# Help with PanPhlAn tutorial **URL:** \ **Category:** PanPhlAn\ **Created:** [April 27, 2020, 7:34pm UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386 "2020-04-27T19:34:37Z")\ **Posts on this page:** 7\ **Page:** 1
**Author:** ![sapphi](https://avatars.discourse-cdn.com/v4/letter/s/9dc877/32.png) [@sapphi](https://forum.biobakery.org/u/sapphi)\ **Post date:** [April 27, 2020, 7:34pm UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/1 "2020-04-27T19:34:37Z")
Hi, I am following the PanPhlAn tutorial online and ran into an issue I can’t solve ([https://github.com/SegataLab/panphlan/wiki/Tutorial#4-build-bowtie2-indexes](https://github.com/SegataLab/panphlan/wiki/Tutorial#4-build-bowtie2-indexes)). I am at step 4 (Build Bowtie2 indexes) but getting an error. Error: Please provide input files, either (–fna & --ffn), (–gff & --fna), or --gff alone. I am confused because the tutorial doesn’t mention -gff files or --ffa files. Thanks ---
**Author:** ![leonard.dubois](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/leonard.dubois/32/750_2.png) [@leonard.dubois](https://forum.biobakery.org/u/leonard.dubois)\ **Post date:** [April 28, 2020, 2:37pm UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/2 "2020-04-28T14:37:58Z")
Hello, there are currently two scripts with similar names. `panphlan_pangenome_generation.py ` and `panphlan_new_pangenome_generation.py ` The first one comes from previous version where the user should provide fasta file and annotation files to generate the pangenome himself (using the USEARCH software). The second script assume that you already have everything and only need. For the next step (mapping with `panphlan_map.py`), one only need to generate the Bowtie2 indexes from fna files. Anyway, I confess this isn’t the best option anyway. We are currently working on a full pangenome database, providing fasta, annotation and Bowtie2 indexes, that should be available soon. ---
**Author:** ![sapphi](https://avatars.discourse-cdn.com/v4/letter/s/9dc877/32.png) [@sapphi](https://forum.biobakery.org/u/sapphi)\ **Post date:** [April 29, 2020, 9:35am UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/3 "2020-04-29T09:35:04Z")
Thank you. I am quite new to bioinformatics so was struggling a bit. I have the new version of panphlan so can I create my own pangenome with fasta files and annotation files? ---
**Author:** ![leonard.dubois](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/leonard.dubois/32/750_2.png) [@leonard.dubois](https://forum.biobakery.org/u/leonard.dubois)\ **Post date:** [April 30, 2020, 9:45am UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/4 "2020-04-30T09:45:58Z")
Hello, creating your own pangenome with fasta and annotation is actually part of the “old code” ( `panphlan_pangenome_generation.py`) from PanPhlAn 1.2. For that you’ll need the USEARCH software for clustering ([https://www.drive5.com/usearch/](https://www.drive5.com/usearch/)) ---
**Author:** ![sapphi](https://avatars.discourse-cdn.com/v4/letter/s/9dc877/32.png) [@sapphi](https://forum.biobakery.org/u/sapphi)\ **Post date:** [May 1, 2020, 9:46am UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/5 "2020-05-01T09:46:46Z")
Hi, Thank you. I have just realized that I was using the 1.2 version so I should be able to make my own pangenome. Also, the panphlan\_erectale16\_pangenome.csv export from ChocoPhlan looks different to the panphlan\_39491\_pangenome.csv in the tutorial. Is there a reason one shows the UniProtID and the other the locus tags instead? ---
**Author:** ![CK\_zhu](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/ck_zhu/32/151_2.png) [@CK\_zhu](https://forum.biobakery.org/u/CK_zhu)\ **Post date:** [May 2, 2020, 12:20pm UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/6 "2020-05-02T12:20:51Z")
Hi , I have been studying panphlan example I download 14 fna and put them into reference\_genomes/ ```auto GCA_000209935.fna GCA_000209955.fna GCA_001404855.fna GCA_001405295.fna GCA_001406375.fna GCA_001406835.fna GCA_003122495.fna GCA_003436035.fna GCA_003436785.fna GCA_003438175.fna GCA_003438365.fna GCA_003438715.fna GCA_003438925.fna GCA_003438965.fna ``` when I run the follwing code I get an error，would you please give `panphlan_39491_pangenome.csv` file? ```auto ./panphlan/panphlan_new_pangenome_generation.py \ -c 39491 --i_fna reference_genomes/ -o . --verbose (panphlan) [ckzhu@vm-login02 panphlan]$ ./panphlan/panphlan_new_pangenome_generation.py \ > -c 39491 --i_fna reference_genomes/ -o . --verbose Error: Pangenome file (reference_genomes/panphlan_39491_pangenome.csv) not found ``` I also try old code ```auto (panphlan) [ckzhu@vm-login02 panphlan]$ ./panphlan/panphlan_pangenome_generation.py -c 39491 --i_fna reference_genomes/ -o . --verbose PanPhlAn pangenome generation version 1.2.3.7 Python version: 3.7.3 System: linux ./panphlan/panphlan_pangenome_generation.py -c 39491 --i_fna reference_genomes/ -o . --verbose [I] Input genome FNA folder: /public/home/sample_lib/ckzhu/software/panphlan/reference_genomes/ [I] Species database name: 39491 [I] Identity threshold percentage: 95.0 %. [I] Output folder: ./ [I] Temporary folder: TMP_panphlan_db/ STEP 1. Checking required software installations ... [I] Bowtie2 is installed, version: 2.4.1, path: /public/home/sample_lib/ckzhu/miniconda3/envs/panphlan/bin/bowtie2 [I] Usearch v.7 is installed, version: v7.0.1090_i86linux32, path: /public/home/sample_lib/ckzhu/software/panphlan/usearch7 STEP 2. Prepare input gene and genome files ... Traceback (most recent call last): File "./panphlan/panphlan_pangenome_generation.py", line 1341, in main() File "./panphlan/panphlan_pangenome_generation.py", line 1294, in main path_genome_fna_files, path_gene_ffn_files = check_genomes(args['i_ffn'], args['i_fna'], VERBOSE) File "./panphlan/panphlan_pangenome_generation.py", line 1089, in check_genomes genefiles = [f for f in os.listdir(ffn_folder) if fnmatch(f,'*.'+FFN)] NotADirectoryError: [Errno 20] Not a directory: False ``` Is there something else I need to download? ---
**Author:** ![leonard.dubois](https://yyz1.discourse-cdn.com/flex027/user_avatar/forum.biobakery.org/leonard.dubois/32/750_2.png) [@leonard.dubois](https://forum.biobakery.org/u/leonard.dubois)\ **Post date:** [May 11, 2020, 9:50am UTC](https://forum.biobakery.org/t/help-with-panphlan-tutorial/386/7 "2020-05-11T09:50:33Z")
Hello, I answered your question here : [https://github.com/SegataLab/panphlan/issues/3](https://github.com/SegataLab/panphlan/issues/3)