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mhammell-laboratory / TEtranscripts

A package for including transposable elements in differential enrichment analysis of sequencing datasets.
http://hammelllab.labsites.cshl.edu/software/#TEtranscripts
GNU General Public License v3.0
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TE transcript output #41

Closed SalimMegat closed 5 years ago

SalimMegat commented 5 years ago

Hi,

I have running TE transcript on my fastq but I only see gene ID in the output so I do not understand. Isn'it supposed to be TE families as well ? I am quite confused. ..Here are the output.

thanks

Salim.

fusWT_KI_gene_TE_analysis.txt fusWT_KI_sigdiff_gene_TE.txt

olivertam commented 5 years ago

Hi Salim,

Could you provide us an example of the command line, and perhaps attach the counts table (XXXX.cntTable)? Thanks.

SalimMegat commented 5 years ago

Figured it out all te count are stuck to 0, how is this possible ? fusWT_KI.cntTable.txt

olivertam commented 5 years ago

Hi Salim,

What genome build are you using (Ensembl GRCm38 or mm10)? Could you check the chromosome name in the BAM file to see if they start with "chr" or if they are just the number/letter of the chromosome? One possibility is that your mapping and gene annotations are using the Ensembl chromosome nomenclature, which does not have the "chr" in the chromosome name, while the TE annotation is in the UCSC format (mm10), which does have the "chr". If you need the GRCm38 TE annotation, it is available here. If you need an older mouse build, please let me know and we can try to make it.

If that does not address the issue, could you provide a small excerpt of your GTF and BAM files? You can send those to tam at cshl dot edu.

Thanks.

SalimMegat commented 5 years ago

Hi,

Many thanks for the advice I was indeed using the wrong gtf ! It worked !

Thank you,

Salim. On Jul 1, 2019, at 6:58 PM, Oliver Tam notifications@github.com wrote:

Hi Salim,

What genome build are you using (Ensembl GRCm38 or mm10)? Could you check the chromosome name in the BAM file to see if they start with "chr" or if they are just the number/letter of the chromosome? One possibility is that your mapping and gene annotations are using the Ensembl chromosome nomenclature, which does not have the "chr" in the chromosome name, while the TE annotation is in the UCSC format (mm10), which does have the "chr". If you need the GRCm38 TE annotation, it is available here. If you need an older mouse build, please let me know and we can try to make it.

If that does not address the issue, could you provide a small excerpt of your GTF and BAM files. You can send those to tam at cshl dot edu.

Thanks.

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub, or mute the thread.

SalimMegat commented 5 years ago

Hi,

Actually I have one more question regarding the output if you do not mind helping on this. Attached is the count table and if I understand correctly, there are gene ID and count for each gene right ? It is weird because the count that I get through TE transcripts are completely different to what I get from STAR geneCount output for instance. Are these count related to transposable elements in any given gene or the count for the whole gene ? I am just really confused and I would really appreciate some help.

Many thanks,

salim.

Le 1 juil. 2019 à 18:58, Oliver Tam notifications@github.com a écrit :

Hi Salim,

What genome build are you using (Ensembl GRCm38 or mm10)? Could you check the chromosome name in the BAM file to see if they start with "chr" or if they are just the number/letter of the chromosome? One possibility is that your mapping and gene annotations are using the Ensembl chromosome nomenclature, which does not have the "chr" in the chromosome name, while the TE annotation is in the UCSC format (mm10), which does have the "chr". If you need the GRCm38 TE annotation, it is available here http://labshare.cshl.edu/shares/mhammelllab/www-data/TEToolkit/TE_GTF/GRCm38_rmsk_TE.gtf.gz. If you need an older mouse build, please let me know and we can try to make it.

If that does not address the issue, could you provide a small excerpt of your GTF and BAM files. You can send those to tam at cshl dot edu.

Thanks.

