Raw reads
We retrieved four 12 files from the iSeqNovaSeq runs. One Four of these contains contain the 1x100bp reads. I am not sure why we received 2 index reads for each run. The file files with the raw reads of interest are ( will be in: /project/gtl/seq/distribution/trout1/Trout1_*/rawreads
).
Swap out “distribution” for “raw” for starting directory.
Trout1_P1_S1_L001_R1_001.fastq.gz (24G) – 457,726,974 reads (1.5 GBytes uncompressed)
Trout1_P2_S2_L002_R1_001.fastq.gz (25G) – 457,726,974 reads (1.5 GBytes uncompressed)
Trout1_P3_S1_L001_R1_001.fastq.gz (39G) – 457,726,974 reads (1.5 GBytes uncompressed)
Trout1_P4_S2_L002_R1_001.fastq.gz (40G) – 457,726,974 reads (1.5 GBytes uncompressed)
Unzip and split
cd /gscratch/grandol1/
mkdir Trout1
cd Trout1
gunzip /project/gtl/seq/raw/trout1/Trout1_1/rawreads/Trout1_P1_S1_L001_R1_001.fastq.gz
...
gunzip /project/gtl/seq/raw/trout1/Trout1_4/rawreads/Trout1_P4_S2_L002_R1_001.fastq.gz
Needs updating below 4-26-22
Demultiplexing
Split into 100000 line files
Code Block |
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mkdir /gscratch/grandol1/trout1
cd /gscratch/grandol1/trout1
cat /project/microbiome/data_queue/seq/trout1/Trout1_Pool3_S1_L001_R1_001.fastq | split -l 1000000 -d --suffix-length=3 --additional-suffix=.fastq - Trout1_Pool3_
|
Remove underscores and extraneous spaces
cat split -l 10000000 -d --suffix-length=3 --additional-suffix=.fastq - Trout1_Pool1_
cat split -l 10000000 -d --suffix-length=3 --additional-suffix=.fastq - Trout1_Pool2_
cat split -l 10000000 -d --suffix-length=3 --additional-suffix=.fastq - Trout1_Pool3_
cat split -l 10000000 -d --suffix-length=3 --additional-suffix=.fastq - Trout1_Pool14
Demultiplexing
Remove underscores and extraneous spaces
cd /project/gtl/seq/raw/trout1/Trout1_1/rawreads
sed 's/_/-/' Trout1Pool1_Demux.csv > Trout1Pool1_Demux1.csv
sed -E 's/^([[:alnum:]-]+),([[:alnum:]-]+),([[:alnum:]-]+).*/\1,\2,\3/' Trout1Pool1_Demux1.csv > Trout1Pool1_Demux_fixed.csv
Remove underscores and extraneous spaces
cd /project/gtl/seq/raw/trout1/Trout1_2/rawreads
sed 's/_/-/' Trout1Pool2_Demux.csv > Trout1Pool2_Demux1.csv
sed -E 's/^([[:alnum:]-]+),([[:alnum:]-]+),([[:alnum:]-]+).*/\1,\2,\3/' Trout1Pool2_Demux1.csv > Trout1Pool2_Demux_fixed.csv
Remove underscores and extraneous spaces
cd /project/gtl/seq/raw/trout1/Trout1_3/rawreads
sed 's/_/-/' Trout1Pool3_Demux.csv > Trout1Pool3_Demux1.csv
sed -E 's/^([[:alnum:]-]+),([[:alnum:]-]+),([[:alnum:]-]+).*/\1,\2,\3/' Trout1Pool3_Demux1.csv > Trout1Pool3_Demux_fixed.csv
Remove underscores and extraneous spaces
cd /project/gtl/seq/raw/trout1/Trout1_4/rawreads
sed 's/_/-/' Trout1Pool4_Demux.csv > Trout1Pool4_Demux1.csv
sed -E 's/^([[:alnum:]-]+),([[:alnum:]-]+),([[:alnum:]-]+).*/\1,\2,\3/' Trout1Pool4_Demux1.csv > Trout1Pool4_Demux_fixed.csv
cd /gscratch/grandol1/
Parse split files
/project/gtl/analyses/gtl/HMAX1seq/raw/trout1/Trout1_1/demultiplex/run_parsebarcodes_onSplitInput.pl
/project/gtl/seq/raw/trout1/Trout1_2/demultiplex/run_parsebarcodes_onSplitInput.pl
/project/gtl/seq/raw/trout1/Trout1_3/demultiplex/run_parsebarcodes_onSplitInput.pl
/project/gtl/seq/raw/trout1/Trout1_4/demultiplex/run_parsebarcodes_onSplitInput.pl
Recombine by sample name and mid
cd /project/gtl/seq/raw/trout1/Trout1_1/demultiplex
./run_splitFastq_gbs.sh
cd /project/gtl/seq/raw/trout1/Trout1_2/demultiplex
./run_splitFastq_gbs.sh
cd /project/gtl/seq/raw/trout1/Trout1_3/demultiplex
./run_splitFastq_gbs.sh
cd /project/gtl/seq/raw/trout1/Trout1_4/demultiplex
./run_splitFastq_gbs.sh