Moderator: Rob
Rob ♦ 3.2k
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Posts by Rob
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Comment:
C: Understanding salmon codes
... How large are they? You could use a file-sharing website, or if it is easier, you should be able to upload the data to [this gdrive link](https://drive.google.com/drive/folders/17N35X4_-6yuR4JMfLpbcreDZhQow1jpM?usp=sharing). ...
written 7 days ago by
Rob ♦ 3.2k
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Comment:
C: Understanding salmon codes
... That's a very surprising difference if the annotations are really the same. Would you be able to share the reads from one of your samples? We'd be interested to take a look. ...
written 8 days ago by
Rob ♦ 3.2k
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Comment:
C: Understanding salmon codes
... Hi Morris,
Can I ask how you verified that these are "human"? I took the sequences you gave above, and searched against the gencode transcriptome using Bowtie2 as the aligner. Both of these reads came back as completely unmapped. I did get a hit with blast (https://www.ncbi.nlm.nih.gov/nucleot ...
written 8 days ago by
Rob ♦ 3.2k
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Comment:
C: Understanding salmon codes
... Hi Morris,
It looks like you have a big space between the `--` and the flag name that should follow immediately after. That is, your command line should be:
```
salmon quant -i gencode_v29_idx -l A -1 Treated_1_m1.fastq -2 Treated_1_m2.fastq -p 4 --validateMappings --writeUnmappedNames -o salmo ...
written 9 days ago by
Rob ♦ 3.2k
1
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Comment:
C: Understanding salmon codes
... Actually, salmon can accept gzipped FASTA files for reference index creation as well as gzipped FASTA/Q files for quantification. I believe that Morris was able to build the index, but is running into problems while quantifying. ...
written 9 days ago by
Rob ♦ 3.2k
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Answer:
C: Understanding salmon codes
... Hi Morris,
You should add the index *directory* as the parameter to `-i`, so, try:
```
(salmon) [@ws7910 RNAseq]$ salmon quant -i gencode_v29_idx -l A -1 Treated_1_m1.fastq -2 Treated_1_m2.fastq -p 4 --validateMappings -o salm_treated1
```
I've also removed a spurious `/` before `gencode_v29_idx ...
written 9 days ago by
Rob ♦ 3.2k
0
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Comment:
C: Salmon transcriptome generation
... Hi Morris,
The `-k` argument has a default value (31) that is used if `-k` is not provided. I you wish to use the default, you don't have to pass that option explicitly. If you want to use another value of k, then you can pass that to the index command. ...
written 11 days ago by
Rob ♦ 3.2k
3
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... Hi Morris,
The easiest way to start using salmon is to download the transcriptome file directly (e.g. from Gencode --- ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_29/gencode.v29.transcripts.fa.gz). Then, you can create a salmon index via:
```
salmon index -t gencode.v29.tran ...
written 11 days ago by
Rob ♦ 3.2k
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... Yes, it seems that the output was sorted separately, using `samtools`, after having been generated by STAR. STAR has this behavior, I believe, because the transcriptome output is designed specifically for use with tools that have similar requirements to salmon (i.e. all alignments for a read are to ...
written 5 weeks ago by
Rob ♦ 3.2k
5
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... The command line you show for running using the STAR alignments:
```
salmon quant -t /hg38_salmon_transcriptome.fa -l IU -p 16 -a some.transcriptome.sorted.bam -o ./
```
suggests that the bam file provided to salmon was (coordinate?) sorted; is this the case? In alignment mode, salmon, like e.g. ...
written 5 weeks ago by
Rob ♦ 3.2k
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