---
# System prepended metadata

title: Protist TEs

---

# Protist TEs

Got a bunch of genome assemblies for holozoan protists from Alex Tice. Little has been done to examine these groups for TE content and impact. Will have Angela examine them for TEs. I will parallel her as a way of teaching her the basics.

She's reading the necessary papers and has some basic information. 

In D:\Dropbox\Angela Ximena Correa Perdomo\
Grau-Bove.2017.elife.pdf
Hehenberger.et.al.2017.pdf
holozoan.metadata.tsv
project ideas.txt

Zipped the files and have renamed them as below. Work will be in /lustre/scratch/daray/protistan_holozoa

Abeoforma_whisleri.fa.gz = aWhi.fa.gz
Capsaspora_owczarzaki.fa.gz = cOwc.fa.gz
Chromosphaera_perkinsii.fa.gz = cPer.fa.gz
Corallochytrium_limacisporum.fa.gz = cLim.fa.gz
Creolimax_fragrantissima.fa.gz = cFra.fa.gz
Ichthyophonus_hoferi.fa.gz = iHof.fa.gz
Ministeria_vibrans.fa.gz = mVib.fa.gz
Monosiga_brevicollis.fa.gz = mBre.fa.gz
Pigoraptor_chileana.fa.gz = pChi.fa.gz
Pigoraptor_vietnamica.fa.gz = pVie.fa.gz
Pirum_gemmata.fa.gz = pGem.fa.gz
Salpingoeca_rosetta.fa.gz = sRos.fa.gz
Sphaeroforma_arctica.fa.gz = sArc.fa.gz

LIST="aWhi cOwc cPer cLim cFra iHof mVib mBre pChi pVie pGem sRos sArc"


edit curation_templates/rmodel_template.sh to:

```
#!/bin/bash 
#SBATCH --job-name=<NAME>_rmod
#SBATCH --output=%x.%j.out
#SBATCH --error=%x.%j.err
#SBATCH --partition=nocona
#SBATCH --nodes=1
#SBATCH --ntasks=64


. ~/conda/etc/profile.d/conda.sh
conda activate

TAXON=<NAME>

cd /lustre/scratch/daray/protistan_holozoa/te_curations

python ../curation_templates/rmodel.py -g /lustre/scratch/daray/protistan_holozoa/assemblies/${TAXON}.fa.gz -p 64 -b 50 -w /lustre/scratch/daray/protistan_holozoa/te_curations/${TAXON}/

```
Do the following to set up.

```
cd /lustre/scratch/daray/protistan_holozoa
LIST="aWhi cOwc cPer cLim cFra iHof mVib mBre pChi pVie pGem sRos sArc"

for NAME in $LIST; do
mkdir -p te_curations/$NAME/te-aid
done

cp -r /lustre/scratch/daray/curation_templates/ . 

for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rmodel_template.sh >te_curations/${NAME}_rmodel.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/RM_template_mod.sh >te_curations/${NAME}_RM.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/extend_align_template.sh >te_curations/${NAME}_extend_align.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/repeatclassifier_template.sh >te_curations/${NAME}_repeatclassifier.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rm2bed_template_mod.sh >te_curations/${NAME}_rm2bed.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done
```

Angela will need to update her files to have her username instead of mine. 

```
cd /lustre/scratch/daray/protistan_holozoa/te_curations
LIST="aWhi cOwc cPer cLim cFra iHof mVib mBre pChi pVie pGem sRos sArc"
for NAME in $LIST; do sed -i "s/daray/acorreap/g" ${NAME}_rmodel.sh; done
for NAME in $LIST; do sed -i "s/daray/acorreap/g" ${NAME}_RM.sh; done
for NAME in $LIST; do sed -i "s/daray/acorreap/g" ${NAME}_extend_align.sh; done
for NAME in $LIST; do sed -i "s/daray/acorreap/g" ${NAME}_repeatclassifier.sh; done
for NAME in $LIST; do sed -i "s/daray/acorreap/g" ${NAME}_rm2bed.sh; done
for NAME in $LIST; do sed -i "s/daray/acorreap/g" ${NAME}_TEcurate.sh; done
```
Start RepeatModeler

```
cd te_curations
for NAME in $LIST; do sbatch ${NAME}_rmodel.sh; done
```

Four more assemblies have been added. Need to add those as well.

pUst, hCla, nSp, nDam

```
LIST="pUst hCla nSp nDam"
cd /lustre/scratch/daray/protistan_holozoa
for NAME in $LIST; do
mkdir -p te_curations/$NAME/te-aid
done

cp -r /lustre/scratch/daray/curation_templates/ . 

for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rmodel_template.sh >te_curations/${NAME}_rmodel.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/RM_template_mod.sh >te_curations/${NAME}_RM.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/extend_align_template.sh >te_curations/${NAME}_extend_align.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/repeatclassifier_template.sh >te_curations/${NAME}_repeatclassifier.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rm2bed_template_mod.sh >te_curations/${NAME}_rm2bed.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done

cd te_curations
for NAME in $LIST; do sbatch ${NAME}_rmodel.sh; done

```

Updated list: LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"

```
LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"
```


Track progress using squeue and when all are done, run the extension script. This script runs RAM and generates the putative TE consensus sequences. 

** Angela will need to set up her conda environment to match mine. **

Setting up extend_env conda environment:

```
cd ~ 
cp /home/daray/extend_env.yml .
conda env create -f extend_env.yml
```

Now, the extend_align script should work. 

```
cd /lustre/scratch/daray/protistan_holozoa/te_curations
LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"

for NAME in $LIST; do sbatch ${NAME}_extend_align.sh; done
```
Paste squeue output here.

I forgot to. All finished.

While the extend_align scripts are running, queue the next steps to run after they're completed.

```
cd /lustre/scratch/daray/protistan_holozoa/te_curations
LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"

for NAME in $LIST; do sbatch ${NAME}_repeatclassifier.sh; done
```

12218469     nocona     654  PD     daray aWhi_clas        0:00      1      1   normal  (Priority)
12218470     nocona     654  PD     daray cOwc_clas        0:00      1      1   normal  (Priority)
12218471     nocona     654  PD     daray cPer_clas        0:00      1      1   normal  (Priority)
12218472     nocona     654  PD     daray cLim_clas        0:00      1      1   normal  (Priority)
12218473     nocona     654  PD     daray cFra_clas        0:00      1      1   normal  (Priority)
12218474     nocona     654  PD     daray hCla_clas        0:00      1      1   normal  (Priority)
12218475     nocona     654  PD     daray iHof_clas        0:00      1      1   normal  (Priority)
12218476     nocona     654  PD     daray mVib_clas        0:00      1      1   normal  (Priority)
12218477     nocona     654  PD     daray mBre_clas        0:00      1      1   normal  (Priority)
12218478     nocona     654  PD     daray nSp_class        0:00      1      1   normal  (Priority)
12218479     nocona     654  PD     daray nDam_clas        0:00      1      1   normal  (Priority)
12218480     nocona     654  PD     daray pChi_clas        0:00      1      1   normal  (Priority)
12218481     nocona     654  PD     daray pVie_clas        0:00      1      1   normal  (Priority)
12218482     nocona     654  PD     daray pGem_clas        0:00      1      1   normal  (Priority)
12218483     nocona     654  PD     daray pUst_clas        0:00      1      1   normal  (Priority)
12218484     nocona     654  PD     daray sRos_clas        0:00      1      1   normal  (Priority)
12218485     nocona     654  PD     daray sArc_clas        0:00      1      1   normal  (Priority)




While the repeatclassifier scripts are running, queue the next steps to run after they're completed.
```
sbatch --dependency=afterok:12218469 aWhi_TEcurate.sh
sbatch --dependency=afterok:12218470 cOwc_TEcurate.sh
sbatch --dependency=afterok:12218471 cPer_TEcurate.sh
sbatch --dependency=afterok:12218472 cLim_TEcurate.sh
sbatch --dependency=afterok:12218473 cFra_TEcurate.sh
sbatch --dependency=afterok:12218474 hCla_TEcurate.sh
sbatch --dependency=afterok:12218475 iHof_TEcurate.sh
sbatch --dependency=afterok:12218476 mVib_TEcurate.sh
sbatch --dependency=afterok:12218477 mBre_TEcurate.sh
sbatch --dependency=afterok:12218478 nSp_TEcurate.sh
sbatch --dependency=afterok:12218479 nDam_TEcurate.sh
sbatch --dependency=afterok:12218480 pChi_TEcurate.sh
sbatch --dependency=afterok:12218481 pVie_TEcurate.sh
sbatch --dependency=afterok:12218482 pGem_TEcurate.sh
sbatch --dependency=afterok:12218483 pUst_TEcurate.sh
sbatch --dependency=afterok:12218484 sRos_TEcurate.sh
sbatch --dependency=afterok:12218485 sArc_TEcurate.sh

