ONT_pilot_w_GDB_samples.md

@@ -565,15 +565,102 @@ mkdir slurm_files
 mv slurm* slurm_files/.
 ```
 
-##############################
-# Notes  - for 2018_GDB data #
-##############################
+## Chapter XII - Work(s) in Progress
+##### CIRCULATORY POTENTIAL #####
+- This is where the brain doesn't know when to shut off, and we went on a tanget
+- The question being - does flye or metaspades_hybrid allow for circulisation of the contigs, especially for binning
+```
+############
+# CIRCULAR #           "TO-DO"
+############
+# checking how many contigs in each bin
+cd /scratch/users/sbusi/ONT/cedric_ont_basecalling/results/Binning
+# made a list with the following. excluded 'flye' 'cos bins are not real
+bwa_lr_metaspades
+bwa_merged_metaspades
+bwa_sr_metaspades
+mmi_lr_metaspades
+mmi_merged_metaspades
+mmi_sr_metaspades
+megahit
+
+for i in `cat list`
+do 
+    for file in "$i"/dastool_output/"$i"_DASTool_bins/*.fa
+    do
+        echo "$file"
+        grep -c '>' "$file"
+    done >> "$i"_bins_contigs
+done 
+# None of the bins came from a single contig
+
+# checking the circularity of the bins
+cd /scratch/users/sbusi/ONT/cedric_ont_basecalling/
+mkdir circular
+cd circular
+
+conda activate circlator
+conda install canu
+circlator all ../results/assembly/flye/lr/merged/barcode07/assembly.fna ../results/basecalled/merged/barcode07.fna flye_circlator_output
+
+# failing in 10-hour interactive so submitted as bash script
+sbatch -p bigmem -t 2-00:00:00 -N 1 -n 24 run_circlator.sh (OR) sbatch -p bigmem -t 0-16:00:00 -N 1 -n 24 run_circlator.sh      # failed after a while, so going to manual methods below
+
+# Checking graph with Bandage
+conda activate bandage
+Bandage info ../results/assembly/metaspades_hybrid/lr_barcode07-sr_NEB2_MG_S17/assembly_graph_with_scaffolds.gfa
+
+# removing singletons == DOES NOT WORK yet..
+cd /scratch/users/sbusi/ONT/cedric_ont_basecalling/circular
+ml swenv/bioinfo-env/v0.1-20170602-production
+ml swenv/default-env/v0.1-20170602-production
+ml bio/BioPerl/1.7.1-foss-2017a-Perl-5.24.1
+
+perl -0076 -ne ‘{chomp; m/(.*):(.*);/; print “>$_” if $1 ne $2};’ < /scratch/users/sbusi/ONT/cedric_ont_basecalling/results/assembly/metaspades_hybrid/lr_barcode07-sr_NEB2_MG_S17/assembly_graph.fastg > megahit_singleton_removed.fastg
+```
+
+##### GraphBin #####
+- https://github.com/Vini2/GraphBin     - for binning from graphs
+- downloaded the "methylation-aware" flye assembly and .gfa files to desktop
+- also downloaded the flye_maxbin_scaffolds2bin.tsv and saved as .csv
+- working on the desktop (MAC)
+```
+cd ~/Documents/Nanopore_ONT
+git clone https://github.com/Vini2/GraphBin.git
+cd GraphBin/
+conda env create -f environment.yml
+conda activate graphbin
+
+python graphbin.py --assembler flye --graph assembly_graph.gfa --paths assembly.fasta \
+  --binned flye_maxbin.scaffolds2bin.csv --output flye_methylation_graphbin --max_iteration 10
+
+# downloaded the spades contigs.fasta, contigs.paths, assembly_graph_with_scaffolds.gfa, and the scaffolds2bin.tsv file from bwa_sr_metaspades_hybrid to desktop
+cd ~/Documents/Nanopore_ONT/GraphBin/spades_input
+conda activate graphbin
+
+# removed the "maxbin_output." from the .tsv file and saved as .csv
+cd /Users/susheel.busi/Documents/Nanopore_ONT/GraphBin
+python graphbin.py --assembler spades --graph spades_input/assembly_graph_with_scaffolds.gfa \
+  --paths spades_input/contigs.paths --binned spades_input/bwa_sr_metaspades_hybrid_maxbin.scaffolds2bin.csv \
+    --output spades_graphbin_output
+```
+
+##### AGB #####
+- Trying AGB: https://almiheenko.github.io/AGB/manual.html
+```
+cd /scratch/users/sbusi/ONT/cedric_ont_basecalling/2019_GDB
+conda create -c almiheenko -c bioconda -n AGB agb
+chmod -R 775 /home/users/sbusi/apps/miniconda3/envs/AGB/
+
+si
+conda activate AGB
+agb.py -i methylation_aware_results/assembly/flye/lr/merged/no_barcode/ -a Flye -o flye_methylation_agb_output
+```
+
+##### Notes  - for 2018_GDB data #####
 1. Flye does not produce metabat bins. Reason == only 2 contigs (less than 1500 bp) have coverages over 1. 
 2. Should we adjust the "minCV" parameter for metabat?
 3. Added the snakemake "--keep-going" or "-k" flag to the launcher to finish the independent jobs, and restarted 
 4. Need to fix checkm and gtdbktk runs
-5. CheckM was throwing file missing erros, so did the following:
-```
-cp /home/users/sbusi/apps/checkm/* /home/users/sbusi/.checkm/. 
-```
+5. CheckM was throwing file missing erros, so did the following: "cp /home/users/sbusi/apps/checkm/* /home/users/sbusi/.checkm/."
 6. Use "checkm data setRoot" to specify the location of CheckM database files.
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