apraga/org - Change CKXX2K6A6RCMJENA3XKDBHO3YULTUDORZ2M5ZXE4UQKIIXHBVIQQC

Two ways to generate notes:

a list generated automatically. Issue: wrong title
a hand made list of HTML link. Issue: hard to maintain

Created by Alexis Praga on May 29, 2023

CKXX2K6A6RCMJENA3XKDBHO3YULTUDORZ2M5ZXE4UQKIIXHBVIQQC

Dependencies

In channels

main

Change contents

Deletion in src/Main.hs at line 41 [7.9907]
B:BD[6.221] → [6.221:222]

Replacement in src/Main.hs at line 42 [7.9907]

B:BD[6.223] → [6.223:265]

    match "notes/*bacteriologie.org" $ do

[6.223]

[8.816]

    match ("notes/*microbio*.org" .||. "notes/medecine/202*.org") $ do

Replacement in src/Main.hs at line 45 [7.9907]

B:BD[9.1229] → [9.1229:1369]

            >>= loadAndApplyTemplate "templates/post.html"    postCtx
            >>= loadAndApplyTemplate "templates/default.html" postCtx

[9.1229]

[9.1369]

            -- >>= loadAndApplyTemplate "templates/post.html"    defaultContext
            >>= loadAndApplyTemplate "templates/default.html" defaultContext

Insertion in src/Main.hs at line 51 [7.9907]

[10.1591]

[11.433]

    create ["notes/medecine.html"] $ do
        route idRoute
        compile $ do
            notes <- loadAll "notes/medecine/*"
            let notesCtx =
                    listField "notes" defaultContext (return notes) `mappend`
                    constField "title" "Notes"            `mappend`
                    defaultContext
            makeItem ""
                >>= loadAndApplyTemplate "templates/notes.html" notesCtx
                >>= loadAndApplyTemplate "templates/default.html" notesCtx
                >>= relativizeUrls
    -- Don't forget to set the path to temporary files

Deletion in src/Main.hs at line 94 [7.9907]

B:BD[12.2118] → [12.2118:2119]

B:BD[12.2119] → [13.370:689]


    -- -- Don't forget to set the path to temporary files
    -- match "genetique.html" $ do
    --     route idRoute
    --     compile $ do
    --         posts <- loadAll "../genetique/*.org"
    --         let indexCtx = listField "notes" defaultContext (return posts)
    --               `mappend` defaultContext

Deletion in src/Main.hs at line 95 [7.9907]

B:BD[7.12186] → [13.690:885]

∅:D[13.885] → [7.12300:12301]

∅:D[9.2432] → [7.12300:12301]

B:BD[7.12300] → [7.12300:12301]

    --         getResourceBody
    --             >>= applyAsTemplate indexCtx
    --             >>= loadAndApplyTemplate "templates/default.html" indexCtx
    --             >>= relativizeUrls

Deletion in src/Main.hs at line 97 [7.9907]
∅:D[10.2082] → [7.12952:12953]
B:BD[7.12952] → [7.12952:12953]

Replacement in projects.org at line 116 [15.123895]

B:BD[14.562] → [3.12:92]

** TODO XAM.jl: PR pour modification record :julia:
SCHEDULED: <2023-05-28 Sun>

[16.28]

[3.92]

** KILL XAM.jl: PR pour modification record :julia:
CLOSED: [2023-05-29 Mon 15:40] SCHEDULED: <2023-05-28 Sun>

Insertion in projects.org at line 119 [15.123895]

[3.128]

[17.8973]

** TODO XAMscissors.jl
:PROPERTIES:
:ID:       966a298c-948a-4694-a6f5-c326b1046a05
:END:
Modification de la séquence dans BAM.
*Pas de mise à jour de CIGAR*
On convertit en fastq et on lance le pipeline pour "corriger"
#+begin_src sh
cd /home/alex/code/bisonex/out/63003856/preprocessing/mapped
samtools view 63003856_S135.bam NC_000022.11 -o 63003856_S135_chr22.bam
cd /home/alex/recherche/bisonex/code/BamScissors.jl
cp ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135_chr22.bam .
samtools index 63003856_chr22.bam
#+end_src
Le script va modifier le bam, le trier et générer le fastq. !!!
Attention: ne pas oublier l'option -n !!!
#+begin_src sh
time julia --project=.. insertVariant.jl
scp 63003856_S135_chr22_{1,2}.fq.gz meso:/Work/Users/apraga/bisonex/tests/bamscissors/
#+end_src
*** TODO Implémenter les SNV avec VAF :snv:
SCHEDULED: <2023-06-05 Mon>
Stratégie :
1. calculer la profondeur sur les positions
2. créer un dictionnaire { nom du reads : position dataframe }
3. itérer sur tous les reads et changer ceux marqués
**** DONE VAF = 1
CLOSED: [2023-05-29 Mon 15:34]
**** DONE VAF selon loi normale
CLOSED: [2023-05-29 Mon 15:35]
Tronquée si > 1
**** TODO Tests unitaires
SCHEDULED: <2023-05-29 Mon>
*** TODO Implémenter les indel avec VAF :indel:
SCHEDULED: <2023-05-29 Mon>
*** TODO Soumission paquet

Replacement in projects/bisonex.org at line 44 [19.35]

B:BD[18.24047] → [18.24047:24367]

B:BD[18.24367] → [20.558:8430]

B:BD[20.8430] → [3.144:8654]

∅:D[3.8654] → [21.8262:16454]

B:BD[21.8262] → [21.8262:16454]

B:BD[21.16454] → [4.27:8903]