```

12218528     nocona     633  PD     daray aWhi_TEcu        0:00      1      1   normal  (Dependency)
12218529     nocona     633  PD     daray cOwc_TEcu        0:00      1      1   normal  (Dependency)
12218530     nocona     633  PD     daray cPer_TEcu        0:00      1      1   normal  (Dependency)
12218531     nocona     633  PD     daray cLim_TEcu        0:00      1      1   normal  (Dependency)
12218532     nocona     633  PD     daray cFra_TEcu        0:00      1      1   normal  (Dependency)
12218533     nocona     633  PD     daray hCla_TEcu        0:00      1      1   normal  (Dependency)
12218534     nocona     633  PD     daray iHof_TEcu        0:00      1      1   normal  (Dependency)
12218535     nocona     633  PD     daray mVib_TEcu        0:00      1      1   normal  (Dependency)
12218536     nocona     633  PD     daray mBre_TEcu        0:00      1      1   normal  (Dependency)
12218537     nocona     633  PD     daray nSp_TEcur        0:00      1      1   normal  (Dependency)
12218538     nocona     633  PD     daray nDam_TEcu        0:00      1      1   normal  (Dependency)
12218539     nocona     633  PD     daray pChi_TEcu        0:00      1      1   normal  (Dependency)
12218540     nocona     633  PD     daray pVie_TEcu        0:00      1      1   normal  (Dependency)
12218541     nocona     633  PD     daray pGem_TEcu        0:00      1      1   normal  (Dependency)
12218542     nocona     633  PD     daray pUst_TEcu        0:00      1      1   normal  (Dependency)
12218543     nocona     633  PD     daray sArc_TEcu        0:00      1      1   normal  (Dependency)
12218544     nocona     633  PD     daray sRos_TEcu        0:00      1      1   normal  (Dependency)



### Oct 2, 2023
Met with Alex and he provided another set of assemblies. n=15. This time they're much more closely related and that could make this a bit more manageable.

All are from Heterolobosea and most are free-living human parasites.

14 are from genus Naeglaria and one from Acasia

From the metadata file provide:

Taxon		Lifstyle	Habitat	Life Cycle States Present	Accession
Neovahlkampfia damarascottae	CCAP 1588/7	freeliving	freshwater	amoeba/cyst	GCA_016618085.1 *illumina only
Heteramoeba clara	N/A	freeliving	brackish water	amoeba/cyst/flagellate	N/A Not public
Paravahlkampfia ustiana	N/A	freeliving	freshwater	amoeba/cyst	N/A Not public
Acrasis kona	ATCC MYA-3509	freeliving	dead plant tissue	amoeba/cyst/sporocarp **exhibits aggregative multicellularity	GCA_026419775.1
Naegleria gruberi	NEG-M	freeliving	freshwater	amoeba/cyst/flagellate	GCA_000004985.1
Naegleria fowleri	ATCC 30863	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_000499105.1
Naegleria fowleri	NF_KFSHRC_1	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_902703645.1
Naegleria fowleri	NF1	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_027563095.1
Naegleria fowleri	AR12	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_027563075.1
Naegleria fowleri	PA34	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_027563055.1
Naegleria fowleri	kurume	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_020891285.1
Naegleria fowleri	Ty	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_014843625.1
Naegleria fowleri	ATCC 30894	freeliving/opportunistic human pathogen	freshwater	amoeba/cyst/flagellate	GCA_008403515.1
Naegleria lovaniensis	Lova6	freeliving	freshwater	amoeba/cyst/flagellate	GCA_027563035.1
Naegleria lovaniensis	Lova7	freeliving	freshwater	amoeba/cyst/flagellate	GCA_027563015.1
Naegleria lovaniensis	F9	freeliving	freshwater	amoeba/cyst/flagellate	GCA_027562995.1
Naegleria lovaniensis	NL_76_15_250	freeliving	freshwater	amoeba/cyst/flagellate	GCA_022530875.1
Naegleria lovaniensis	ATCC 30569	freeliving	freshwater	amoeba/cyst/flagellate	GCA_003324165.2

new list 
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"

The plan is to concentrate on these for the moment because of the, presumed, reduced diversity in TE content. Will make it easier on Angela.

```
cd /lustre/scratch/daray/protistan_holozoa/
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"

for NAME in $LIST; do
mkdir -p te_curations/$NAME/te-aid
done

cp -r /lustre/scratch/daray/curation_templates/ . 

for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rmodel_template.sh >te_curations/${NAME}_rmodel.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/RM_template_mod.sh >te_curations/${NAME}_RM.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/extend_align_template.sh >te_curations/${NAME}_extend_align.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/repeatclassifier_template.sh >te_curations/${NAME}_repeatclassifier.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rm2bed_template_mod.sh >te_curations/${NAME}_rm2bed.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done

cd te_curations
for NAME in $LIST; do sbatch ${NAME}_rmodel.sh; done
```
12249694     nocona    1078  PD     daray aKon_rmod        0:00      1     64   normal  (Priority)
12249695     nocona    1078  PD     daray nFow_AR12        0:00      1     64   normal  (Priority)
12249696     nocona    1078  PD     daray nFow_ATCC        0:00      1     64   normal  (Priority)
12249697     nocona    1078  PD     daray nFow_kuru        0:00      1     64   normal  (Priority)
12249698     nocona    1078  PD     daray nFow_NF1_        0:00      1     64   normal  (Priority)
12249699     nocona    1078  PD     daray nFow_NF_K        0:00      1     64   normal  (Priority)
12249700     nocona    1078  PD     daray nFow_PA34        0:00      1     64   normal  (Priority)
12249701     nocona    1078  PD     daray nFow_Ty_r        0:00      1     64   normal  (Priority)
12249702     nocona    1078  PD     daray nGru_NEGM        0:00      1     64   normal  (Priority)
12249703     nocona    1078  PD     daray nFow_ATCC        0:00      1     64   normal  (Priority)
12249704     nocona    1078  PD     daray nLov_ATCC        0:00      1     64   normal  (Priority)
12249705     nocona    1078  PD     daray nLov_F9_r        0:00      1     64   normal  (Priority)
12249706     nocona    1078  PD     daray nLov_Lova        0:00      1     64   normal  (Priority)
12249707     nocona    1078  PD     daray nLov_Lova        0:00      1     64   normal  (Priority)
12249708     nocona    1078  PD     daray nLov_NL_7        0:00      1     64   normal  (Priority)


## Thought about it and decided that 10 copies is way too few for these organisms. Reset MINCOPY variable in the TEcurate_template file to MINCOPY=2.

Now need to re-run all of my curations for the species that are already complete.
```
cd /lustre/scratch/daray/protistan_holozoa/te_curations
LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"

cd /lustre/scratch/daray/protistan_holozoa/
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done

cd /lustre/scratch/daray/protistan_holozoa/te_curations
for NAME in $LIST; do rm -rf te-aid/*/; sbatch ${NAME}_TEcurate.sh; done

```

Also need to set up my curation scripts for the new genome assemblies
```
cd /lustre/scratch/daray/protistan_holozoa/
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"

cd /lustre/scratch/daray/protistan_holozoa/
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done

```

Those will actually be run later, after all the preparation steps are complete.

