blahah/transrate

Understand your transcriptome assembly

Stuart Willis

@stuartwillis

So I guess v1.0.4 is ready to rock? with updated dependencies? Any timeframe for updating the ruby gem?

Stuart Willis

@stuartwillis

ok, so deleted old local version of salmon (0.4.2) that the gem installed, copied updated version of salmon (0.8.2) there so it's in the path, and ran transrate (1.0.4) install-deps just for kicks. Got this: Checking dependencies
Installing salmon (0.4)...
[ERROR] 2017-07-20 17:13:29 : Failed to install:

salmon:0.4

Kate Wathen-Dunn

@katewd

@stuartwillis , I am having the same problem. Tried various salmon versions (0.4.2, 0.6.0 and 0.8.2),but I can't get TransRate to recognise them and use them, so currently I can only do the basic assembly metric analysis without the reads, etc. I would really love to get this to work, so any advice would be appreciated!

Lisa K. Johnson

@ljcohen

I had the same problem. A comment above about removing the /bin/librt.so.1 file and assistance from @bluegenes helped me out. this worked on Ubuntu 16.04:

cd 
sudo curl -SL https://bintray.com/artifact/download/blahah/generic/transrate-1.0.3-linux-x86_64.tar.gz | tar -xz
cd transrate-1.0.3-linux-x86_64 
./transrate --install-deps ref
rm -f bin/librt.so.1
echo 'export PATH=$PATH:"$HOME/transrate-1.0.3-linux-x86_64"'

, transcriptome eval tutorial here: http://dibsi-rnaseq.readthedocs.io/en/latest/evaluation.html

Kate Wathen-Dunn

@katewd

Thanks @ljcohen , you're a legend! I ran 'sudo gem uninstall transrate' , unpacked the transrate-1.0.3 tarball I already had downloaded then ran the './transrate --install-deps ref' and the 'rm librt.so.1' commands. I added the 'export PATH' line for both transrate-1.0.3 and salmon-0.8.2 to my ~/.profile and then TransRate ran perfectly. Thank you so much!

Lisa K. Johnson

@ljcohen

great, glad it worked!

Upendra Kumar Devisetty

@upendrak

Hi, I am running transrate on an Trinity assembly and eventhough the Transrate finished running and generated the outputs, I see the salmon.log file has several rows that shows this error
[2017-07-27 16:11:56.857] [jointLog] [info] Sampling alignments; outputting results to ./postSample.bam
[2017-07-27 16:11:57.184] [jointLog] [warning] [Sampler.hpp]: Failed to sample an alignment for this read; this shouldn't happen
currentMass = -nan, r = 0.293905
[2017-07-27 16:11:57.185] [jointLog] [warning] [Sampler.hpp]: Failed to sample an alignment for this read; this shouldn't happen
currentMass = -nan, r = 0.665585"
Consequently the Transrate score is quite low. Can anyone let me what this error means? Thanks..

JakeGoodall

@JakeGoodall

Hi All, I was wondering if someone could maybe weight in on an 'issue' I'm having. I've been using Translate a lot recently (great program), running lots of different assembly variants (some are TPM filter, some TransRate filtered etc, etc).

Now a consistent issue I've been having is the initial generation of the .bam file. It takes a long time (not suprising since its a massive transcriptome), but once the .bam completes (presumably) TransRate gets stuck, never moving past the .bam and/or onto the downstream analysis. Its not a massive issue, since cancelling the run, then restarting will progress the program to completion.

I wouldn’t think much off it but some of my results recently have been ‘interesting’. My best transcriptome’ initial score rated 0.8, whilst the good.assembly 1.5. Is it even possible to get TransRate scores >1? Certainly I’ve improved my ability to construct good transcriptomes as of late, but I’m more inclined to think somethings going wrong in the analysis. Should I be trusting these scores or is TransRate getting stuck affecting the outputs?

Anne Webb

@webbchen

Good morning. I've made a couple of assembly with a mixture of single- and paired end reads. The question of SE_reads came up earlier, e.g. here:

@preeti333 : yes, this is currently restricted to paired-end data. The information provided by single-end data is much less useful and reliable for assembly evaluation.