           61        65         126
  21 │ NC_000015.10  74891539  g.74891539C>T  snv          heterozygous  C          T               118       124         242
  22 │ NC_000015.10  48488433  g.48488433A>G  snv          heterozygous  A          G               367       122         492
  23 │ NC_000015.10  89
318565  g.89318565A>G  snv          heterozygous  A          G               303        98         404
  24 │ NC_000015.10  89323426  g.89323426C>G  snv          heterozygous  C          G                93       109         202
  25 │ NC_000015.10  89318595  g.89318595T>C  snv          heterozygous  T          C               321       128         453
  26 │ NC_000015.10  48488437  g.48488437T>C  snv          heterozygous  T          C               356       132         488
CLOSED: [2023-05-01 Mon 17:18]
***** KILL Chromosome1 15 :Test haplotype caller : échec car CIGARE non mis à jour
CLOSED: [2023-05-13 Sat 18:29] SCHEDULED: <2023-05-01 Mon>
#+begin_src
julia -Jbisonex.so --project=. insertVariants.jl `63003856_S135_chr15.bam` 63003856_S135_chr15_inserted.bam
scp 63003856_S135_chr15_inserted.bam* meso:/Work/Users/apraga/bisonex/tests/synthetic/
#+end_src
#+begin_src sh :dir /ssh:meso:/Work/Users/apraga/bisonex/tests/synthetic :results silent
ln -s /Work/Projects/bisonex/data/dbSNP/GRCh38.p13/dbSNP.gz .
ln -s /Work/Projects/bisonex/data/dbSNP/GRCh38.p13/dbSNP.gz.tbi
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef.dict .
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef.fna .
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef.fna.fai .
#+end_src
puis
#+begin_src
gatk --java-options "-Xmx3072M" HaplotypeCaller --input 63003856_S135_chr15_inserted.bam --output testchr15.vcf.gz --reference genomeRef.fna  --tmp-dir . -L NC_000015.10
#+end_src
scp meso:/Work/Users/apraga/bisonex/tests/synthetic/testchr15.vcf.gz haplotypecaller-chr15.vcf.gz
Aucun variant inséré
- base quality ok
  -
****** DONE bam out : non appelé
CLOSED: [2023-05-01 Mon 21:57]
gatk --java-options "-Xmx3072M" HaplotypeCaller --input 63003856_S135_chr15_inserted.bam     --output haplotypecaller-chr15.vcf.gz --reference genomeRef.f
na  --tmp-dir . -L NC_000015.10  --bam-output debug.bam
****** DONE --linked-de-bruijn-graph : idem
CLOSED: [2023-05-01 Mon 21:57]
readlink testchr15.vcf.gz -f^C
[apraga@mesointeractive synthetic]$ gatk --java-options "-Xmx3072M" HaplotypeCaller --input 63003856_S135_chr15_inserted.bam     --output haplotypecaller-chr15.vcf.gz --reference genomeRef.fna  --tmp-dir . -L NC_000015.10  --linked-de-bruijn-graph
****** KILL regénérer fastq
CLOSED: [2023-05-13 Sat 18:29]
Non
***** KILL Générer bam données pour tous les chromosomes
CLOSED: [2023-05-13 Sat 18:29]
 timeit julia -Jbisonex.so --project=. insertVariants.jl ~/code/bisonex/out/63003856/preprocessing/63003856_S135.bam 63003856_S135_inserted.bam
40min 516ms 835µs 405ns
Avertissement:
 [W::bam_hdr_read] EOF marker is absent. The input is probably truncated
Inserted.bam et excluded.bam (fichier avant le merge)  ont l'air ok...
On réessaie à la main : ça passe
#+begin_src
samtools merge test-all.bam inserted.bam excluded.bam
❯ mv test-all.bam `63003856_S135_inserted.bam` -f
❯ mv test-all.bam.bai `63003856_S135_chr15_inserted.bam.bai` -f
#+end_src
***** DONE BAm2fastq pour avoir CIGAR à jour : échec (variants "cachés")
CLOSED: [2023-05-04 Thu 20:30] SCHEDULED: <2023-05-01 Mon>
On lance la génération de bam depuis le mesocentro (la copie plante via le VPN)
#+begin_src sh
cd /Work/Users/apraga/recherche/bisonex/generate
julia --project=. insertVariants.jl  ../../../bisonex/out/63003856_S135/preprocessing/applybqsr/63003856_S135.bam 63003856_S135_inserted.bam
#+end_src
Workflow après avec désactivé storeDir pour SAMTOOLS_BAM2FQ dans nextflow.config (pourquoi ??)
#+begin_src nextflow
include { SAMTOOLS_BAM2FQ }                            from "${params.modulesDir}/samtools/bam2fq/main"
include { SAMTOOLS_SORT as sortBamByName }             from "${params.modulesDir}/samtools/sort/main"
workflow {
    f = Channel.fromPath("${params.dataDir}/synthetic/63003856_S135_inserted.bam",
                         checkIfExists: true).map{it -> [["id": "synthetic_63003856"], it]}
    // Important: use "-n" option !!
    sortBamByName(f)
    SAMTOOLS_BAM2FQ(sortBamByName.out.bam, true)
}
#+end_src
Puis
#+begin_src
cp work/34/fb2fc136f6f6d7f42d0960512f06de/*.fq.gz /Work/Groups/bisonex/data/synthetic/
#+end_src
***** KILL Lancer pipeline
CLOSED: [2023-05-04 Thu 20:30] SCHEDULED: <2023-05-01 Mon>
NXF_OPTS=-D"user.name=apraga" nextflow run   main.nf -c nextflow.config  -profile standard,helios -bg --input="/Work/Groups/bisonex/data/synthetic/synthetic_63003856_{1,2}.fq.gz" --outdir out/synthetic_63003856
*** TODO Bamsurgeon :bamsurgeon:
**** TODO Package nix
1. Patcher la recherche du génome de référence pour bien trouver les index (en utilisant une regexp comme nf-core)
2. Rajouter le chemin de picard dans les arguments
3. Option -O3 pour performance
***** DONE Erreur ValueError: quality and sequence mismatch
CLOSED: [2023-05-19 Fri 18:44]
****** DONE Idem avec dernière version sur github
CLOSED: [2023-05-18 Thu 14:36]
****** DONE Version 1.3: ok mais
CLOSED: [2023-05-19 Fri 18:44]
Test sur chr22: variants ok mais VAF=1...
S'exécute "normalement" (échec selon nextflow) mais le bam de sorstie quasiment vide
La fusion du bam avec les variants et du fichier de référence n'a fonctionné correctement.
******* DONE Lancer replacereads.py à la main
CLOSED: [2023-05-18 Thu 21:41]
Dans /Work/Users/apraga/bisonex/work/f3/ce044f80ca91016d68d1bc4f4f5301
#+begin_src sh
 /nix/store/xw277la6w4sjqlsvw9h32cvrlacrfkgm-python3-3.10.9-env/bin/python3.10 /nix/store/abzangf0q8k37053p776cfkw181dzjn3-bamsurgeon-1.3/bin/bamsurgeon/replacereads.py -b cento.bam -r addsnv.e14561be-4fdd-45cc-9989-048ab6da6cc6.muts.bam -o snv-manual.bam
#+end_src
Puis
#+begin_src
samtools sort snv-manual.bam -o snv-manual-sorted.bam
#+end_src
A l'air de fonctionne
***** TODO Corriger run nextflow pour éviter les erreurs
SCHEDULED: <2023-05-21 Sun>
Trop de message d'erreur en sortie ?
***** DONE Test sur mini-bam: échec
CLOSED: [2023-05-14 Sun 21:12]
❯ samtools view -h ~/code/bisonex/simuscop-centogene-200x/cento/preprocessing/mapped/cento.bam | head -n1000 | samtools view -Sb - > mini.bam
❯ samtools index mini.bam
Sans spécfier le variant:
#+begin_quote
NC_000001.11	17651	17651
#+end_quote
./result/bin/addsnv -v snv.txt -f mini.bam -r ../data/genomeRef.fna -o test.bam
***** DONE Test chr22
CLOSED: [2023-05-15 Mon 23:24]
Pas assez de reads, on prend le chromosome 22
#+begin_src sh
samtools view ../simuscop-centogene-200x/cento/preprocessing/mapped/cento.bam NC_000022.11 -b -o chr22.bam
samtools index chr22.bam
#+end_src
Mésocentre
dans tests/bamsurgeno
#+begin_src
addsnv -v snv.txt -f chr22.bam -r ../genomeRef.fna -o test.bam --aligner mem
#+end_src
****** DONE SNV aléatoire:
CLOSED: [2023-05-15 Mon 23:13]
NC_000022.11	17499704	17499704    0.2
On retrouve bien un variant à cette position A > T
****** DONE SNV avec ALT prédéfini : retrouvée dans IGV (mais pas dans pileup)
CLOSED: [2023-05-15 Mon 23:13]
NC_000022.11	17499704	17499704    0.2 G
****** DONE Variants patients chr22: ok IGV
CLOSED: [2023-05-15 Mon 23:23]
Fichier non trié donc
samtools sort test.bam -o test-sorted.bam
samtools index test-sorted.bam
****** DONE Vérifier qu'il faut POS et POS+1: non
CLOSED: [2023-05-14 Sun 21:21]
**** TODO Variants cento
***** STRT SNV
SCHEDULED: <2023-05-15 Mon>
Attention à la mémoire: 32G ne semble pas suffire avec 12 threads
#+begin_src sh
NXF_OPTS=-D"user.name=${USER}" nextflow run workflows/bamsurgeon.nf -profile standard,helios --input=tests/bamsurgeon/snv-cento.tsv -bg
#+end_src
ET
#+begin_src nextflow
workflow {
    f = Channel.fromPath(params.input, checkIfExists: true)
    bam = Channel.fromPath("simuscop-centogene-200x/cento/preprocessing/mapped/cento.bam",
                         