Ran into trouble with permissions. Copied everything to a new folder "protists". Make appropriate changes to paths for downstream work.


```
cd /lustre/scratch/daray/protists/te_curations
LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"

cd /lustre/scratch/daray/protists/
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done

cd /lustre/scratch/daray/protists/te_curations
for NAME in $LIST; do rm -rf ${NAME}/te-aid/*/; done

for NAME in $LIST; do sbatch ${NAME}_TEcurate.sh; done

```

Also need to set up my curation scripts for the new genome assemblies
```
cd /lustre/scratch/daray/protists/
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"

for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rmodel_template.sh >te_curations/${NAME}_rmodel.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/RM_template_mod.sh >te_curations/${NAME}_RM.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/extend_align_template.sh >te_curations/${NAME}_extend_align.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/repeatclassifier_template.sh >te_curations/${NAME}_repeatclassifier.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/rm2bed_template_mod.sh >te_curations/${NAME}_rm2bed.sh; done
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" curation_templates/TEcurate_template.sh >te_curations/${NAME}_TEcurate.sh; done

```


RepeatModeler was already run before copying all the files to the new directory. Start with extend align.

```
cd /lustre/scratch/daray/protists/te_curations
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"

for NAME in $LIST; do sbatch ${NAME}_extend_align.sh; done
```
12251717    nocona nLov_NL_    daray PD       0:00      1 (Priority)
12251716    nocona nLov_Lov    daray PD       0:00      1 (Priority)
12251715    nocona nLov_Lov    daray PD       0:00      1 (Priority)
12251714    nocona nLov_F9_    daray PD       0:00      1 (Priority)
12251713    nocona nLov_ATC    daray PD       0:00      1 (Priority)
12251712    nocona nFow_ATC    daray PD       0:00      1 (Priority)
12251711    nocona nGru_NEG    daray PD       0:00      1 (Priority)
12251710    nocona nFow_Ty_    daray PD       0:00      1 (Priority)
12251709    nocona nFow_PA3    daray PD       0:00      1 (Priority)
12251708    nocona nFow_NF_    daray PD       0:00      1 (Priority)
12251707    nocona nFow_NF1    daray PD       0:00      1 (Priority)
12251706    nocona nFow_kur    daray PD       0:00      1 (Priority)
12251705    nocona nFow_ATC    daray PD       0:00      1 (Priority)
12251704    nocona nFow_AR1    daray PD       0:00      1 (Priority)
12251703    nocona aKon_ext    daray PD       0:00      1 (Priority)

```
sbatch --dependency=afterok:12251703 aKon_repeatclassifier.sh
sbatch --dependency=afterok:12251704 nFow_AR12_repeatclassifier.sh
sbatch --dependency=afterok:12251705 nFow_ATCC30894_repeatclassifier.sh
sbatch --dependency=afterok:12251706 nFow_kurume_repeatclassifier.sh
sbatch --dependency=afterok:12251707 nFow_NF1_repeatclassifier.sh
sbatch --dependency=afterok:12251708 nFow_NF_KFSHRC_repeatclassifier.sh
sbatch --dependency=afterok:12251709 nFow_PA34_repeatclassifier.sh
sbatch --dependency=afterok:12251710 nFow_Ty_repeatclassifier.sh
sbatch --dependency=afterok:12251711 nGru_NEGM_repeatclassifier.sh
sbatch --dependency=afterok:12251712 nFow_ATCC30863_repeatclassifier.sh
sbatch --dependency=afterok:12251713 nLov_ATCC30569_repeatclassifier.sh
sbatch --dependency=afterok:12251714 nLov_F9_repeatclassifier.sh
sbatch --dependency=afterok:12251715 nLov_Lova6_repeatclassifier.sh
sbatch --dependency=afterok:12251716 nLov_Lova7_repeatclassifier.sh
sbatch --dependency=afterok:12251717 nLov_NL_76_15_250_repeatclassifier.sh

```
12251746    nocona nLov_NL_    daray PD       0:00      1 (Dependency)
12251745    nocona nLov_Lov    daray PD       0:00      1 (Dependency)
12251744    nocona nLov_Lov    daray PD       0:00      1 (Dependency)
12251743    nocona nLov_F9_    daray PD       0:00      1 (Dependency)
12251742    nocona nLov_ATC    daray PD       0:00      1 (Dependency)
12251741    nocona nFow_ATC    daray PD       0:00      1 (Dependency)
12251740    nocona nGru_NEG    daray PD       0:00      1 (Dependency)
12251739    nocona nFow_Ty_    daray PD       0:00      1 (Dependency)
12251738    nocona nFow_PA3    daray PD       0:00      1 (Dependency)
12251737    nocona nFow_NF_    daray PD       0:00      1 (Dependency)
12251736    nocona nFow_NF1    daray PD       0:00      1 (Dependency)
12251735    nocona nFow_kur    daray PD       0:00      1 (Dependency)
12251734    nocona nFow_ATC    daray PD       0:00      1 (Dependency)
12251733    nocona nFow_AR1    daray PD       0:00      1 (Dependency)
12251732    nocona aKon_cla    daray PD       0:00      1 (Dependency)


LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"

```
sbatch --dependency=afterok:12251732 aKon_TEcurate.sh
sbatch --dependency=afterok:12251733 nFow_AR12_TEcurate.sh
sbatch --dependency=afterok:12251734 nFow_ATCC30894_TEcurate.sh
sbatch --dependency=afterok:12251735 nFow_kurume_TEcurate.sh
sbatch --dependency=afterok:12251736 nFow_NF1_TEcurate.sh
sbatch --dependency=afterok:12251737 nFow_NF_KFSHRC_TEcurate.sh
sbatch --dependency=afterok:12251738 nFow_PA34_TEcurate.sh
sbatch --dependency=afterok:12251739 nFow_Ty_TEcurate.sh
sbatch --dependency=afterok:12251740 nGru_NEGM_TEcurate.sh
sbatch --dependency=afterok:12251741 nFow_ATCC30863_TEcurate.sh
sbatch --dependency=afterok:12251742 nLov_ATCC30569_TEcurate.sh
sbatch --dependency=afterok:12251743 nLov_F9_TEcurate.sh
sbatch --dependency=afterok:12251744 nLov_Lova6_TEcurate.sh
sbatch --dependency=afterok:12251745 nLov_Lova7_TEcurate.sh
sbatch --dependency=afterok:12251746 nLov_NL_76_15_250_TEcurate.sh
```

Not a lot of stuff is being recognized. Suggested to Angela that she start more targeted searches. Will begin with an LTR search using MegaLTR.

https://bioinformatics.um6p.ma/MegaLTR/documentation

megaltr_template.sh
```
#!/bin/bash 
#SBATCH --job-name=<NAME>_mLTR
#SBATCH --output=%x.%j.out
#SBATCH --error=%x.%j.err
#SBATCH --partition=nocona
#SBATCH --nodes=1
#SBATCH --ntasks=12

. ~/conda/etc/profile.d/conda.sh
conda activate MegaLTR

TAXON=<NAME>
WORKDIR=/lustre/scratch/daray/protists/te_curations/$TAXON
mkdir -p $WORKDIR/megaltr
cd $WORKDIR/megaltr
SOFTWARE=/lustre/work/daray/software

#Run megaltr
bash $SOFTWARE/MegaLTR/MegaLTR.sh -A 1 -t 12 -P ${TAXON}_results -F $WORKDIR/assemblies_dir/${TAXON}.fa


```

```
cp megaltr_template.sh ../curation_templates/
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" megaltr_template.sh >${NAME}_megaltr.sh; done
for NAME in $LIST; do sbatch ${NAME}_megaltr.sh; done

LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" megaltr_template.sh >${NAME}_megaltr.sh; done
for NAME in $LIST; do sbatch ${NAME}_megaltr.sh; done
```


Forgot that this software requires a tRNA database. I can't find anything for protists so I'm going to make my own using tRNAscan-SE.

trnascan_template.sh
```
#!/bin/bash 
#SBATCH --job-name=<NAME>_tRNA
#SBATCH --output=%x.%j.out
#SBATCH --error=%x.%j.err
#SBATCH --partition=nocona
#SBATCH --nodes=1
#SBATCH --ntasks=1

. ~/conda/etc/profile.d/conda.sh
conda activate trnascan

TAXON=<NAME>
WORKDIR=/lustre/scratch/daray/protists/te_curations/$TAXON
mkdir -p $WORKDIR/trnascan
cd $WORKDIR/trnascan
SOFTWARE=/lustre/work/daray/software

#Run trnascan
tRNAscan-SE $WORKDIR/assemblies_dir/${TAXON}.fa
```
```
LIST="aKon nFow_AR12 nFow_ATCC30894 nFow_kurume nFow_NF1 nFow_NF_KFSHRC nFow_PA34 nFow_Ty nGru_NEGM nFow_ATCC30863 nLov_ATCC30569 nLov_F9 nLov_Lova6 nLov_Lova7 nLov_NL_76_15_250"
for NAME in $LIST; do sed "s/<NAME>/$NAME/g" trnascan_template.sh >${NAME}_trnascan.sh; done
for NAME in $LIST; do sbatch ${NAME}_trnascan.sh; done

64363  LIST="aWhi cOwc cPer cLim cFra hCla iHof mVib mBre nSp nDam pChi pVie pGem pUst sRos sArc"
64364  for NAME in $LIST; do sed "s/<NAME>/$NAME/g" trnascan_template.sh >${NAME}_trnascan.sh; done
for NAME in $LIST; do sbatch ${NAME}_trnascan.sh; done

```

Forgot LOTS of things about MegaLTR. It needs:
1. to be modified to take a custom directory to the tRNA database generated by trnascan-se
2. to be directed to rexdb-metazoa rather than to a plant database by default.

