfmt6 \
-task blastn-short \
-evalue 1 \
-max_hsps 1 \
-max_target_seqs 1 \
-outfmt 6 \
-num_threads ${threads} \
2> /dev/null
```



# Results

## Check BLASTn Default e-value results
```{r blastn-default-evalue-results, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

head ${output_dir_top}/*eval_1000.outfmt6

echo ""
echo "-----------------------------------------"
echo ""

wc -l ${output_dir_top}/*eval_1000.outfmt6
```

## Check BLASTn e-value = 10 results
```{r blastn-10-evalue-results, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

head ${output_dir_top}/*eval_10.outfmt6

echo ""
echo "-----------------------------------------"
echo ""

wc -l ${output_dir_top}/*eval_10.outfmt6
```

## Check BLASTn e-value = 1 results
```{r blastn-1-evalue-results, engine='bash', eval=TRUE}
# Load bash variables into memory
source .bashvars

head ${output_dir_top}/*eval_1.outfmt6

echo ""
echo "-----------------------------------------"
echo ""

wc -l ${output_dir_top}/*eval_1.outfmt6
```
The default e-value results in alignments for ~50% of sequences.

Decreasing the e-value to `10` resulted in fewer query alignments, as we'd expect, but only minimally.

Decreasing the e-value to `1` resulted in ~25% of query alignments.

For further analysis, we should probably discuss a reasonable e-value to use for filtering, as it might be worthwhile to test even lower e-value thresholds.

------------------------------------------------------------------------

# Citations


10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase

Sam White

2024-03-29

1 Create a Bash variables file

2 Download MirGeneDB Fasta

2.1 Inspect miRNA FastAs

3 Convert `U` to `T` in miRNA FastAs

4 Create BLAST Databases

5 Prepare reads for BLASTing

5.1 Concatenate all trimmed reads

5.2 Collapse reads to FastA

6 Run BLASTn Default E-value

6.1 miRBase BLASTn Default e-value

6.2 MirGene BLASTn Default e-value

7 BLASTn E-value = 10

7.1 miRBase BLASTn e-value = 10

7.2 MirGene BLASTn e-value = 10

8 BLASTn E-value = 1

8.1 miRBase BLASTn e-value = 1

8.2 MirGene BLASTn e-value = 1

9 Results

9.1 Check BLASTn Default e-value results

9.2 Check BLASTn e-value = 10 results

9.3 Check BLASTn e-value = 1 results

Citations

10.1-Apul-sRNAseq-BLASTn-31bp-fastp-merged-cnidarian_miRBase

Sam White

2024-03-29

1 Create a Bash variables file

2 Download MirGeneDB Fasta

2.1 Inspect miRNA FastAs

3 Convert U to T in miRNA FastAs

4 Create BLAST Databases

5 Prepare reads for BLASTing

5.1 Concatenate all trimmed reads

5.2 Collapse reads to FastA

6 Run BLASTn Default E-value

6.1 miRBase BLASTn Default e-value

6.2 MirGene BLASTn Default e-value

7 BLASTn E-value = 10

7.1 miRBase BLASTn e-value = 10

7.2 MirGene BLASTn e-value = 10

8 BLASTn E-value = 1

8.1 miRBase BLASTn e-value = 1

8.2 MirGene BLASTn e-value = 1

9 Results

9.1 Check BLASTn Default e-value results

9.2 Check BLASTn e-value = 10 results

9.3 Check BLASTn e-value = 1 results

Citations

3 Convert `U` to `T` in miRNA FastAs