  checkIfExists: true)
    bamIndex = bam.map { it -> it + ".bai" }
    downloadGenome | indexGenome
    indexGenome.out.index | view
    addSNV(f, bam, bamIndex, downloadGenome.out, indexGenome.out.index, indexGenome.out.dict, indexGenome.out.fai)
}
#+end_src
****** DONE v1.3: Lancer le pipeline pour vérifier qu'on retrouve les variants
CLOSED: [2023-05-19 Fri 18:41] SCHEDULED: <2023-05-18 Thu>
****** TODO Corrigier position pour avoir une bonne VAF
SCHEDULED: <2023-05-19 Fri>
POS POS+1 VAF ALT
Attention, la base corrigée est à POS+1...
****** DONE Comparaison manuelle avec julia (VAF = 1...)
CLOSED: [2023-05-19 Fri 21:58]
552 found over 585
***** TODO del
***** TODO ins
*** TODO Julia : Bamscissors :bamscissors:
Modification de la séquence dans BAM.
*Pas de mise à jour de CIGAR*
On convertit en fastq et on lance le pipeline pour "corriger"
#+begin_src sh
cd /home/alex/code/bisonex/out/63003856/preprocessing/mapped
samtools view 63003856_S135.bam NC_000022.11 -o 63003856_S135_chr22.bam
cd /home/alex/recherche/bisonex/code/BamScissors.jl
cp ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135_chr22.bam .
samtools index 63003856_chr22.bam
#+end_src
Le script va modifier le bam, le trier et générer le fastq. !!!
Attention: ne pas oublier l'option -n !!!
#+begin_src sh
time julia --project=.. insertVariant.jl
scp 63003856_S135_chr22_{1,2}.fq.gz meso:/Work/Users/apraga/bisonex/tests/bamscissors/
#+end_src
**** TODO Implémenter les SNV avec VAF
SCHEDULED: <2023-05-27 Sat>
Stratégie :
1. calculer la profondeur sur les positions
2. créer un dictionnaire { nom du reads : position dataframe }
3. itérer sur tous les reads et changer ceux marqués
**** TODO Implémenter les indel avec VAF
SCHEDULED: <2023-05-29 Mon>
**** TODO Phase 1 : chr22, VAF=1
***** DONE 1 SNV  : ok !
CLOSED: [2023-05-20 Sat 19:35] SCHEDULED: <2023-05-20 Sat>
#+begin_src
 make run READS="tests/bamscissors/corrected_{1,2}.fq.gz"
Puis on lance le pipeline sur correct_1
- [X] Variant visible dans IGV
- [X] Variant visible après alignement
- [X] Variant visible après appel de variant
***** TODO Tester SNV chromosome 22
SCHEDULED: <2023-05-20 Sat>
**** DONE PHase 2 : chr22, VAF variable: de nombreux reads sont perdus -> ok sur un SNV en alignant sur chromosome 22
CLOSED: [2023-05-24 Wed 23:19]
Problème dansr le BAM car sans les insertion
***** DONE Filtrer les reads sans pair : idem
CLOSED: [2023-05-23 Tue 00:01]
FOUND:Found 16662 unpaired mates
	at htsjdk.samtools.SAMUtils.processValidationError(SAMUtils.java:470)
	at picard.sam.SamToFastq.doWork(SamToFastq.java:224)
	at picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:308)
	at picard.cmdline.PicardCommandLine.instanceMain(PicardCommandLine.java:103)
	at picard.cmdline.PicardCommandLine.main(PicardCommandLine.java:113)
ex: chr22:42213078
On essaie
samtools view ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135.bam NC_000022.11  -hf 0x2 -o 63003856_chr22.bam
Ne rale pas...
SCHEDULED: <2023-05-22 Mon>
***** DONE Problème de la conversion BAM -> fastq ?
CLOSED: [2023-05-24 Wed 23:19]
****** DONE Test bamtofastq sur le BAM originel: idem
CLOSED: [2023-05-23 Tue 01:16]
Dans ~/recherche/code/bisonex/Bamscissors.jl
samtools view ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135.bam NC_000022.11  -hf 0x2 -o 63003856_chr22.bam
On trie bien par nom de read
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
Conversion en fastq. Attention à bien prendre le *fichier trié* !
samtools bam2fq  -1 63003856_chr22_init1.fq.gz -2 63003856_chr22_init2.fq.gz  -n 63003856_chr22_sorted.bam
[M::bam2fq_mainloop] discarded 0 singletons
[M::bam2fq_mainloop] processed 2592110 reads
On envoie sur le mésocentre
scp 63003856_chr22_init{1,2}.fq.gz meso:/Work/Users/apraga/bisonex/tests/bamscissors
On démarre un job avec 24 coeurs pour aller vite
srun -c 24 -p smp -t 1:00:00 --pty bash
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_init1.fq.gz 63003856_chr22_init2.fq.gz | samtools sort -@24 -o output.bam -
Dans /home/alex/recherche/bisonex/code/BamScissors.jl/aligned
❯ scp meso:/Work/Users/apraga/bisonex/tests/bamscissors/output.bam*
****** DONE Avec samtools fastq
CLOSED: [2023-05-23 Tue 01:16]
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
samtools fastq 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -o 63003856_chr22_sorted.bam
scp 63003856_chr22_{1,2}.fq.gz  meso:/Work/Users/apraga/bisonex/tests/bamscissors/
mesocentre
 bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz | samtools sort -@24 -o output.bam
 scp meso:/Work/Users/apraga/bisonex/tests/bamscissors/output.bam\* aligned/
****** DONE En rajoutant .fna dans le doserrie (+samtools fastq): idem
CLOSED: [2023-05-23 Tue 01:16]
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef* .
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef* .
bwa mem -t 24 genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz | samtools sort -@24 -o output.bam
****** DONE Test picard : idem
CLOSED: [2023-05-23 Tue 01:16]
Dans bisonex/code/BamScissors.jl
❯ picard SamToFastq -I 63003856_chr22_sorted.bam -F testpicard1.fastq -F2 testpicard2.fastq
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef testpicard1.fastq.gz testpicard2.fastq.gz | samtools sort -@24 -o testpicard.bam -
****** DONE Il ne manque pas des reads dans les fastq :
CLOSED: [2023-05-23 Tue 01:26]
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ samtools  flagstat aligned/output.bam
1306273 + 0 in total (QC-passed reads + QC-failed reads)
1296055 + 0 primary
0 + 0 secondary
10218 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
1304871 + 0 mapped (99.89% : N/A)
1294653 + 0 primary mapped (99.89% : N/A)
0 + 0 paired in sequencing
0 + 0 read1
0 + 0 read2
0 + 0 properly paired (N/A : N/A)
0 + 0 with itself and mate mapped
0 + 0 singletons (N/A : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ samtools flagstat 63003856_chr22.bam
2609214 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
17104 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2609214 + 0 mapped (100.00% : N/A)
2592110 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2592110 + 0 properly paired (100.00% : N/A)
2592110 + 0 with itself and mate mapped
0 + 0 singletons (0.00% : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
❯ echo $(zcat 63003856_chr22_init2.fq.gz | wc -l)/4 | bc
1296055
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0 took 2s
❯ echo $(zcat 63003856_chr22_init1.fq.gz | wc -l)/4 | bc
1296055
❯ samtools  flagstat 63003856_chr22_sorted.bam
2609214 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
17104 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2609214 + 0 mapped (100.00% : N/A)
2592110 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2592110 + 0 properly paired (100.00% : N/A)
2592110 + 0 with itself and mate mapped
0 + 0 singletons (0.00% : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
****** DONE Avec sort -O BAM idem
CLOSED: [2023-05-23 Tue 01:21]
****** DONE Singleton ou non mapped ? nonVirtPosition
CLOSED: [2023-05-23 Tue 01:22]
❯ samtools bam2fq  -1 63003856_chr22_init1.fq.gz -2 63003856_chr22_init2.fq.gz -0 both -s single.fq.gz  -n 63003856_chr22_sorted.bam
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ wc -l 63003856_chr22_init* both single.fq.gz
   403540 63003856_chr22_init1.fq.gz
   404788 63003856_chr22_init2.fq.gz
        0 both
        0 single.fq.gz
   808328 total
****** Problème d'aligner car les reads sont bien dans le .fastq ?
Ex:
 zgrep "A00853:477:HMLWYDSX3:2:2624:2826:18630" 63003856_chr22_{1,2}.fq.gz
63003856_chr22_1.fq.gz:@A00853:477:HMLWYDSX3:2:2624:2826:1863
0
63003856_chr22_2.fq.gz:@A00853:477:HMLWYDSX3:2:2624:2826:18630
***** DONE Refaire la manip sur bam chr22 non modifié + mail Alexis
CLOSED: [2023-05-23 Tue 23:27]
[1]_samtools view ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135.bam NC_000022.11  -f 0x2 -o 63003856_chr22.bam
 Les reads sont bien tous mappé
samtools flagstat 63003856_chr22.bam
2609214 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
17104 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2609214 + 0 mapped (100.00% : N/A)
[2] samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
flagstat idem
[3] samtools fastq -1 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -n 63003856_chr22_sorted.bam
[M::bam2fq_mainloop] discarded 0 singletons
[M::bam2fq_mainloop] processed 2592110 reads
Nombre de reads ok (= la moitié) et les séquences ne sont pas identiques pour le premier read (= on n'a pas 2x la même chose)
 echo $(zcat 63003856_chr22_1.fq.gz | wc -l)/4 | bc
1296055
 echo $(zcat 63003856_chr22_2.fq.gz | wc -l)/4 | bc
1296055
 [4]
 bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz  -o wtf.bam
 [M::bwa_idx_load_from_disk] read 0 ALT contigs
[M::process] read 1752492 sequences (240000014 bp)...
[M::mem_pestat] # candidate unique pairs for (FF, FR, RF, RR): (12, 702821, 0, 18)
[M::mem_pestat] analyzing insert size distribution for orientation FF...
[M::mem_pestat] (25, 50, 75) percentile: (54, 197, 268)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 696)
[M::mem_pestat] mean and std.dev: (163.92, 129.71)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 910)
[M::mem_pestat] analyzing insert size distribution for orientation FR...
[M::mem_pestat] (25, 50, 75) percentile: (129, 175, 233)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 441)
[M::mem_pestat] mean and std.dev: (185.56, 75.37)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 545)
[M::mem_pestat] skip orientation RF as there are not enough pairs
[M::mem_pestat] analyzing insert size distribution for orientation RR...
[M::mem_pestat] (25, 50, 75) percentile: (56, 192, 239)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 605)
[M::mem_pestat] mean and std.dev: (158.00, 110.30)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 788)
[M::mem_pestat] skip orientation FF
[M::mem_pestat] skip orientation RR
[M::process] read 839618 sequences (114952336 bp)...
[M::mem_process_seqs] Processed 1752492 reads in 375.714 CPU sec, 17.645 real sec
[M::mem_pestat] # candidate unique pairs for (FF, FR, RF, RR): (0, 336379, 0, 5)
[M::mem_pestat] skip orientation FF as there are not enough pairs
[M::mem_pestat] analyzing insert size distribution for orientation FR...
[M::mem_pestat] (25, 50, 75) percentile: (128, 174, 232)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 440)
[M::mem_pestat] mean and std.dev: (184.73, 74.63)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 544)
[M::mem_pestat] skip orientation RF as there are not enough pairs
[M::mem_pestat] skip orientation RR as there are not enough pairs
[M::mem_process_seqs] Processed 839618 reads in 183.039 CPU sec, 7.961 real sec
[main] Version: 0.7.17-r1188
[main] CMD: bwa mem -t 24 -o wtf.bam /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz
[main] Real time: 38.278 sec; CPU: 565.821 sec
Bon nombre de reads pourtant