To solve these problems I modified the MegaLTR script in the github cloned directory. New version is below. Specific changes are on lines 
1 - commented out original
56 - Added documentation line
91 - Added option "I"
99 - Added option "I"

New script is:
```
#!/bin/bash
##Last update 31-01-2023 by Morad Mokhtar
   if [ $# -eq 0 ]
      then
         echo "Parameters -A and -F is required, use [bash MegaLTR.sh -help] for more detalis"
         exit
   fi
#tRNAdb= DRay changed this to an option. See options in text below.
LTR_FINDER=/lustre/work/daray/software/MegaLTR/bin/LTR_FINDER_parallel/LTR_FINDER_parallel
LTR_HARVEST=/lustre/work/daray/software/MegaLTR/bin/LTR_HARVEST_parallel/LTR_HARVEST_parallel
LTRretriever=/lustre/work/daray/software/MegaLTR/bin/LTR_retriever/LTR_retriever
chmod 775 $LTRretriever #Give execulting permissions for LTRretriever
RUN=/lustre/work/daray/software/MegaLTR/bin/RUN
chmod 775 $RUN/usearch11.0.667_i86linux32 #Give execulting permissions
eval "$(conda shell.bash hook)"
############################################################
# Set variables                                            #
############################################################
Analysistype=3
Fastafilepath=""
gffpath=""
trna=Arabidopsis_thaliana_trna.fa
outfileprefix=results
minlenltr=100
maxlenltr=7000
mindisltr=1000
maxdisltr=15000
similar=85
matchpairs=20
TEsorterdb=rexdb
TEsortercov=20
TEsortereval=0.001
TEsorterrule=80-80-80
TEsorterhmm=rexdb
RateOfEvolution=0.000000015
up=5000
down=5000
density1=1000000
numberofchromosom=9
threads=4
############################################################
# Help                                                     #
############################################################
Help()
{
   # Display Help
   echo "MegaLTR is a pipeline designed for high-throughput identification and classification of LTR-retrotransposons, insertion age estimation, detection of LTR-gene chimeras, and determination of nearby genes."
   echo
   echo "     MegaLTR v2.0."
   echo
   echo "     Options:"
   echo "     -h     Print this Help."
   echo "     -A     The analysis type [1 or 2 or 3] 1 (for Intact LTR-RT identification and annotation of internal domains 'This analysis needs FASTA file only') 2 (for Intact LTR-RT Identification and annotation of internal domains plus determination of insertion time 'This analysis needs FASTA file only') 3 (for Intact LTR-RT Identification, annotation of internal domains, determination of insertion time, LTR-RT gene-chimera analysis and visualization of gene density and LTR-RTs across chromosomes 'This analysis needs FASTA and GFF files') , default is 3"
   echo "     -F     Your path to Fasta file."
   echo "     -G     Your path to GFF file."
   echo "     -I     Path to fasta file with tRNA sequences."
   echo "     -T     tRNA sequence file (Locate the filename from the tRNA folder or provide your own tRNA sequence in FASTA format, default is Arabidopsis_thaliana_trna.fa)."
   echo "     -P     Outfileprefix, default is results."
   echo "     -l     Min length of 5'&3'LTR, default is 100."
   echo "     -L     Max length of 5'&3'LTR, default is 7000."
   echo "     -d     Min distance between 5'&3'LTR, default is 1000."
   echo "     -D     Max distance between 5'&3'LTR, default is 15000."
   echo "     -S     Specify similaritythreshold, default is 85."
   echo "     -M     Min length of exact match pair, default is 20."
   echo "     -B     TE Database that TEsorter will use it {gydb,rexdb,rexdb-plant,rexdb-metazoa,sine}, default is rexdb."
   echo "     -C     mininum coverage for protein domains in HMMScan output, default is 20."
   echo "     -V     maxinum E-value for protein domains in HMMScan output, default is 0.001."
   echo "     -Q     classifying rule [identity-coverage-length] based on similarity, default is 80-80-80]."
   echo "     -E     hmm-database that TEsorter will use it {gydb,rexdb,rexdb-plant,rexdb-metazoa,sine}, default is rexdb]."
   echo "     -R     neutral mutation rate of the target species (per bp per ya), e.g., rice: 1.3e-8 [0.000000013] (Ma and Bennetzen 2004); mammal: 2.2e-9 [0.0000000022] (S. Kumar 2002); Drosophila: 1.6e-8 [0.000000016](Bowen and McDonald 2001), default is 0.000000013."
   echo "     -U     the distance upstream LTR retrotransposons, default is 5000."
   echo "     -X     the distance downstream LTR retrotransposons, default is 5000."
   echo "     -W     window size to extract gene density from the GFF file, default is 1000000."
   echo "     -N     Number of chromosomes specified in FASTA file to visualize density of genes and LTRs, default is 12."
   echo "     -v     Print MegaLTR version and exit."
   echo "     -t     Indicate how many CPU/threads you want to run MegaLTR, default is 4."
   echo
   echo "Default parameters:bash MegaLTR.sh -A $Analysistype -F Your_path_to_FASTA_file -G Your_path_to_GFF_file -T $trna -P $outfileprefix -l $minlenltr -L $maxlenltr -d $mindisltr -D $maxdisltr -S $similar -M $matchpairs -B $TEsorterdb -C $TEsortercov -V $TEsortereval -Q $TEsorterrule -E $TEsorterhmm -R $RateOfEvolution -U $up -X $down -W $density1 -N $numberofchromosom -t $threads"
   exit
}
version()
{
   echo "MegaLTR v1.06."
   echo
   exit
}
############################################################
# Process the input options.                               #
############################################################
check=0
while getopts h:A:F:G:T:P:l:L:d:D:S:M:B:C:V:Q:E:R:U:X:W:N:v:t:I: options
do
   case $options in
      h) Help;;
      v) version;;
      A) Analysistype=$OPTARG;((check=check+1));;
      F) Fastafilepath=$OPTARG;((check=check+1));;
      G) gffpath=$OPTARG;;
      I) tRNAdb=$OPTARG;;
	  T) trna=$OPTARG;;
      P) outfileprefix=$OPTARG;;
      l) minlenltr=$OPTARG;;
      L) maxlenltr=$OPTARG;;
      d) mindisltr=$OPTARG;;
      D) maxdisltr=$OPTARG;;
      S) similar=$OPTARG;;
      M) matchpairs=$OPTARG;;
      B) TEsorterdb=$OPTARG;;
      C) TEsortercov=$OPTARG;;
      V) TEsortereval=$OPTARG;;
      Q) TEsorterrule=$OPTARG;;
      E) TEsorterhmm=$OPTARG;;
      R) RateOfEvolution=$OPTARG;;
      U) up=$OPTARG;;
      X) down=$OPTARG;;
      W) density1=$OPTARG;;
      N) numberofchromosom=$OPTARG;;
      t) threads=$OPTARG;;
     *) echo "Error: Invalid option" ;;
   esac
done

########################################################
division=100
similarFinder=$(perl -e "print $similar/$division") ###convert the similarity to LTR_FINDER format
process_id=$outfileprefix
userpath= mkdir -p $(pwd)/$process_id
userpath=$(pwd)/$process_id
LAI= mkdir -p $userpath/LAI
LAI=$userpath/LAI
ltrdigest= mkdir -p $userpath/LTRdigest
ltrdigest=$userpath/LTRdigest
mkdir -p $userpath/TEsorter
TEsorter=$userpath/TEsorter
mkdir -p $userpath/Others
Others=$userpath/Others
mkdir -p $userpath/Time
time=$userpath/Time
mkdir -p $userpath/R_plots
Rplots=$userpath/R_plots
mkdir -p $userpath/inside_genes
inside_genes=$userpath/inside_genes
mkdir -p $userpath/near_genes
near_genes=$userpath/near_genes
mkdir -p $userpath/density
densitypath=$userpath/density
process_id=$outfileprefix
mkdir -p $userpath/Collected_Files
Collected_Files=$userpath/Collected_Files
mkdir -p $userpath/FASTA