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub https://github.com/mhammell-laboratory/tetoolkit/issues/41?email_source=notifications&email_token=AKO24HQ65IB2R2ZSDL2I7GDP5IZSZA5CNFSM4H4UDCYKYY3PNVWWK3TUL52HS4DFVREXG43VMVBW63LNMVXHJKTDN5WW2ZLOORPWSZGODY6XWNY#issuecomment-507345719, or mute the thread https://github.com/notifications/unsubscribe-auth/AKO24HUL4JU2TIVCMVHT6U3P5IZSZANCNFSM4H4UDCYA.

gene/TE fuski_2.bamAligned.sortedByCoord.out.bam.T fuski_5.bamAligned.sortedByCoord.out.bam.T fuski_3.bamAligned.sortedByCoord.out.bam.T fuski_1.bamAligned.sortedByCoord.out.bam.T fusWT_3.bamAligned.sortedByCoord.out.bam.C fusWT_4.bamAligned.sortedByCoord.out.bam.C fusWT_2.bamAligned.sortedByCoord.out.bam.C fusWT_5.bamAligned.sortedByCoord.out.bam.C "ENSMUSG00000000001.4" 1 1 0 0 2 4 1 0 "ENSMUSG00000000003.15" 0 0 0 0 0 0 0 0 "ENSMUSG00000000028.15" 0 2 0 0 0 0 0 0 "ENSMUSG00000000031.16" 0 0 0 0 0 0 0 0 "ENSMUSG00000000037.16" 0 0 0 0 0 0 0 0 "ENSMUSG00000000049.11" 0 0 0 0 0 0 0 0 "ENSMUSG00000000056.7" 1 3 3 5 2 3 2 3 "ENSMUSG00000000058.6" 1 1 2 1 1 0 0 1 "ENSMUSG00000000078.7" 1 1 5 0 9 7 1 5 "ENSMUSG00000000085.16" 0 0 1 1 4 1 1 0 "ENSMUSG00000000088.7" 3 3 1 1 6 3 2 3 "ENSMUSG00000000093.6" 0 0 0 0 0 0 0 0 "ENSMUSG00000000094.12" 0 0 0 0 0 0 0 0 "ENSMUSG00000000103.12" 0 0 0 0 0 0 0 0 "ENSMUSG00000000120.6" 0 0 0 0 0 0 0 0 "ENSMUSG00000000125.5" 0 1 0 1 0 0 0 0 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olivertam commented 5 years ago

Hi Salim,

The counts are for each gene, and aggregated to the gene ID. As for why there are differences with STAR quantMode geneCounts, there could be several reasons: 1) TEtranscripts disambiguate reads if they are annotated to multiple features, and so annotate more reads than geneCounts 2) TEtranscripts require the user to specify if the library is stranded, reverse-stranded, or unstranded. Each of these modes could give different counts. geneCounts does all three at once, but again, they cannot handle ambiguous annotations. 3) geneCounts will not handle multi-mapping reads correctly, and will count each alignment as a separate read. This could lead to overcounting (if a read maps to multiple regions that matches different gene).

Without knowing how different the read counts are (whether there is a global decrease/increase, or gene-specific variations), it is difficult to determine which of the scenarios is/are at play.

Thanks.

SalimMegat commented 5 years ago

Hi,

The 2nd option was actually the good one ! My library was reverse stranded so I added it too the command line and now it looks closer to what I got from STAR.

Thank you for your help !

Salim.

Le 2 juil. 2019 à 18:32, Oliver Tam notifications@github.com a écrit :

Hi Salim,

The counts are for each gene, and aggregated to the gene ID. As for why there are differences with STAR quantMode geneCounts, there could be several reasons:

TEtranscripts disambiguate reads if they are annotated to multiple features, and so annotate more reads than geneCounts TEtranscripts require the user to specify if the library is stranded, reverse-stranded, or unstranded. Each of these modes could give different counts. geneCounts does all three at once, but again, they cannot handle ambiguous annotations. geneCounts will not handle multi-mapping reads correctly, and will count each alignment as a separate read. This could lead to overcounting (if a read maps to multiple regions that matches different gene). Without knowing how different the read counts are (whether there is a global decrease/increase, or gene-specific variations), it is difficult to determine which of the scenarios is/are at play.

Thanks.

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