 samtools flagstat wtf.bam
2611059 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
18949 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2611058 + 0 mapped (100.00% : N/A)
2592109 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2590970 + 0 properly paired (99.96% : N/A)
2592108 + 0 with itself and mate mapped
1 + 0 singletons (0.00% : N/A)
458 + 0 with mate mapped to a different chr
63 + 0 with mate mapped to a different chr (mapQ>=5)
$ samtools sort -@24 -o wtf_sorted.bam wtf.sam
[bam_sort_core] merging from 0 files and 24 in-memory blocks...
 samtools flagstat wtf.bam
2611059 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
18949 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2611058 + 0 mapped (100.00% : N/A)
2592109 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2590970 + 0 properly paired (99.96% : N/A)
2592108 + 0 with itself and mate mapped
1 + 0 singletons (0.00% : N/A)
458 + 0 with mate mapped to a different chr
63 + 0 with mate mapped to a different chr (mapQ>=5)
Effectivement, ce n'est pas un problème d'IGV
❯ samtools mpileup 63003856_chr22.bam -r NC_000022.11:42213078-42213078
[mpileup] 1 samples in 1 input files
NC_000022.11	42213078	N	19	TTtTTtTTTtTTtTtTttt	kkk_kkkkFkFF_FkFkQk
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ samtools mpileup aligned/wtf_sorted.bam -r NC_000022.11:42213078-42213078
[mpileup] 1 samples in 1 input files
NC_000022.11	42213078	N	5	TTtTT	_FkFF
***** DONE Regarder où ont été aligné les reads (nouveau run)
CLOSED: [2023-05-24 Wed 23:18]
****** DONE Préparation
CLOSED: [2023-05-24 Wed 21:59]
On relance le pipeline pour avoir un BAM propre
On garde les reads non mappé à partsir de la sortie d'applybqsr
#+begin_src sh
NXF_OPTS=-D"user.name=apraga" nextflow run main.nf -c nextflow.config  -profile standard,helios --input="/Work/Projects/bisonex/centogene/fastq/2200467051_63003856/63003856_S135_R{1,2}_001.fastq.gz" --outdir=out -bg
cd out/63003856_S135_R/preprocessing/applybqsr/
samtools view 63003856_S135_R.bam NC_000022.11  -f 0x2 -o 63003856_chr22.bam
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
samtools fastq -1 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -n 63003856_chr22_sorted.bam
make run BG= READS="out/63003856_S135_R/preprocessing/applybqsr/63003856_chr22_{1,2}.fq.gz"
cd out/63003856_chr22/preprocessing/mapped/
samtools index 63003856_chr22.bam
samtools mpileup 63003856_chr22.bam -r NC_000022.11:42213078-42213078
#+end_src
On récupère les 2 bam dans
#+begin_src
cd /home/alex/recherche/bisonex/code/BamScissors.jl/
rsync -avz meso:/Work/Users/apraga/bisonex/out/63003856_chr22/preprocessing/mapped data
rsync -avz meso:/Work/Users/apraga/bisonex/out/63003856_S135_R/preprocessing/applybqsr/ data/init/
#+end_src
****** Vérification que le reads est ailleurs
On cherche un read manquant dans le second alignement
#+begin_src sh
samtools view data/init/63003856_chr22.bam | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
#+end_src
#+RESULTS:
: A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NC_000022.11	42212845	0	151M	=	42212883	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	ACC+FBCDCBBBAEAEDEEBBCCCECACBAEBEBDCCBCBFDCCCCFACEBEBCEEDCCCCFDCAEDCACBCEBBCFEACCFBDCACDCBCEBDBBCFEEDCCCFAFEACECCCECAEEDCADCBEDC7BEBCCCFBAFDCECCFBEAACA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
: A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NC_000022.11	42212883	0	151M	=	42212845	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	AADECCCBDCBFCE<?CDEEEEBDEACDEAC;:BFBCBCDCCBEAEACAEFCCEAFBCBCCDEECBDBCECBEECCEACDEEBBFGDEFGCCFFFFCFCCEFBFDCFCDAAEBEE:CECBABBEBEE;DBFCCCDBCDBCCBBC?@BEEDA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
#+begin_src sh
samtools view data/mapped/63003856_chr22.bam | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
#+end_src
#+RESULTS:
: A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NW_014040930.1	115017	0	151M	=	115055	189	CCCAGGGGCCCCAGTGGGGATTTTCT
AATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	ACC+FBCDCBBBAEAEDEEBBCCCECACBAEBEBDCCBCBFDCCCCFACEBEBCEEDCCCCFDCAEDCACBCEBBCFEACCFBDCACDCBCEBDBBCFEEDCCCFAFEACECCCECAEEDCADCBEDC7BEBCCCFBAFDCECCFBEAACA	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
: A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NW_014040930.1	115055	0	151M	=	115017	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	AADECCCBDCBFCE<?CDEEEEBDEACDEAC;:BFBCBCDCCBEAEACAEFCCEAFBCBCCDEECBDBCECBEECCEACDEEBBFGDEFGCCFFFFCFCCEFBFDCFCDAAEBEE:CECBABBEBEE;DBFCCCDBCDBCCBBC?@BEEDA	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
Effectivement, on aligne sur une zonne supprimée !
***** DONE Corriger la qualité: non
CLOSED: [2023-05-24 Wed 22:19]
****** DONE Comparaison avec le fastq de référénce : qualité !!
CLOSED: [2023-05-24 Wed 22:17]
#+begin_src sh
cd /Work/Users/apraga/bisonex/work/6e/8548fc90263830bf677f36585f11dc
zgrep -A 3 "A00853:477:HMLWYDSX3:1:1413:4390:28573" 63003856_chr22_1.fq.gz
#+end_src
@A00853:477:HMLWYDSX3:1:1413:4390:28573
AGGGTTACCACCACCACCCTGACAGGAGATATTCTAGGAGTACTCAAGAGCATCAGGGGATGGCTGGTAGCCTAGAAGGAACCACAAGGCCCAATGTCTTGGTTAGTCAAACCAATGAATTAGCTAGCAGGGGCCTTCTGAACAAAAGCAT
+
ADEEB@?CBBCCBDCBDCCCFBD;EEBEBBABCEC:EEBEAADCFCDFBFECCFCFFFFCCGFEDGFBBEEDCAECCEEBCECBDBCEEDCCBCBFAECCFEACAEAEBCCDCBCBFB:;CAEDCAEDBEEEEDC?<ECFBCDBCCCEDAA
#+begin_src
zgrep -A 3 "A00853:477:HMLWYDSX3:1:1413:4390:28573" /Work/Projects/bisonex/centogene/fastq/2200467051_63003856/63003856_S135_R1_001.fastq.gz
#+end_src
#+RESULTS:
: @A00853:477:HMLWYDSX3:1:1413:4390:28573 1:N:0:ATTCCACACA+TAGGCGATTG
AGGGTTACCACCACCACCCTGACAGGAGATATTCTAGGAGTACTCAAGAGCATCAGGGGATGGCTGGTAGCCTAGAAGGAACCACAAGGCCCAATGTCTTGGTTAGTCAAACCAATGAATTAGCTAGCAGGGGCCTTCTGAACAAAAGCAT
: +
: FFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFF
****** DONE Regarder la qualité après bwa mem vs applybqsr: différente
CLOSED: [2023-05-24 Wed 22:18]
Sur le mésocentre, dans /Work/Users/apraga/bisonex/out/63003856_S135_R/preprocessing
$ samtools view mapped/63003856_S135_R.bam NC_000022.11 | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NC_000022.11	42212845	0	151M	=	42212883	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	FFF,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NC_000022.11	42212883	0	151M	=	42212845	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	FFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
samtools view applybqsr/63003856_S135_R.bam NC_000022.11 | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NC_000022.11	42212845	0	151M	=	42212883	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	ACC+FBCDCBBBAEAEDEEBBCCCECACBAEBEBDCCBCBFDCCCCFACEBEBCEEDCCCCFDCAEDCACBCEBBCFEACCFBDCACDCBCEBDBBCFEEDCCCFAFEACECCCECAEEDCADCBEDC7BEBCCCFBAFDCECCFBEAACA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NC_000022.11	42212883	0	151M	=	42212845	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	AADECCCBDCBFCE<?CDEEEEBDEACDEAC;:BFBCBCDCCBEAEACAEFCCEAFBCBCCDEECBDBCECBEECCEACDEEBBFGDEFGCCFFFFCFCCEFBFDCFCDAAEBEE:CECBABBEBEE;DBFCCCDBCDBCCBBC?@BEEDA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
****** DONE Réaligner à partir de la sortie de bwa mem
CLOSED: [2023-05-24 Wed 22:32]
#+begin_src sh
cd out/63003856_S135_R/preprocessing/mapped/
samtools view 63003856_S135_R.bam NC_000022.11  -f 0x2 -o 63003856_chr22.bam
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
samtools fastq -1 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -n 63003856_chr22_sorted.bam
#+end_src
ON vérifie la qualité
#+begin_src
zgrep -A 3 "A00853:477:HMLWYDSX3:1:1413:4390:28573" 63003856_chr22_1.fq.gz
#+end_src
#+RESULTS:
: @A00853:477:HMLWYDSX3:1:1413:4390:28573
: AGGGTTACCACCACCACCCTGACAGGAGATATTCTAGGAGTACTCAAGAGCATCAGGGGATGGCTGGTAGCCTAGAAGGAACCACAAGGCCCAATGTCTTGGTTAGTCAAACCAATGAATTAGCTAGCAGGGGCCTTCTGAACAAAAGCAT
: +
: FFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFF
#+begin_src
NXF_OPTS=-D"user.name=apraga" nextflow run main.nf -c nextflow.config  -profile standard,helios -resume  --input="out/63003856_S135_R/preprocessing/mapped/63003856_chr22_{1,2}.fq.gz" --outdir=out/63003856_chr22-from-mapped
#+end_src
Puis ::
#+begin_src
cd /Work/Users/apraga/bisonex/out/63003856_chr22-from-mapped/63003856_chr22/preprocessing/mapped
samtools view 63003856_chr22.bam | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
#+end_src
#+RESULTS:
: A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NW_014040930.1	115017	0	151M	=	115055	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	FFF,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
: A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NW_014040930.1	115055	0	151M	=	115017	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	FFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
***** DONE Aligner sur génome de référence limité au chromosome 22
CLOSED: [2023-05-24 Wed 23:18]
****** KILL Test données non modifiées
CLOSED: [2023-05-24 Wed 23:18]
/Work/Users/apraga/bisonex/tests/bamscissors
#+begin_src
cd /Work/Groups/bisonex/data/genome/GRCh38.p13/
mkdir chr22/
samtools faidx genomeRef.fna NC_000022.11 > chr22/chr22.fna
cd chr22
samtools faidx chr22.fna
bwa index chr22.fna
#+end_src
#+begin_src
cd /Work/Users/apraga/bisonex/tests/bamscissors
ln -s ../../out/63003856_S135_R/preprocessing/applybqsr/63003856_chr22_{1,2}.fq.gz .
srun -c 24 -p smp -t 1:00:00 --pty bash
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/chr22/chr22.fna 63003856_chr22_1.fq.gz 63003856_chr22_1.fq.gz -o smallref.sam
#+end_src
****** DONE Test données modifiées: ok
CLOSED: [2023-05-24 Wed 23:18]
Données dans data/init
#+begin_src sh
time julia insertVariant.jl
rsync -avz data/init/*.fq.gz meso:/Work/Users/apraga/bisonex/tests/bamscissors/
#+end_src
#+begin_src
srun -c 24 -p smp -t 1:00:00 --pty bash
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/chr22/chr22.fna 63003856_chr22_1.fq.gz 63003856_chr22_1.fq.gz | samtools sort -@24 - -o smallref.bam
#+end_src
#+begin_src
rsync -avz meso:/Work/Users/apraga/bisonex/tests/bamscissors/smallref.bam mapped/
#+end_src
**** TODO Phase 3 : insertion SNV chromosome 22 sur BAM complet + test haplotypecaller
NXF_OPTS=-D"user.name=${USER}" nextflow run workflows/runInserted.nf -profile standard,helios  --input="tests/bamscissors/63003856_chr22_{1,2}.fq.gz" -resume
On vérifie le nombre de reads :
cd work/13/3faf1c868bfe61d22da346319b27aa/
samtools mpileup 63003856_chr22.bam -r NC_000022.11:42213078-42213078
[mpileup] 1 samples in 1 input files
NC_000022.11	42213078	N	18	GgGGgGGGgGGgGgGggg	bbZbbbbCbCC\Aa@bTb
rsync -avz meso:/Work/Users/apraga/bisonex/work/8d/b76d9079cc59a5daaefa74653d2a9d/63003856_chr22.bam\* mapped
ok
Après haplotypecaller
 rsync -avz meso:/Work/Users/apraga/bisonex/work/68/e50b99ecc879039086c4c1fc2f24d3/63003856_chr22.vcf.gz\* variantCalling
 On retrouve bien le variant !
*** Divers
**** DONE Vérifier nombre de reads fastq - bam
CLOSED: [2022-10-09 Sun 22:31]