FASTA=$userpath/FASTA
mkdir -p $userpath/LTR_FINDER
FINDER=$userpath/LTR_FINDER
mkdir -p $userpath/LTR_HARVEST
LTRHARVEST=$userpath/LTR_HARVEST
mkdir -p $userpath/LTRFiles
LTRFiles=$userpath/LTRFiles
USERCH= mkdir -p $userpath/USERCH
USERCH=$userpath/USERCH

###############################################################
#### MegaLTR process the start.                               #
###############################################################
now="$(date)" 
echo
printf "\t#############################################\n\t##############  MegaLTR v2.0  ##############\n\t#############################################\n\n\tContributors: Morad M Mokhtar, Achraf El Allali\n\n"
printf "\t$now \t Start time %s\n"  ### print current date
echo
printf "\tParameters: -A $Analysistype -F $Fastafilepath -G $gffpath -T $trna -I $tRNAdb-P $outfileprefix -l $minlenltr -L $maxlenltr -d $mindisltr -D $maxdisltr -S $similar -M $matchpairs -B $TEsorterdb -C $TEsortercov -V $TEsortereval -Q $TEsorterrule -E $TEsorterhmm -R $RateOfEvolution -U $up -X $down -W $density1 -N $numberofchromosom -t $threads\n\n"
conda activate MegaLTR
   if [ $check -ne 2 ]
      then
         printf "\tParameters -A and -F is required, use [bash MegaLTR.sh -help] for more detalis\n"
         exit
      fi
      conda activate MegaLTR
   if [[ $Fastafilepath =~ \.gz$ ]]; then
         cd $userpath
         echo
         printf "\tCheck the FASTA File format.\n"
         gunzip -c "$Fastafilepath" >$userpath/$process_id.fna
         perl $RUN/checkfasta.pl $userpath/$process_id.fna ### Check the FASTA File format
      elif [[ $Fastafilepath =~ \.zip$ ]]; then
         echo
         printf "\tCheck the FASTA File format.\n"
         gunzip -c "$Fastafilepath" >$userpath/$process_id.fna
         perl $RUN/checkfasta.pl $userpath/$process_id.fna ### Check the FASTA File format
      elif ([ $(stat -c%s "$Fastafilepath") -gt 500 ]); then
         printf "\tCheck the FASTA File format.\n"
         perl $RUN/checkfasta.pl $Fastafilepath ### Check the FASTA File format
         cp $Fastafilepath $userpath/$process_id.fna #### copy fasta file tRNA files
      else
         printf "\n\tCheck the FASTA File format.\n"
         printf "\n\tPlease use -F to provide a valid FASTA file [-F FASTA_file_path].\n"
         exit
   fi

   if ([ $Analysistype -eq 3 ] && [[ $gffpath == "" ]]); then
         printf "\n\tPlease use -G to provide a valid GFF file [-G GFF_file_path].\n"
         exit
   fi
   if ([[ $Analysistype -eq 3 ]] && [ $(stat -c%s "$gffpath") -lt 500 ]); then
         printf "\n\tPlease use -G to provide a valid GFF file [-G GFF_file_path].\n"
      exit
   fi

   if ([ $Analysistype -eq 3 ] && [[ $gffpath =~ \.gz$ ]]); ### mode 3
      then
            gunzip -c "$gffpath" >$userpath/$process_id.gff
         elif ([ $Analysistype -eq 3 ] && [[ $gffpath =~ \.zip$ ]]); then
            gunzip -c "$gffpath" >$userpath/$process_id.gff
         elif ([[ $Analysistype -eq 3 ]] && [ $(stat -c%s "$gffpath") -gt 500 ]); then
            cp $gffpath $userpath/$process_id.gff #### copy gff file
      fi
   if ([ $Analysistype -eq 1 ] || [ $Analysistype -eq 2 ] || [ $Analysistype -eq 3 ]); ### mode 2
      then
         # : <<'END' END
         # printf "\t$now \t Start time %s\n"  ### print current date
         # echo
            ############## copy and split fasta #############
            cp $tRNAdb/$trna $userpath/$trna  #### copy tRNA file 
            sed -i 's/|/_/' $userpath/$process_id.fna  ### remove unnecessary details from Fasta sequence headers
            sed -i 's/|/ /' $userpath/$process_id.fna  ### remove unnecessary details from Fasta sequence headers
            sed -i 's/ .*//' $userpath/$process_id.fna  ### remove unnecessary details from Fasta sequence headers
            sed -i 's/\t.*//g' $userpath/$trna  ### remove unnecessary details from tRNA sequence headers
            now2="$(date)"
            printf "\n\t$now2 \tLTR_FINDER Started %s\n" 
            ################## LTR_Finder & LTR_HARVEST ########### updated 31-01-2023
         cd $FASTA
         perl $LTR_FINDER -seq $userpath/$process_id.fna  -threads $threads -harvest_out -size 1000000 -time 500  $tRNAdb/$trna $trna $maxdisltr $mindisltr $maxlenltr $minlenltr $matchpairs $similarFinder  $FASTA  $similar $process_id > /dev/null 2>/dev/null
         now51="$(date)"
         printf "\n\t$now51 \tLTR_HARVEST Started %s\n"

         conda config --show envs_dirs >$densitypath/condapath
         sed -i '1d' $densitypath/condapath
         sed -i 's/  - //g' $densitypath/condapath
            for condapath in `less $densitypath/condapath`
            do
               perl $LTR_HARVEST -seq $userpath/$process_id.fna -threads $threads -size 1000000 -time 500 -gt $condapath/MegaLTR/bin/gt $minlenltr $maxlenltr $similar > /dev/null 2>/dev/null
            done
         cat $FASTA/$process_id.fna.harvest.combine.scn  $FASTA/$process_id.fna.finder.combine.scn >$FASTA/$process_id.all.harvest.finder.combine

         now5="$(date)"
         echo
         printf "\t$now5 \tLTR_FINDER & LTR_HARVEST Done %s\n"   
         now50="$(date)"
         echo
         printf "\t$now50 \tLTR_retriever Started %s\n" 
         
            #######LTR_retriever ##########
         cd $LAI 
         $LTRretriever -genome $userpath/$process_id.fna -inharvest $FASTA/$process_id.all.harvest.finder.combine -threads $threads -minlen $minlenltr >>$userpath/screen.txt
         if [ -f "$LAI/$process_id.fna.pass.list" ]; then 
            cp $LAI/$process_id.fna.pass.list $Collected_Files
         fi
         if [ -f "$LAI/$process_id.fna.mod.pass.list" ]; then 
            cp $LAI/$process_id.fna.mod.pass.list $Collected_Files/$process_id.fna.pass.list
         fi 
         if [ -f "$LAI/$process_id.fna.nmtf.pass.list" ]; then 
            cp $LAI/$process_id.fna.nmtf.pass.list $Collected_Files
         fi
         if [ -f "$LAI/$process_id.fna.mod.nmtf.pass.list" ]; then 
            cp $LAI/$process_id.fna.mod.nmtf.pass.list $Collected_Files/$process_id.fna.nmtf.pass.list
         fi
         if [ -f "$LAI/$process_id.fna.pass.list.gff3" ]; then 
            cp $LAI/$process_id.fna.pass.list.gff3 $Collected_Files
         fi
         if [ -f "$LAI/$process_id.fna.mod.pass.list.gff3" ]; then 
            cp $LAI/$process_id.fna.mod.pass.list.gff3 $Collected_Files/$process_id.fna.pass.list.gff3
         fi
         if [ -f "$LAI/$process_id.fna.out" ]; then 
            cp $LAI/$process_id.fna.out $Collected_Files
         fi
         if [ -f "$LAI/$process_id.fna.mod.out" ]; then 
            cp $LAI/$process_id.fna.out $Collected_Files/$process_id.fna.out
         fi
         now7="$(date)"
         echo
         printf "\t$now7 \tLTR_retriever Done %s\n"
         #######ltrdigest ###########
         conda activate MegaLTR
         now8="$(date)"
         echo
         printf "\t$now8 \tLTRdigest Started %s\n"
         bash $RUN/ltrdigest.sh $process_id $userpath $trna $LAI $ltrdigest
         now9="$(date)"
         echo
         printf "\t$now9 \tLTRdigest Done %s\n"
         #######TEsorter ###########
         now10="$(date)"
         echo
         printf "\t$now10 \tTEsorter Started %s\n"