[18.24047]

           61        65         126
  21 │ NC_000015.10  74891539  g.74891539C>T  snv          heterozygous  C          T               118       124         242
  22 │ NC_000015.10  48488433  g.48488433A>G  snv          heterozygous  A          G               367       122         492
  23 │ NC_000015.10  89318565  g.89318565A>G  snv          heterozygous  A          G               303        98         404
  24 │ NC_000015.10  89323426  g.89323426C>G  snv          heterozygous  C          G                93       109         202
  25 │ NC_000015.10  89318595  g.89318595T>C  snv          heterozygous  T          C               321       128         453
  26 │ NC_000015.10  48488437  g.48488437T>C  snv          heterozygous  T          C               356       132         488
CLOSED: [2023-05-01 Mon 17:18]
***** KILL Chromosome1 15 :Test haplotype caller : échec car CIGARE non mis à jour
CLOSED: [2023-05-13 Sat 18:29] SCHEDULED: <2023-05-01 Mon>
#+begin_src
julia -Jbisonex.so --project=. insertVariants.jl `63003856_S135_chr15.bam` 63003856_S135_chr15_inserted.bam
scp 63003856_S135_chr15_inserted.bam* meso:/Work/Users/apraga/bisonex/tests/synthetic/
#+end_src
#+begin_src sh :dir /ssh:meso:/Work/Users/apraga/bisonex/tests/synthetic :results silent
ln -s /Work/Projects/bisonex/data/dbSNP/GRCh38.p13/dbSNP.gz .
ln -s /Work/Projects/bisonex/data/dbSNP/GRCh38.p13/dbSNP.gz.tbi
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef.dict .
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef.fna .
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef.fna.fai .
#+end_src
puis
#+begin_src
gatk --java-options "-Xmx3072M" HaplotypeCaller --input 63003856_S135_chr15_inserted.bam --output testchr15.vcf.gz --reference genomeRef.fna  --tmp-dir . -L NC_000015.10
#+end_src
scp meso:/Work/Users/apraga/bisonex/tests/synthetic/testchr15.vcf.gz haplotypecaller-chr15.vcf.gz
Aucun variant inséré
- base quality ok
  -
****** DONE bam out : non appelé
CLOSED: [2023-05-01 Mon 21:57]
gatk --java-options "-Xmx3072M" HaplotypeCaller --input 63003856_S135_chr15_inserted.bam     --output haplotypecaller-chr15.vcf.gz --reference genomeRef.f
na  --tmp-dir . -L NC_000015.10  --bam-output debug.bam
****** DONE --linked-de-bruijn-graph : idem
CLOSED: [2023-05-01 Mon 21:57]
readlink testchr15.vcf.gz -f^C
[apraga@mesointeractive synthetic]$ gatk --java-options "-Xmx3072M" HaplotypeCaller --input 63003856_S135_chr15_inserted.bam     --output haplotypecaller-chr15.vcf.gz --reference genomeRef.fna  --tmp-dir . -L NC_000015.10  --linked-de-bruijn-graph
****** KILL regénérer fastq
CLOSED: [2023-05-13 Sat 18:29]
Non
***** KILL Générer bam données pour tous les chromosomes
CLOSED: [2023-05-13 Sat 18:29]
 timeit julia -Jbisonex.so --project=. insertVariants.jl ~/code/bisonex/out/63003856/preprocessing/63003856_S135.bam 63003856_S135_inserted.bam
40min 516ms 835µs 405ns
Avertissement:
 [W::bam_hdr_read] EOF marker is absent. The input is probably truncated
Inserted.bam et excluded.bam (fichier avant le merge)  ont l'air ok...
On réessaie à la main : ça passe
#+begin_src
samtools merge test-all.bam inserted.bam excluded.bam
❯ mv test-all.bam `63003856_S135_inserted.bam` -f
❯ mv test-all.bam.bai `63003856_S135_chr15_inserted.bam.bai` -f
#+end_src
***** DONE BAm2fastq pour avoir CIGAR à jour : échec (variants "cachés")
CLOSED: [2023-05-04 Thu 20:30] SCHEDULED: <2023-05-01 Mon>
On lance la génération de bam depuis le mesocentro (la copie plante via le VPN)
#+begin_src sh
cd /Work/Users/apraga/recherche/bisonex/generate
julia --project=. insertVariants.jl  ../../../bisonex/out/63003856_S135/preprocessing/applybqsr/63003856_S135.bam 63003856_S135_inserted.bam
#+end_src
Workflow après avec désactivé storeDir pour SAMTOOLS_BAM2FQ dans nextflow.config (pourquoi ??)
#+begin_src nextflow
include { SAMTOOLS_BAM2FQ }                            from "${params.modulesDir}/samtools/bam2fq/main"
include { SAMTOOLS_SORT as sortBamByName }             from "${params.modulesDir}/samtools/sort/main"
workflow {
    f = Channel.fromPath("${params.dataDir}/synthetic/63003856_S135_inserted.bam",
                         checkIfExists: true).map{it -> [["id": "synthetic_63003856"], it]}
    // Important: use "-n" option !!
    sortBamByName(f)
    SAMTOOLS_BAM2FQ(sortBamByName.out.bam, true)
}
#+end_src
Puis
#+begin_src
cp work/34/fb2fc136f6f6d7f42d0960512f06de/*.fq.gz /Work/Groups/bisonex/data/synthetic/
#+end_src
***** KILL Lancer pipeline
CLOSED: [2023-05-04 Thu 20:30] SCHEDULED: <2023-05-01 Mon>
NXF_OPTS=-D"user.name=apraga" nextflow run   main.nf -c nextflow.config  -profile standard,helios -bg --input="/Work/Groups/bisonex/data/synthetic/synthetic_63003856_{1,2}.fq.gz" --outdir out/synthetic_63003856
*** TODO Bamsurgeon :bamsurgeon:
**** TODO Package nix
1. Patcher la recherche du génome de référence pour bien trouver les index (en utilisant une regexp comme nf-core)
2. Rajouter le chemin de picard dans les arguments
3. Option -O3 pour performance
***** DONE Erreur ValueError: quality and sequence mismatch
CLOSED: [2023-05-19 Fri 18:44]
****** DONE Idem avec dernière version sur github
CLOSED: [2023-05-18 Thu 14:36]
****** DONE Version 1.3: ok mais
CLOSED: [2023-05-19 Fri 18:44]
Test sur chr22: variants ok mais VAF=1...
S'exécute "normalement" (échec selon nextflow) mais le bam de sorstie quasiment vide
La fusion du bam avec les variants et du fichier de référence n'a fonctionné correctement.
******* DONE Lancer replacereads.py à la main
CLOSED: [2023-05-18 Thu 21:41]
Dans /Work/Users/apraga/bisonex/work/f3/ce044f80ca91016d68d1bc4f4f5301
#+begin_src sh
 /nix/store/xw277la6w4sjqlsvw9h32cvrlacrfkgm-python3-3.10.9-env/bin/python3.10 /nix/store/abzangf0q8k37053p776cfkw181dzjn3-bamsurgeon-1.3/bin/bamsurgeon/replacereads.py -b cento.bam -r addsnv.e14561be-4fdd-45cc-9989-048ab6da6cc6.muts.bam -o snv-manual.bam
#+end_src
Puis
#+begin_src
samtools sort snv-manual.bam -o snv-manual-sorted.bam
#+end_src
A l'air de fonctionne
***** TODO Corriger run nextflow pour éviter les erreurs
Trop de message d'erreur en sortie ?
***** DONE Test sur mini-bam: échec
CLOSED: [2023-05-14 Sun 21:12]
❯ samtools view -h ~/code/bisonex/simuscop-centogene-200x/cento/preprocessing/mapped/cento.bam | head -n1000 | samtools view -Sb - > mini.bam
❯ samtools index mini.bam
Sans spécfier le variant:
#+begin_quote
NC_000001.11	17651	17651
#+end_quote
./result/bin/addsnv -v snv.txt -f mini.bam -r ../data/genomeRef.fna -o test.bam
***** DONE Test chr22
CLOSED: [2023-05-15 Mon 23:24]
Pas assez de reads, on prend le chromosome 22
#+begin_src sh
samtools view ../simuscop-centogene-200x/cento/preprocessing/mapped/cento.bam NC_000022.11 -b -o chr22.bam
samtools index chr22.bam
#+end_src
Mésocentre
dans tests/bamsurgeno
#+begin_src
addsnv -v snv.txt -f chr22.bam -r ../genomeRef.fna -o test.bam --aligner mem
#+end_src
****** DONE SNV aléatoire:
CLOSED: [2023-05-15 Mon 23:13]
NC_000022.11	17499704	17499704    0.2
On retrouve bien un variant à cette position A > T
****** DONE SNV avec ALT prédéfini : retrouvée dans IGV (mais pas dans pileup)
CLOSED: [2023-05-15 Mon 23:13]
NC_000022.11	17499704	17499704    0.2 G
****** DONE Variants patients chr22: ok IGV
CLOSED: [2023-05-15 Mon 23:23]
Fichier non trié donc
samtools sort test.bam -o test-sorted.bam
samtools index test-sorted.bam
****** DONE Vérifier qu'il faut POS et POS+1: non
CLOSED: [2023-05-14 Sun 21:21]
**** HOLD Variants cento
***** STRT SNV
Attention à la mémoire: 32G ne semble pas suffire avec 12 threads
#+begin_src sh
NXF_OPTS=-D"user.name=${USER}" nextflow run workflows/bamsurgeon.nf -profile standard,helios --input=tests/bamsurgeon/snv-cento.tsv -bg
#+end_src
ET
#+begin_src nextflow
workflow {
    f = Channel.fromPath(params.input, checkIfExists: true)
    bam = Channel.fromPath("simuscop-centogene-200x/cento/preprocessing/mapped/cento.bam",
                           checkIfExists: true)
    bamIndex = bam.map { it -> it + ".bai" }
    downloadGenome | indexGenome
    indexGenome.out.index | view
    addSNV(f, bam, bamIndex, downloadGenome.out, indexGenome.out.index, indexGenome.out.dict, indexGenome.out.fai)
}
#+end_src
****** DONE v1.3: Lancer le pipeline pour vérifier qu'on retrouve les variants
CLOSED: [2023-05-19 Fri 18:41] SCHEDULED: <2023-05-18 Thu>
****** HOLD Corrigier position pour avoir une bonne VAF
POS POS+1 VAF ALT
Attention, la base corrigée est à POS+1...
****** DONE Comparaison manuelle avec julia (VAF = 1...)
CLOSED: [2023-05-19 Fri 21:58]
552 found over 585
***** TODO del
***** TODO ins
*** TODO [[id:966a298c-948a-4694-a6f5-c326b1046a05][XAMscissors.jl]] :xamscissors:
**** TODO Test SNV
SCHEDULED: <2023-05-30 Tue>
***** DONE Phase 1 : chr22, VAF=1
CLOSED: [2023-05-29 Mon 15:36]
****** DONE 1 SNV  : ok !
CLOSED: [2023-05-20 Sat 19:35] SCHEDULED: <2023-05-20 Sat>
#+begin_src
 make run READS="tests/bamscissors/corrected_{1,2}.fq.gz"
Puis on lance le pipeline sur correct_1
- [X] Variant visible dans IGV
- [X] Variant visible après alignement
- [X] Variant visible après appel de variant
****** KILL Tester SNV chromosome 22
CLOSED: [2023-05-29 Mon 15:36] SCHEDULED: <2023-05-20 Sat>
***** DONE PHase 2 : chr22, VAF variable
****** DONE de nombreux reads sont perdus -> ok sur un SNV en alignant sur chromosome 22
CLOSED: [2023-05-29 Mon 15:38]
Problème dansr le BAM car sans les insertion
******* DONE Filtrer les reads sans pair : idem
CLOSED: [2023-05-23 Tue 00:01]
FOUND:Found 16662 unpaired mates
	at htsjdk.samtools.SAMUtils.processValidationError(SAMUtils.java:470)
	at picard.sam.SamToFastq.doWork(SamToFastq.java:224)