         cd $TEsorter
         bash $RUN/TEsorter.sh $TEsorter $ltrdigest/"$process_id"_complete.fas $TEsorterdb $TEsortercov $TEsortereval $TEsorterrule $TEsorterhmm $RUN $process_id $Collected_Files $userpath $threads
         now11="$(date)"
         echo
         printf "\t$now11 \tTEsorter Done %s\n"
         now12="$(date)"
         echo
         printf "\t$now12 \tFiltering TEsorter results Started %s\n"
         awk -F '\t' '$2 == "LTR"' $TEsorter/"$process_id"_complete.fas.$TEsorterhmm.cls.tsv >$Others/$process_id.LTR.tsv  ##### search inside a specific column
         awk -F '\t' '$2 != "LTR"' $TEsorter/"$process_id"_complete.fas.$TEsorterhmm.cls.tsv >$Others/$process_id.others.tsv  ##### search inside a specific column remove any other elements (like nested, LINE, ... any thing eles LTR)
         perl $RUN/print.pl $ltrdigest/"$process_id"_tabout.csv >$Others/$process_id.tabout.tsv ###### $Others/$"$process_id".tabout.tsv header ########## element	id element start	element end	element length	sequence	lLTR start	lLTR end	lLTR length	rLTR start	rLTR end	rLTR length	lTSD start	lTSD end	lTSD motif	rTSD start	rTSD end	rTSD motif	PPT start	PPT end	PPT motif	PPT strand	PPT offset	PBS start	PBS end	PBS strand	tRNA	tRNA motif	PBS offset	tRNA offset	PBS/tRNA edist
         now13="$(date)"
         echo
         printf "\t$now13 \tMergeing of LTR_retriever, LTRdigest, and TEsorter results %s\n"
         perl $RUN/TEsorter_Digest.pl  $Others/$process_id.tabout.tsv $Others/$process_id.LTR.tsv >$Others/2LTR_Table_TEsorter_Digest.tsv
         python3 $RUN/classification_NEW_LTR_2.py $Others/2LTR_Table_TEsorter_Digest.tsv  $Others/new_LTR_Table_TEsorter_Digest.tsv
         awk -F'\t' '{print $2"\t"$3"\t"$4"\t"$5"\t"$6"\t"$7"\t"$8"\t"$9"\t"$10"\t"$11"\t"$12"\t"$13"\t"$14"\t"$15"\t"$16"\t"$17"\t"$18"\t"$19"\t"$20"\t"$21"\t"$22"\t"$23"\t"$24"\t"$25"\t"$26"\t"$27"\t"$28"\t"$29"\t"$30"\t"$31"\t"$32"\t"$33"\t"$1"\t"$35"\t"$36"\t"$37"\t"$38}' $Others/new_LTR_Table_TEsorter_Digest.tsv > $Others/LTR_Table_TEsorter_Digest.tsv
         cp $Others/LTR_Table_TEsorter_Digest.tsv $Collected_Files ### LTR_retriever, LTRdigest, and TEsorter results in one file 
         awk -F "\t" '{ print  $2"\t"$33"\t"$3"\t"$4"\t"$5 }' $Others/LTR_Table_TEsorter_Digest.tsv > $Others/$process_id.length.ids.forstat
         python3 $RUN/super_familly_stat.py $Others/$process_id.length.ids.forstat $Collected_Files/$process_id.statistics.tsv ##LTR superfamily summary    
         awk -F "\t" '{ print  $2"\t"$3"\t"$4"\t"$5"\t"$32"\t"$33"\t"$34 }' $Others/LTR_Table_TEsorter_Digest.tsv > $Others/$process_id.ids.extract_seq
         cd $LTRFiles
         split -n l/100 $Others/$process_id.ids.extract_seq
         python3 $RUN/LTR_Seq_threads.py $userpath/$process_id.fna  $LTRFiles $Collected_Files $threads $RUN/extractseq-id-start-end.pl
         cp $ltrdigest/"$process_id"_pbs.fas $Collected_Files/$process_id.PBS.Sequence.fa
         cp $ltrdigest/"$process_id"_ppt.fas $Collected_Files/$process_id.PPT.Sequence.fa      
         sed  -i '1i LTR-RT id\tPseudomolecules/scaffolds\tLTR-RT start\tLTR-RT end\tLTR-RT length\tlLTR start\tlLTR end\tlLTR length\trLTR start\trLTR end\trLTR length\tlTSD start\tlTSD end\tlTSD sequence\trTSD start\trTSD end\trTSD sequence\tPPT start\tPPT end\tPPT motif\tStrand\tPPT offset\tPBS start\tPBS end\tStrand\ttRNA id\ttRNA motif\tPBS offset\ttRNA offset\tPBS/tRNA\t\tClass\tSuperfamily\tClade\tComplete\tStrand\tDomains' $Collected_Files/LTR_Table_TEsorter_Digest.tsv
		   ######## for LTR.non-redundant.fa #################

		   cp $Collected_Files/LTR-RT_Sequence.fa $USERCH/LTR-RT_Sequence.fa
		   $RUN/usearch11.0.667_i86linux32  -sortbylength $USERCH/LTR-RT_Sequence.fa --fastaout $USERCH/LTR-RT_Sequence_sorted.fa --log $USERCH/usearch.log
		   $RUN/usearch11.0.667_i86linux32  -cluster_fast  $USERCH/LTR-RT_Sequence_sorted.fa --id 0.9 --centroids $USERCH/LTR-RTs_non-redundant.fa --uc $USERCH/result.uc -consout $USERCH/LTR-RTs_conses.fa -msaout $USERCH/aligned.fasta --log $USERCH/usearch2.log
		   rm $USERCH/aligned.* 
		   cp $USERCH/LTR-RTs_non-redundant.fa $Collected_Files/LTR-RTs_non-redundant_library.fasta
		   now100="$(date)"
		   printf "\n\n\t$now101 \tNon-redundant LTR library done%s\n\n"
      
      fi
      if ([ $Analysistype -eq 2 ] || [ $Analysistype -eq 3 ]) ### mode 2
         then
            ########LTR insertion time ###########
            now14="$(date)"
            echo
            printf "\t$now14 \tCalculation of the LTR-RT insertion time %s\n"
            python3 $RUN/get_region.py $userpath/$process_id.fna $Others/LTR_Table_TEsorter_Digest.tsv $time
            echo "" > $Others/$process_id.time.txt
            echo "LTRNAME	K_Kimura	K_Ksd	K_TajimaNei	K_TNsd	timeK	timeKsd	numComparedSites	transitions	numComparedSites	transversions	numComparedSites	timeTN	transitions_numComparedSites	transversions_numComparedSites" >>$Others/$process_id.time.txt
               for fst in  $time/*.fasta 
               do
                  clustalw -infile="$fst" >>$Others/$process_id.clustalw.txt  ### Alignment of the LTR using clustalw
                  fname=$(basename $fst ".fasta")
                  ltrk=$(perl $RUN/estimate_K.pl $time/$fname".aln" $RateOfEvolution) ## estimate LTR-RT insertion time using Kimura and Tajima&Nei methods (this script is part of REannotate program)
                  echo -e $fname "\t""$ltrk"  >> $Others/$process_id.time.txt
               done
            perl $RUN/TEsorterandtable_time.pl  $Others/LTR_Table_TEsorter_Digest.tsv $Others/$process_id.time.txt >$Others/$process_id.Digest_TEsorter_Time.tsv  ############# $Others/$process_id.Digest_TEsorter_Time (header) org_name	acc	element_start	element_end	element_length			strand	lTSD_start	lTSD_end	lLTR_start	lLTR_end	rLTR_start	rLTR_end	rTSD_start	rTSD_end	EDTA_infoo	PPT start	PPT end	PPT motif	PPT offset	PBS start	PBS end	tRNA	tRNA motif	PBS offset	tRNA offset	PBS/tRNA	edist Order	Superfamily	Clade	Complete	Strand	Domains	LTRNAME	K_Kimura	K_Ksd	K_TajimaNei	K_TNsd	timeK	timeKsd	numComparedSites	transitions	numComparedSites	transversions	numComparedSites	timeTN	transitions_numComparedSites	transversions_numComparedSites
            cp $Others/$process_id.Digest_TEsorter_Time.tsv $Collected_Files ### LTR_retriever, LTRdigest, TEsorter, and insertion time results in one file
            sed  -i '1i LTR-RT id\tPseudomolecules/scaffolds\tLTR-RT start\tLTR-RT end\tLTR-RT length\tlLTR start\tlLTR end\tlLTR length\trLTR start\trLTR end\trLTR length\tlTSD start\tlTSD end\tlTSD sequence\trTSD start\trTSD end\trTSD sequence\tPPT start\tPPT end\tPPT motif\tStrand\tPPT offset\tPBS start\tPBS end\tStrand\ttRNA id\ttRNA motif\tPBS offset\ttRNA offset\tPBS/tRNA\t\tClass\tSuperfamily\tClade\tComplete\tStrand\tDomains\tK_Kimura\tK_Ksd\tK_TajimaNei\tK_TNsd\ttimeK\ttimeKsd\tnumComparedSites\ttransitions\tnumComparedSites\ttransversions\tnumComparedSites\ttimeTN\ttransitions_numComparedSites\ttransversions_numComparedSites' $Collected_Files/$process_id.Digest_TEsorter_Time.tsv
            ###### preparing the files for R plots #
            now15="$(date)"
            echo
            printf "\t$now15 \tPreparing R plots files %s\n"
            awk -F "\t" '{ print  $1"\t"$33"\t"$42 }' $Others/$process_id.Digest_TEsorter_Time.tsv > $Rplots/$process_id.time.ids
            awk -F "\t" '{ print  $2"\t"$33"\t"$3"\t"$4"\t"$5 }' $Others/$process_id.Digest_TEsorter_Time.tsv > $Rplots/$process_id.length.ids
            awk -F "\t" '{ print  $1"\t""Autonomous and Nonautonomous""\t"$3 }' $Rplots/$process_id.time.ids > $Rplots/$process_id.time.ids2
            awk -F "\t" '{ print  $1"\t""Autonomous and Nonautonomous""\t"$2"\t"$3"\t"$4"\t"$5 }' $Rplots/$process_id.length.ids >$Rplots/$process_id.length.ids2
            Rscript $RUN/chart_plot.r $Rplots/$process_id.length.ids $Rplots/$process_id.time.ids  MegaLTR..$process_id $RUN 2>/dev/null ### Both LTR-RT insertion time and LTR-RT length destributions plots based on the LTR-RT superfamilies
            Rscript $RUN/chart_plot.r $Rplots/$process_id.length.ids2 $Rplots/$process_id.time.ids2  MegaLTR..$process_id  $RUN 2>/dev/null ### Both LTR-RT insertion time and LTR-RT length destributions plots for the whole genome
            cp $Rplots/*.png $Collected_Files ## Both LTR-RT insertion time and LTR-RT length destributions plots (.png)
            now16="$(date)"
            echo
            printf "\t$now16 \tLTR-RT insertion time plots done %s\n"
            ##exit
         fi

      if [ $Analysistype -eq 3 ] ### mode 3      # #####  TE-gene chimeras #########
         then
            now17="$(date)"
            echo
            printf "\t$now17 \tLTR-RT-gene chimeras started %s\n"
            grep -P "\tgene\t" $userpath/$process_id.gff > $userpath/$process_id.grep.gene ## retrieve gene start and end from GFF file
            grep -P "\tpseudogene\t" $userpath/$process_id.gff > $userpath/$process_id.grep.pseudogene ## retrieve pseudogene start and end from GFF file
            cat $userpath/$process_id.grep.gene  $userpath/$process_id.grep.pseudogene > $userpath/$process_id.gene_pseudogene.gff  ## combine gene and pseudogene in one file
            perl $RUN/get-TE-within-gene.pl $Others/$process_id.Digest_TEsorter_Time.tsv  $userpath/$process_id.gene_pseudogene.gff $process_id $inside_genes > $inside_genes/$process_id.LTR_inside_genes.table  ## determine the LTR-RT located inside the gene start and end
            awk -F "\t" '{ print $1 }' $inside_genes/$process_id.LTR_inside_genes.table >$inside_genes/$process_id.LTR_inside_genes.table.ids
            awk -F "\t" '{ print $1 }' $Others/$process_id.Digest_TEsorter_Time.tsv >$inside_genes/$process_id.LTR_Table_Digest_TEsorter_Time.ids
            grep -F -x -v -f $inside_genes/$process_id.LTR_inside_genes.table.ids $inside_genes/$process_id.LTR_Table_Digest_TEsorter_Time.ids >$inside_genes/LTR_Table_Digest_TEsorter_Time_process   ###### print found in file 2 an not found in file 1 
            grep -f $inside_genes/LTR_Table_Digest_TEsorter_Time_process $Others/$process_id.Digest_TEsorter_Time.tsv > $inside_genes/LTR_Table_Digest_TEsorter_Time_nongene1
            perl $RUN/No.pl $inside_genes/LTR_Table_Digest_TEsorter_Time_nongene1 >$inside_genes/LTR_Table_Digest_TEsorter_Time_nongene  ## determine which LTR-RT located outside the gene start and end
            cat $inside_genes/$process_id.LTR_inside_genes.table $inside_genes/LTR_Table_Digest_TEsorter_Time_nongene >$inside_genes/LTR_Table_Digest_TEsorter_Time_nongene_and_gene.tsv ## combine the LTR-RT located insid and outside the gene start and end in one file
            cp $inside_genes/LTR_Table_Digest_TEsorter_Time_nongene_and_gene.tsv $Collected_Files
            sed  -i '1i LTR-RT id\tPseudomolecules/scaffolds\tLTR-RT start\tLTR-RT end\tLTR-RT length\tlLTR start\tlLTR end\tlLTR length\trLTR start\trLTR end\trLTR length\tlTSD start\tlTSD end\tlTSD sequence\trTSD start\trTSD end\trTSD sequence\tPPT start\tPPT end\tPPT motif\tStrand\tPPT offset\tPBS start\tPBS end\tStrand\ttRNA id\ttRNA motif\tPBS offset\ttRNA offset\tPBS/tRNA\tClass\tSuperfamily\tClade\tComplete\tStrand\tDomains\tK_Kimura\tK_Ksd\tK_TajimaNei\tK_TNsd\ttimeK\ttimeKsd\tnumComparedSites\ttransitions\tnumComparedSites\ttransversions\tnumComparedSites\ttimeTN\ttransitions_numComparedSites\ttransversions_numComparedSites\tinside gene status\tgene/pseudogene\tGene start\tGene end\tstrand\tgene annotation' $Collected_Files/LTR_Table_Digest_TEsorter_Time_nongene_and_gene.tsv
            now18="$(date)"
            echo
            printf "\t$now18 \tLTR-RT-gene chimeras done %s\n"
            #####*** TE near genes***
            now19="$(date)"
            echo
            printf "\t$now19 \tLTR-RT near genes started %s\n"         
            sed  's/\t?\t/\t+\t/g' $userpath/$process_id.gene_pseudogene.gff >$near_genes/$process_id.gene_pseudogene.gff2
            sed  's/\t?\t/\t+\t/g' $Others/$process_id.Digest_TEsorter_Time.tsv >$near_genes/$process_id.Digest_TEsorter_Time2
            awk -F '\t' '$36=="+"'  $near_genes/$process_id.Digest_TEsorter_Time2 >$near_genes/$process_id.Digest_TEsorter_Time5+
            awk -F '\t' '$36=="-"'  $near_genes/$process_id.Digest_TEsorter_Time2 >$near_genes/$process_id.Digest_TEsorter_Time5-
            perl $RUN/get-TE-near-gene-minuse1k.pl $near_genes/$process_id.Digest_TEsorter_Time5+ $near_genes/$process_id.gene_pseudogene.gff2 $up $down >$near_genes/up_genes+
            perl $RUN/get-TE-near-gene-pluse-1k.pl $near_genes/$process_id.Digest_TEsorter_Time5+ $near_genes/$process_id.gene_pseudogene.gff2 $up $down >$near_genes/down_genes+
            perl $RUN/get-TE-near-gene-minuse-1k.pl $near_genes/$process_id.Digest_TEsorter_Time5- $near_genes/$process_id.gene_pseudogene.gff2 $up $down >$near_genes/down_genes-
            perl $RUN/get-TE-near-gene-pluse+1k.pl $near_genes/$process_id.Digest_TEsorter_Time5- $near_genes/$process_id.gene_pseudogene.gff2 $up $down >$near_genes/up_genes-
            cat $near_genes/up_genes+ $near_genes/down_genes+ $near_genes/down_genes- $near_genes/up_genes- >$near_genes/genes_up_and_down_LTR.tsv
            cp $near_genes/genes_up_and_down_LTR.tsv  $Collected_Files/$process_id.genes_up_and_down_LTR.tsv
            sed  -i '1i LTR-RT id\tUp/Downstream\tPseudomolecules/scaffolds\tLTR-RT start\tLTR-RT end\tLTR-RT length\tlLTR start\tlLTR end\tlLTR length\trLTR start\trLTR end\trLTR length\tlTSD start\tlTSD end\tlTSD sequence\trTSD start\trTSD end\trTSD sequence\tPPT start\tPPT end\tPPT motif\tStrand\tPPT offset\tPBS start\tPBS end\tStrand\ttRNA id\ttRNA motif\tPBS offset\ttRNA offset\tPBS/tRNA\t\tClass\tSuperfamily\tClade\tComplete\tStrand\tDomains\tK_Kimura\tK_Ksd\tK_TajimaNei\tK_TNsd\ttimeK\ttimeKsd\tnumComparedSites\ttransitions\tnumComparedSites\ttransversions\tnumComparedSites\ttimeTN\ttransitions_numComparedSites\ttransversions_numComparedSites\tgene/pseudogene\tGene start\tGene end\tgene length\tstrand\tgene annotation' $Collected_Files/$process_id.genes_up_and_down_LTR.tsv
            now20="$(date)"
            echo
            printf "\t$now20 \tLTR-RT near genes done %s\n"
            ################### For chromosome LTR-RT distribution ###########
            now20="$(date)"
            echo
            printf "\t$now20 \tVisualization of gene density and LTR-RTs across chromosomes %s\n"