	at picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:308)
	at picard.cmdline.PicardCommandLine.instanceMain(PicardCommandLine.java:103)
	at picard.cmdline.PicardCommandLine.main(PicardCommandLine.java:113)
ex: chr22:42213078
On essaie
samtools view ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135.bam NC_000022.11  -hf 0x2 -o 63003856_chr22.bam
Ne rale pas...
SCHEDULED: <2023-05-22 Mon>
******* DONE Problème de la conversion BAM -> fastq ?
CLOSED: [2023-05-24 Wed 23:19]
******** DONE Test bamtofastq sur le BAM originel: idem
CLOSED: [2023-05-23 Tue 01:16]
Dans ~/recherche/code/bisonex/Bamscissors.jl
samtools view ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135.bam NC_000022.11  -hf 0x2 -o 63003856_chr22.bam
On trie bien par nom de read
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
Conversion en fastq. Attention à bien prendre le *fichier trié* !
samtools bam2fq  -1 63003856_chr22_init1.fq.gz -2 63003856_chr22_init2.fq.gz  -n 63003856_chr22_sorted.bam
[M::bam2fq_mainloop] discarded 0 singletons
[M::bam2fq_mainloop] processed 2592110 reads
On envoie sur le mésocentre
scp 63003856_chr22_init{1,2}.fq.gz meso:/Work/Users/apraga/bisonex/tests/bamscissors
On démarre un job avec 24 coeurs pour aller vite
srun -c 24 -p smp -t 1:00:00 --pty bash
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_init1.fq.gz 63003856_chr22_init2.fq.gz | samtools sort -@24 -o output.bam -
Dans /home/alex/recherche/bisonex/code/BamScissors.jl/aligned
❯ scp meso:/Work/Users/apraga/bisonex/tests/bamscissors/output.bam*
******** DONE Avec samtools fastq
CLOSED: [2023-05-23 Tue 01:16]
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
samtools fastq 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -o 63003856_chr22_sorted.bam
scp 63003856_chr22_{1,2}.fq.gz  meso:/Work/Users/apraga/bisonex/tests/bamscissors/
mesocentre
 bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz | samtools sort -@24 -o output.bam
 scp meso:/Work/Users/apraga/bisonex/tests/bamscissors/output.bam\* aligned/
******** DONE En rajoutant .fna dans le doserrie (+samtools fastq): idem
CLOSED: [2023-05-23 Tue 01:16]
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef* .
ln -s /Work/Projects/bisonex/data/genome/GRCh38.p13/genomeRef* .
bwa mem -t 24 genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz | samtools sort -@24 -o output.bam
******** DONE Test picard : idem
CLOSED: [2023-05-23 Tue 01:16]
Dans bisonex/code/BamScissors.jl
❯ picard SamToFastq -I 63003856_chr22_sorted.bam -F testpicard1.fastq -F2 testpicard2.fastq
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef testpicard1.fastq.gz testpicard2.fastq.gz | samtools sort -@24 -o testpicard.bam -
******** DONE Il ne manque pas des reads dans les fastq :
CLOSED: [2023-05-23 Tue 01:26]
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ samtools  flagstat aligned/output.bam
1306273 + 0 in total (QC-passed reads + QC-failed reads)
1296055 + 0 primary
0 + 0 secondary
10218 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
1304871 + 0 mapped (99.89% : N/A)
1294653 + 0 primary mapped (99.89% : N/A)
0 + 0 paired in sequencing
0 + 0 read1
0 + 0 read2
0 + 0 properly paired (N/A : N/A)
0 + 0 with itself and mate mapped
0 + 0 singletons (N/A : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ samtools flagstat 63003856_chr22.bam
2609214 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
17104 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2609214 + 0 mapped (100.00% : N/A)
2592110 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2592110 + 0 properly paired (100.00% : N/A)
2592110 + 0 with itself and mate mapped
0 + 0 singletons (0.00% : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
❯ echo $(zcat 63003856_chr22_init2.fq.gz | wc -l)/4 | bc
1296055
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0 took 2s
❯ echo $(zcat 63003856_chr22_init1.fq.gz | wc -l)/4 | bc
1296055
❯ samtools  flagstat 63003856_chr22_sorted.bam
2609214 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
17104 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2609214 + 0 mapped (100.00% : N/A)
2592110 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2592110 + 0 properly paired (100.00% : N/A)
2592110 + 0 with itself and mate mapped
0 + 0 singletons (0.00% : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
******** DONE Avec sort -O BAM idem
CLOSED: [2023-05-23 Tue 01:21]
******** DONE Singleton ou non mapped ? nonVirtPosition
CLOSED: [2023-05-23 Tue 01:22]
❯ samtools bam2fq  -1 63003856_chr22_init1.fq.gz -2 63003856_chr22_init2.fq.gz -0 both -s single.fq.gz  -n 63003856_chr22_sorted.bam
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ wc -l 63003856_chr22_init* both single.fq.gz
   403540 63003856_chr22_init1.fq.gz
   404788 63003856_chr22_init2.fq.gz
        0 both
        0 single.fq.gz
   808328 total
******** Problème d'aligner car les reads sont bien dans le .fastq ?
Ex:
 zgrep "A00853:477:HMLWYDSX3:2:2624:2826:18630" 63003856_chr22_{1,2}.fq.gz
63003856_chr22_1.fq.gz:@A00853:477:HMLWYDSX3:2:2624:2826:18630
63003856_chr22_2.fq.gz:@A00853:477:HMLWYDSX3:2:2624:2826:18630
******* DONE Refaire la manip sur bam chr22 non modifié + mail Alexis
CLOSED: [2023-05-23 Tue 23:27]
[1]_samtools view ~/code/bisonex/out/63003856/preprocessing/mapped/63003856_S135.bam NC_000022.11  -f 0x2 -o 63003856_chr22.bam
 Les reads sont bien tous mappé
samtools flagstat 63003856_chr22.bam
2609214 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
17104 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2609214 + 0 mapped (100.00% : N/A)
[2] samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
flagstat idem
[3] samtools fastq -1 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -n 63003856_chr22_sorted.bam
[M::bam2fq_mainloop] discarded 0 singletons
[M::bam2fq_mainloop] processed 2592110 reads
Nombre de reads ok (= la moitié) et les séquences ne sont pas identiques pour le premier read (= on n'a pas 2x la même chose)
 echo $(zcat 63003856_chr22_1.fq.gz | wc -l)/4 | bc
1296055
 echo $(zcat 63003856_chr22_2.fq.gz | wc -l)/4 | bc
1296055
 [4]
 bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz  -o wtf.bam
 [M::bwa_idx_load_from_disk] read 0 ALT contigs
[M::process] read 1752492 sequences (240000014 bp)...
[M::mem_pestat] # candidate unique pairs for (FF, FR, RF, RR): (12, 702821, 0, 18)
[M::mem_pestat] analyzing insert size distribution for orientation FF...
[M::mem_pestat] (25, 50, 75) percentile: (54, 197, 268)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 696)
[M::mem_pestat] mean and std.dev: (163.92, 129.71)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 910)
[M::mem_pestat] analyzing insert size distribution for orientation FR...
[M::mem_pestat] (25, 50, 75) percentile: (129, 175, 233)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 441)
[M::mem_pestat] mean and std.dev: (185.56, 75.37)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 545)
[M::mem_pestat] skip orientation RF as there are not enough pairs
[M::mem_pestat] analyzing insert size distribution for orientation RR...
[M::mem_pestat] (25, 50, 75) percentile: (56, 192, 239)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 605)
[M::mem_pestat] mean and std.dev: (158.00, 110.30)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 788)
[M::mem_pestat] skip orientation FF
[M::mem_pestat] skip orientation RR
[M::process] read 839618 sequences (114952336 bp)...
[M::mem_process_seqs] Processed 1752492 reads in 375.714 CPU sec, 17.645 real sec
[M::mem_pestat] # candidate unique pairs for (FF, FR, RF, RR): (0, 336379, 0, 5)
[M::mem_pestat] skip orientation FF as there are not enough pairs
[M::mem_pestat] analyzing insert size distribution for orientation FR...
[M::mem_pestat] (25, 50, 75) percentile: (128, 174, 232)
[M::mem_pestat] low and high boundaries for computing mean and std.dev: (1, 440)
[M::mem_pestat] mean and std.dev: (184.73, 74.63)
[M::mem_pestat] low and high boundaries for proper pairs: (1, 544)
[M::mem_pestat] skip orientation RF as there are not enough pairs
[M::mem_pestat] skip orientation RR as there are not enough pairs
[M::mem_process_seqs] Processed 839618 reads in 183.039 CPU sec, 7.961 real sec
[main] Version: 0.7.17-r1188
[main] CMD: bwa mem -t 24 -o wtf.bam /Work/Projects/bisonex/data/genome/GRCh38.p13/bwa/genomeRef 63003856_chr22_1.fq.gz 63003856_chr22_2.fq.gz
[main] Real time: 38.278 sec; CPU: 565.821 sec
Bon nombre de reads pourtant
 samtools flagstat wtf.bam
2611059 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
18949 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2611058 + 0 mapped (100.00% : N/A)
2592109 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2590970 + 0 properly paired (99.96% : N/A)
2592108 + 0 with itself and mate mapped
1 + 0 singletons (0.00% : N/A)
458 + 0 with mate mapped to a different chr
63 + 0 with mate mapped to a different chr (mapQ>=5)
$ samtools sort -@24 -o wtf_sorted.bam wtf.sam
[bam_sort_core] merging from 0 files and 24 in-memory blocks...
 samtools flagstat wtf.bam
2611059 + 0 in total (QC-passed reads + QC-failed reads)
2592110 + 0 primary
0 + 0 secondary
18949 + 0 supplementary
0 + 0 duplicates
0 + 0 primary duplicates
2611058 + 0 mapped (100.00% : N/A)
2592109 + 0 primary mapped (100.00% : N/A)
2592110 + 0 paired in sequencing
1296055 + 0 read1
1296055 + 0 read2
2590970 + 0 properly paired (99.96% : N/A)
2592108 + 0 with itself and mate mapped
1 + 0 singletons (0.00% : N/A)
458 + 0 with mate mapped to a different chr
63 + 0 with mate mapped to a different chr (mapQ>=5)
Effectivement, ce n'est pas un problème d'IGV
❯ samtools mpileup 63003856_chr22.bam -r NC_000022.11:42213078-42213078
[mpileup] 1 samples in 1 input files
NC_000022.11	42213078	N	19	TTtTTtTTTtTTtTtTttt	kkk_kkkkFkFF_FkFkQk
bisonex/code/BamScissors.jl on  bamscissors [!?] via ஃ v1.9.0
❯ samtools mpileup aligned/wtf_sorted.bam -r NC_000022.11:42213078-42213078
[mpileup] 1 samples in 1 input files
NC_000022.11	42213078	N	5	TTtTT	_FkFF
******* DONE Regarder où ont été aligné les reads (nouveau run)
CLOSED: [2023-05-24 Wed 23:18]
******** DONE Préparation
CLOSED: [2023-05-24 Wed 21:59]
On relance le pipeline pour avoir un BAM propre
On garde les reads non mappé à partsir de la sortie d'applybqsr
#+begin_src sh
NXF_OPTS=-D"user.name=apraga" nextflow run main.nf -c nextflow.config  -profile standard,helios --input="/Work/Projects/bisonex/centogene/fastq/2200467051_63003856/63003856_S135_R{1,2}_001.fastq.gz" --outdir=out -bg
cd out/63003856_S135_R/preprocessing/applybqsr/
samtools view 63003856_S135_R.bam NC_000022.11  -f 0x2 -o 63003856_chr22.bam
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
samtools fastq -1 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -n 63003856_chr22_sorted.bam
make run BG= READS="out/63003856_S135_R/preprocessing/applybqsr/63003856_chr22_{1,2}.fq.gz"
cd out/63003856_chr22/preprocessing/mapped/
samtools index 63003856_chr22.bam
samtools mpileup 63003856_chr22.bam -r NC_000022.11:42213078-42213078
#+end_src
On récupère les 2 bam dans
#+begin_src
cd /home/alex/recherche/bisonex/code/BamScissors.jl/
rsync -avz meso:/Work/Users/apraga/bisonex/out/63003856_chr22/preprocessing/mapped data
rsync -avz meso:/Work/Users/apraga/bisonex/out/63003856_S135_R/preprocessing/applybqsr/ data/init/
#+end_src
******** Vérification que le reads est ailleurs
On cherche un read manquant dans le second alignement
#+begin_src sh
samtools view data/init/63003856_chr22.bam | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
#+end_src
#+RESULTS:
: A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NC_000022.11	42212845	0	151M	=	42212883	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	ACC+FBCDCBBBAEAEDEEBBCCCECACBAEBEBDCCBCBFDCCCCFACEBEBCEEDCCCCFDCAEDCACBCEBBCFEACCFBDCACDCBCEBDBBCFEEDCCCFAFEACECCCECAEEDCADCBEDC7BEBCCCFBAFDCECCFBEAACA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
: A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NC_000022.11	42212883	0	151M	=	42212845	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	AADECCCBDCBFCE<?CDEEEEBDEACDEAC;:BFBCBCDCCBEAEACAEFCCEAFBCBCCDEECBDBCECBEECCEACDEEBBFGDEFGCCFFFFCFCCEFBFDCFCDAAEBEE:CECBABBEBEE;DBFCCCDBCDBCCBBC?@BEEDA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
#+begin_src sh
samtools view data/mapped/63003856_chr22.bam | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
#+end_src
#+RESULTS:
: A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NW_014040930.1	115017	0	151M	=	115055	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	ACC+FBCDCBBBAEAEDEEBBCCCECACBAEBEBDCCBCBFDCCCCFACEBEBCEEDCCCCFDCAEDCACBCEBBCFEACCFBDCACDCBCEBDBBCFEEDCCCFAFEACECCCECAEEDCADCBEDC7BEBCCCFBAFDCECCFBEAACA	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
: A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NW_014040930.1	115055	0	151M	=	115017	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	AADECCCBDCBFCE<?CDEEEEBDEACDEAC;:BFBCBCDCCBEAEACAEFCCEAFBCBCCDEECBDBCECBEECCEACDEEBBFGDEFGCCFFFFCFCCEFBFDCFCDAAEBEE:CECBABBEBEE;DBFCCCDBCDBCCBBC?@BEEDA	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
Effectivement, on aligne sur une zonne supprimée !
******* DONE Corriger la qualité: non
CLOSED: [2023-05-24 Wed 22:19]
******** DONE Comparaison avec le fastq de référénce : qualité !!
CLOSED: [2023-05-24 Wed 22:17]
#+begin_src sh
cd /Work/Users/apraga/bisonex/work/6e/8548fc90263830bf677f36585f11dc
zgrep -A 3 "A00853:477:HMLWYDSX3:1:1413:4390:28573" 63003856_chr22_1.fq.gz
#+end_src
@A00853:477:HMLWYDSX3:1:1413:4390:28573
AGGGTTACCACCACCACCCTGACAGGAGATATTCTAGGAGTACTCAAGAGCATCAGGGGATGGCTGGTAGCCTAGAAGGAACCACAAGGCCCAATGTCTTGGTTAGTCAAACCAATGAATTAGCTAGCAGGGGCCTTCTGAACAAAAGCAT
+
ADEEB@?CBBCCBDCBDCCCFBD;EEBEBBABCEC:EEBEAADCFCDFBFECCFCFFFFCCGFEDGFBBEEDCAECCEEBCECBDBCEEDCCBCBFAECCFEACAEAEBCCDCBCBFB:;CAEDCAEDBEEEEDC?<ECFBCDBCCCEDAA
#+begin_src
zgrep -A 3 "A00853:477:HMLWYDSX3:1:1413:4390:28573" /Work/Projects/bisonex/centogene/fastq/2200467051_63003856/63003856_S135_R1_001.fastq.gz
#+end_src
#+RESULTS:
: @A00853:477:HMLWYDSX3:1:1413:4390:28573 1:N:0:ATTCCACACA+TAGGCGATTG
AGGGTTACCACCACCACCCTGACAGGAGATATTCTAGGAGTACTCAAGAGCATCAGGGGATGGCTGGTAGCCTAGAAGGAACCACAAGGCCCAATGTCTTGGTTAGTCAAACCAATGAATTAGCTAGCAGGGGCCTTCTGAACAAAAGCAT
: +
: FFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFF
******** DONE Regarder la qualité après bwa mem vs applybqsr: différente
CLOSED: [2023-05-24 Wed 22:18]
Sur le mésocentre, dans /Work/Users/apraga/bisonex/out/63003856_S135_R/preprocessing
$ samtools view mapped/63003856_S135_R.bam NC_000022.11 | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NC_000022.11	42212845	0	151M	=	42212883	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	FFF,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NC_000022.11	42212883	0	151M	=	42212845	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	FFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
samtools view applybqsr/63003856_S135_R.bam NC_000022.11 | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NC_000022.11	42212845	0	151M	=	42212883	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	ACC+FBCDCBBBAEAEDEEBBCCCECACBAEBEBDCCBCBFDCCCCFACEBEBCEEDCCCCFDCAEDCACBCEBBCFEACCFBDCACDCBCEBDBBCFEEDCCCFAFEACECCCECAEEDCADCBEDC7BEBCCCFBAFDCECCFBEAACA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NC_000022.11	42212883	0	151M	=	42212845	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	AADECCCBDCBFCE<?CDEEEEBDEACDEAC;:BFBCBCDCCBEAEACAEFCCEAFBCBCCDEECBDBCECBEECCEACDEEBBFGDEFGCCFFFFCFCCEFBFDCFCDAAEBEE:CECBABBEBEE;DBFCCCDBCDBCCBBC?@BEEDA	MC:Z:151M	MD:Z:151	PG:Z:MarkDuplicates	RG:Z:sample	NM:i:0	AS:i:151	XS:i:151
******** DONE Réaligner à partir de la sortie de bwa mem
CLOSED: [2023-05-24 Wed 22:32]
#+begin_src sh
cd out/63003856_S135_R/preprocessing/mapped/
samtools view 63003856_S135_R.bam NC_000022.11  -f 0x2 -o 63003856_chr22.bam
samtools sort -n 63003856_chr22.bam -o 63003856_chr22_sorted.bam
samtools fastq -1 63003856_chr22_1.fq.gz -2 63003856_chr22_2.fq.gz -0 /dev/null -s /dev/null -n 63003856_chr22_sorted.bam
#+end_src
ON vérifie la qualité
#+begin_src
zgrep -A 3 "A00853:477:HMLWYDSX3:1:1413:4390:28573" 63003856_chr22_1.fq.gz
#+end_src
#+RESULTS:
: @A00853:477:HMLWYDSX3:1:1413:4390:28573
: AGGGTTACCACCACCACCCTGACAGGAGATATTCTAGGAGTACTCAAGAGCATCAGGGGATGGCTGGTAGCCTAGAAGGAACCACAAGGCCCAATGTCTTGGTTAGTCAAACCAATGAATTAGCTAGCAGGGGCCTTCTGAACAAAAGCAT
: +
: FFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFF
#+begin_src
NXF_OPTS=-D"user.name=apraga" nextflow run main.nf -c nextflow.config  -profile standard,helios -resume  --input="out/63003856_S135_R/preprocessing/mapped/63003856_chr22_{1,2}.fq.gz" --outdir=out/63003856_chr22-from-mapped
#+end_src
Puis ::
#+begin_src
cd /Work/Users/apraga/bisonex/out/63003856_chr22-from-mapped/63003856_chr22/preprocessing/mapped
samtools view 63003856_chr22.bam | rg "A00853:477:HMLWYDSX3:1:1413:4390:28573"
#+end_src
#+RESULTS:
: A00853:477:HMLWYDSX3:1:1413:4390:28573	163	NW_014040930.1	115017	0	151M	=	115055	189	CCCAGGGGCCCCAGTGGGGATTTTCTAATAGAGACCCAATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACT	FFF,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
: A00853:477:HMLWYDSX3:1:1413:4390:28573	83	NW_014040930.1	115055	0	151M	=	115017	-189	ATGCTTTTGTTCAGAAGGCCCCTGCTAGCTAATTCATTGGTTTGACTAACCAAGACATTGGGCCTTGTGGTTCCTTCTAGGCTACCAGCCATCCCCTGATGCTCTTGAGTACTCCTAGAATATCTCCTGTCAGGGTGGTGGTGGTAACCCT	FFFFFFFFFFFFFF::FFFFFFFFFFFFFFF::FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF:FFFFFFFFFFF:FFFFFFFFFFFFFFFFFFFFFFF	NM:i:0	MD:Z:151	MC:Z:151M	AS:i:151	XS:i:151	RG:Z:sample
******* DONE Aligner sur génome de référence limité au chromosome 22
CLOSED: [2023-05-24 Wed 23:18]
******** KILL Test données non modifiées
CLOSED: [2023-05-24 Wed 23:18]
/Work/Users/apraga/bisonex/tests/bamscissors
#+begin_src
cd /Work/Groups/bisonex/data/genome/GRCh38.p13/
mkdir chr22/
samtools faidx genomeRef.fna NC_000022.11 > chr22/chr22.fna
cd chr22
samtools faidx chr22.fna
bwa index chr22.fna
#+end_src
#+begin_src
cd /Work/Users/apraga/bisonex/tests/bamscissors
ln -s ../../out/63003856_S135_R/preprocessing/applybqsr/63003856_chr22_{1,2}.fq.gz .
srun -c 24 -p smp -t 1:00:00 --pty bash
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/chr22/chr22.fna 63003856_chr22_1.fq.gz 63003856_chr22_1.fq.gz -o smallref.sam
#+end_src
******** DONE Test données modifiées: ok
CLOSED: [2023-05-24 Wed 23:18]
Données dans data/init
#+begin_src sh
time julia insertVariant.jl
rsync -avz data/init/*.fq.gz meso:/Work/Users/apraga/bisonex/tests/bamscissors/
#+end_src
#+begin_src
srun -c 24 -p smp -t 1:00:00 --pty bash
bwa mem -t 24 /Work/Projects/bisonex/data/genome/GRCh38.p13/chr22/chr22.fna 63003856_chr22_1.fq.gz 63003856_chr22_1.fq.gz | samtools sort -@24 - -o smallref.bam
#+end_src
#+begin_src
rsync -avz meso:/Work/Users/apraga/bisonex/tests/bamscissors/smallref.bam mapped/
#+end_src
****** DONE Test haplotypecaller 1 variant
CLOSED: [2023-05-29 Mon 15:38]
****** DONE Test haplotypecaller tous les variants
***** TODO PHase 3 : tous les SNV
**** TODO Test Indel
*** Divers
**** DONE Vérifier nombre de reads fastq - bam
CLOSED: [2022-10-09 Sun 22:31]