            awk -F "\t" '{ print  $33"\t"$33"\t"$2"\t"$3"\t"$4"\t"$33 }' $Others/$process_id.Digest_TEsorter_Time.tsv > $densitypath/$process_id.figure.distrbution1
            sed -i -E 's/Autonomous://g' $densitypath/$process_id.figure.distrbution1
            sed -i -E 's/Nonautonomous://g' $densitypath/$process_id.figure.distrbution1
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("Copia", "triangle", $2); print }' $densitypath/$process_id.figure.distrbution1 > $densitypath/$process_id.figure.distrbution2
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("Copia", "6a3d9a", $6); print }' $densitypath/$process_id.figure.distrbution2 > $densitypath/$process_id.figure.distrbution3
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("Gypsy", "triangle", $2); print }' $densitypath/$process_id.figure.distrbution3 > $densitypath/$process_id.figure.distrbution4
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("Gypsy", "33a02c", $6); print }' $densitypath/$process_id.figure.distrbution4 > $densitypath/$process_id.figure.distrbution5
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("BARE-2", "triangle", $2); print }' $densitypath/$process_id.figure.distrbution5 > $densitypath/$process_id.figure.distrbution6
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("BARE-2", "ff7f00", $6); print }' $densitypath/$process_id.figure.distrbution6 > $densitypath/$process_id.figure.distrbution7
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("TR-GAG", "triangle", $2); print }' $densitypath/$process_id.figure.distrbution7 > $densitypath/$process_id.figure.distrbution8
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("TR-GAG", "ff0080", $6); print }' $densitypath/$process_id.figure.distrbution8 > $densitypath/$process_id.figure.distrbution9
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("Unknown", "triangle", $2); print }' $densitypath/$process_id.figure.distrbution9 > $densitypath/$process_id.figure.distrbution10
            awk -F '\t' 'BEGIN { OFS=FS } { gsub("Unknown", "9d8477", $6); print }' $densitypath/$process_id.figure.distrbution10 > $densitypath/$process_id.figure.distrbution11

            grep "##sequence-region" $userpath/$process_id.gff > $densitypath/"$process_id"_karyotype
            sed -i 's/##sequence-region //g' $densitypath/"$process_id"_karyotype
            sed -i 's/ /\t/g' $densitypath/"$process_id"_karyotype
            awk -F "\t" '{print $1"\t"$4"\t"$5"\t""1"}' $userpath/$process_id.gene_pseudogene.gff >$densitypath/gene_anno
            python3 $RUN/counter.py $densitypath/gene_anno $density1 $densitypath/gene_anno_counter
            ulimit -i >$densitypath/limts
            for limt in `less $densitypath/limts`
            do
               ulimit -s $limt
               cd $densitypath
               if [ $numberofchromosom -le 8 ] ### the numbers of chromosomes less than or equal 8 to justfay the image
                  then  
                     sort  -k3,3nr $densitypath/"$process_id"_karyotype >$densitypath/"$process_id"_karyotype_sort
                     echo "Chr	Start	End" >$densitypath/"$process_id"_karyotype_sort_head
                     head -8 $densitypath/"$process_id"_karyotype_sort >>$densitypath/"$process_id"_karyotype_sort_head
                     awk -F "\t" '{print $1}' $densitypath/"$process_id"_karyotype_sort_head >$densitypath/"$process_id"_karyotype_sort_head.ids
                     echo "Chr	Start	End	Value" >$densitypath/gene_anno_counter_ok
                     grep -f $densitypath/"$process_id"_karyotype_sort_head.ids $densitypath/gene_anno_counter >>$densitypath/gene_anno_counter_ok
                     echo "Type	Shape	Chr	Start	End	color" >$densitypath/figure_distrbution5_ok
                     grep -f $densitypath/"$process_id"_karyotype_sort_head.ids $densitypath/$process_id.figure.distrbution11 >>$densitypath/figure_distrbution5_ok
                     python3 $RUN/figure_legend.py $densitypath/figure_distrbution5_ok $densitypath/gene_anno_counter_ok $densitypath/"$process_id"_karyotype_sort_head  $Collected_Files/"Map of Gene density and LTR-RTs distribution Figure.tsv"
                     Rscript $RUN/density_width.r $densitypath/"$process_id"_karyotype_sort_head $densitypath/gene_anno_counter_ok $densitypath/figure_distrbution5_ok
                     cp $densitypath/chromosome.svg $Collected_Files/"Gene density and LTR-RTs distribution.svg"
                     cp $densitypath/chromosome.png $Collected_Files/"Gene density and LTR-RTs distribution.png"
               fi
               if [ $numberofchromosom -ge 8 ] ### the numbers of chromosomes more than or equal 8 to justfay the image
                  then
                     sort  -k3,3nr $densitypath/"$process_id"_karyotype >$densitypath/"$process_id"_karyotype_sort
                     echo "Chr	Start	End" >$densitypath/"$process_id"_karyotype_sort_head
                     head -20 $densitypath/"$process_id"_karyotype_sort >>$densitypath/"$process_id"_karyotype_sort_head
                     awk -F "\t" '{print $1}' $densitypath/"$process_id"_karyotype_sort_head >$densitypath/"$process_id"_karyotype_sort_head.ids
                     echo "Chr	Start	End	Value" >$densitypath/gene_anno_counter_ok
                     grep -f $densitypath/"$process_id"_karyotype_sort_head.ids $densitypath/gene_anno_counter >>$densitypath/gene_anno_counter_ok
                     echo "Type	Shape	Chr	Start	End	color" >$densitypath/figure_distrbution5_ok
                     grep -f $densitypath/"$process_id"_karyotype_sort_head.ids $densitypath/$process_id.figure.distrbution11 >>$densitypath/figure_distrbution5_ok
                     python3 $RUN/figure_legend.py $densitypath/figure_distrbution5_ok $densitypath/gene_anno_counter_ok $densitypath/"$process_id"_karyotype_sort_head  $Collected_Files/"Map of Gene density and LTR-RTs distribution Figure.tsv"
                     Rscript $RUN/density.r $densitypath/"$process_id"_karyotype_sort_head $densitypath/gene_anno_counter_ok $densitypath/figure_distrbution5_ok
                     cp $densitypath/chromosome.svg $Collected_Files/"Gene density and LTR-RTs distribution.svg"
                     cp $densitypath/chromosome.png $Collected_Files/"Gene density and LTR-RTs distribution.png"               
                  fi
            done
         fi
         rm $userpath/$process_id.fna
         rm -rf $FASTA
         rm $userpath/$process_id.gff
         now21="$(date)"
         echo
         printf "\t$now21 \tMegaLTR Done, The results saved in ($Collected_Files) %s\n"
```

Also modified the megaltr_template.sh script to accommodate these changes:

```
#!/bin/bash 
#SBATCH --job-name=<NAME>_mLTR
#SBATCH --output=%x.%j.out
#SBATCH --error=%x.%j.err
#SBATCH --partition=nocona
#SBATCH --nodes=1
#SBATCH --ntasks=12

. ~/conda/etc/profile.d/conda.sh
conda activate MegaLTR

TAXON=<NAME>
WORKDIR=/lustre/scratch/daray/protists/te_curations/$TAXON
mkdir -p $WORKDIR/megaltr
cd $WORKDIR/megaltr
SOFTWARE=/lustre/work/daray/software

#Run megaltr
bash $SOFTWARE/MegaLTR/MegaLTR.sh \
	-A 1 \
	-t 12 \
	-P ${TAXON}_results \
	-T ${TAXON}.tRNA.fa \
	-F $WORKDIR/assemblies_dir/${TAXON}.fa \
	-I $WORKDIR/trnascan \
	-B rexdb-metazoa \
	-E rexdb-metazoa 
```










###### tags: `Protist TEs`