File addition: paludisme.org (----------)

[13.1207]

#+title: Paludisme
#+options: toc:nil author:nil date:nil
#+latex_header: \usepackage{tabularx}
#+latex_header: \usepackage{booktabs}
#+latex_header: \usepackage{enumitem}
#+latex_header: \usepackage[margin=1cm]{geometry}
#+latex_header_extra: \usepackage{adjustbox}
#+LATEX: \def\dec{$\searrow{}$}
#+LATEX: \def\inc{$\nearrow{}$}
#+LATEX: \newcommand{\tabitem}{~~\llap{\textbullet}~~}
#+LATEX: \newcommand{\ttabitem}{~~~~~~\llap{$\square$}~~}
#+LATEX: \newcommand{\tttabitem}{~~~~~~~~\llap{-}~~}
#+begin_table
#+LATEX: \centering
#+LATEX: \adjustbox{max width=\linewidth}{
#+ATTR_LATEX: :center nil
|                            |                             | Accès simple                    |                             |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
|                            | Adulte                      | Femme enceinte                  | Enfants                     | Remarques                      |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Combinaison d'artémisinine | 1 intention au choix        | Hospitalisation                 | 1re intention               |                                |
| – artémether               | – 3 jours                   | – CI 1er trimestre              |                             | – ATCD d'atteintes cardiaques  |
| -luméfantrine              | Avec un repas gras          | et allaitement,                 |                             | congénitales                   |
|                            | (malabsorption              |                                 |                             | – ATCD de QT long              |
| – (arténimol               | À jeun depuis 3 heures      | -  CI pendant                   |                             |                                |
| pipéraquine)               |                             | la grossesse                    |                             | – Pas plus de 2 traitements/an |
|                            |                             | et allaitement                  |                             | – 2 mois au moins              |
|                            |                             |                                 |                             | entre 2 traitements            |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Atovaquone-                | 2e intention                | Alternative pour le             | 2e intention                | IR sévère                      |
| proguanil                  | – Si CI aux CTA             | 1er trimestre                   |                             |                                |
|                            | – 3 j avec un repas gras    |                                 |                             |                                |
|                            | (mauvaise biodisponibilité) |                                 |                             |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Méfloquine                 | Non recommandée             | Non recommandée                 | 3e intention                |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Chloroquine :              | 3 jours                     | 3 jours                         | 3 jours                     |                                |
| espèce autre que           |                             |                                 |                             |                                |
| P. falciparum              |                             |                                 |                             |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Quinine                    | 3e intention si IV          | 2e intention                    | 3e intention si IV          |                                |
|                            | nécessaire :                | Utilisable tout au              | nécessaire                  |                                |
|                            | Relais PO dès que possible  | long de la grossesse            |                             |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
|                            |                             | Accès grave                     |                             |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Artésunate                 | 7 jours                     | Balance bénéfice/embryotoxicité |                             |                                |
|----------------------------+-----------------------------+---------------------------------+-----------------------------+--------------------------------|
| Quinine                    | Si artésunate indisponible  |                                 | Si artésunate  indisponible |                                |
#+LATEX: }
#+end_table

Replacement in notes/medecine/20230528225221-classification_bacteries.org at line 43 [5.20341]
B:BD[5.21797] → [5.21797:21838]
```
[[../images/microbiologie/aerobies.png]]
```
[5.21797]
[5.21838]
```
[[../../images/microbiologie/aerobies.png]]
```
Replacement in notes/medecine/20230528225221-classification_bacteries.org at line 54 [5.20341]
B:BD[5.22166] → [5.22166:22209]
```
[[../images/microbiologie/anaerobies.png]]
```
[5.22166]
[5.22209]
```
[[../../images/microbiologie/anaerobies.png]]
```
Replacement in notes/medecine/20230528225221-classification_bacteries.org at line 68 [5.20341]
B:BD[5.22660] → [5.22660:22699]
```
[[../images/microbiologie/autres.png]]
```
[5.22660]
[5.22699]
```
[[../../images/microbiologie/autres.png]]
```

Replacement in notes/index.org at line 1 [15.368291]

B:BD[15.368291] → [2.140:199]

[[file:20230511180745-bacteriologie.html][Bactériologie]]

[15.368291]

Bactériologie
- [[./medecine/20230528225221-classification_bacteries.html][Classification bacteries]]
- [[./medecine/20230528225531-bacteries.html][Bacteries]]
- [[./medecine/20230528235213-maladies_infectieuses.html][Maladies infectieuses]]
- [[./medecine/20230528235406-antibiotiques.html][Antibiotiques]]
- [[./medecine/20230528235124-culture.html][Culture]]
Virus
- [[./medecine/20230528233600-norovirus.html][Norovirus]]

File move: 20230511180745-bacteriologie.org → 20230511180745-microbiologie.org
BF:BFD[22.29] → [23.4578:4634]
BF:BF[23.4634] → [23.3565:3565]
[22.29]
[23.3565]

Replacement in notes/20230511180745-microbiologie.org at line 8 [23.3565]

B:BD[5.24303] → [5.24303:24567]

[[file:medecine/20230528225221-classification_bacteries.org]]
[[file:medecine/20230528225531-bacteries.org]]
[[file:medecine/20230528235124-culture.org]]
[[file:medecine/20230528235213-maladies_infectieuses.org]]
[[file:medecine/20230528235406-antibiotiques.org]]

[5.24303]

[5.24567]

- [[file:medecine/20230528225221-classification_bacteries.org][Classification bactéries]]
- [[file:medecine/20230528225531-bacteries.org][Bactéries]]
- [[file:medecine/20230528235124-culture.org][Culture]]
- [[file:medecine/20230528235213-maladies_infectieuses.org][Maladies infectieuses]]
- [[file:medecine/20230528235406-antibiotiques.org][Antibiotiques]]

Insertion in notes/20230511171011-gentoo.org at line 86 [25.75]

[24.856]

* Erreurs
** Missing keyword
!!! All ebuilds that could satisfy "=dev-lang/julia-1.9.0" have been masked.
!!! One of the following masked packages is required to complete your request:
- dev-lang/julia-1.9.0::gentoo (masked by: missing keyword)
Utiliser emerge --autounmask

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      need [src, "notes/20230511180745-bacteriologie.org"]

[26.1669]

[26.1728]

      need [src, "notes/20230511180745-microbiologie.org"]