With transrate I've included the single end reads with the forward reads as I did for the assemblies. Will that cause transrate to fail? At the moment it's getting struck at the read diagnostic stage as described by Jake Goodall. Tried transrate 1.0.2 and 1.0.3 so far.

Single-ended reads are at the end of the forward-reads file so they should not interfere with the ordering of the paired-end files.

bubblegirl

@bubblegirl

@ljcohen & @katewd Thank you two! Same problem, fixed ^___^

FleurGaBru

@FleurGaBru

Hi all! I am new to transrate and have a question about the conda build. I installed transrate-tools by "conda install -c bioconda transrate-tools" but the only commando that I can execute (and can find in the conda recipes) is bam-read. I am actually looking for the commando to execute transrate to analyse my trinity assembly based on the sequences and reads. Is the conda build incomplete or do I miss something very crucial? Thanks a lot!

Matt MacManes

@macmanes

Sorry @FleurGaBru, transrate-tools is not the same as transrate.

you can grab the binary here: http://hibberdlab.com/transrate/installation.html

FleurGaBru

@FleurGaBru

Thanks!

Are you planning to make a conda build for transrate?

Matt MacManes

@macmanes

that is a question for @blahah

FleurGaBru

@FleurGaBru

I actually just scrolled up a bit and saw that @notestaff is working on that. Any update on the progress?

sextuplev

@sextuplev

Dear Transrate authors and users, I am using TransRate to evaluate RNA-seq de-novo assembly and surprisingly, I got a very low mapping rate (< 1%). To check the problem, I directly used snap-aligner to map the same dataset and got a high mapping rate (>90%). In another dataset, however, TransRate gave a high mapping rate as well as snap-aligner.
I am confused now and any suggestions or solution will be appreciated.

sextuplev

@sextuplev

Hi all, I noticed that a parameter '0.95' in "bam-read in.bam out.csv 0.95", and Transrate authors use 0.7 instead. Who knows it stands for?

ddurai

@ddurai

Dear transrate authors and users. I am trying to evaluate assemblies generated by transABySS and Oases. The datasets were normalized and then assembled. I checked the transrate score for assemblies generated from various reduction levels of the normalization algorithm. The number of contigs decreased with the decrease in the number of input reads. But surprisingly, the transrate assembly score increased with the decrease in number of contigs. I am a bit confused about it. Shouldnt the transrate decrease with the decrease in the number of contigs. Or am I understanding the algorithm incorrectly. I used the original unreduced dataset for evaluation using transrate

Balázs Hajgató

@hajgato

Dear All, I would like to know how important Salmon version for transrate 1.0.3. It seems that changing from the bundled 0.6.0 to 0.8.2 gives completely different results. Which version should be used then?

Abhijeet Shah

@abshah

Hi transrate devs! I have quick question. Is there a "quick and dirty" way of getting hold of possible chimeric contigs using s(Cseg) scores ? More specifically, I mean, can I just chuck out the contigs with "low" s(Cseg) scores ?

Abhijeet Shah

@abshah

@blahah @rob-p, bump!

Rik

@blahah

@abshah sorry I haven't been getting notifications from this channel for some reason

Yes you should be able to do that. It's not a perfect filter but it fits the "quick and dirty" requirement :)

Anton Kulaga

@antonkulaga

I have misterious "terminal_columns': undefined methodwinsize' for nil:NilClass " when I run transrate blahah/transrate#216

AsterChen

@AsterChen

Hi. I recently used transrate to check my transcriptome assemblies derived from Trinity and the assembly scores were incredibly low (around 0.02). I dont know whether I did something wrongly when I run the program or my assemblies are really bad. I once changed my read files manually from .fastqsanger to .fastq since transrate seems doesnt recognize .fastqsanger. Is that a problem?

svanscarfo

@svanscarfo

Hi,
I've installed transrate on my mac to assess the quality of different assemblies. Everything seem to work fine for the contig metrics (2sec for 38K contigs ranging from 200-15000bp). This effectively run either with example data or my own assemblies. However, when I tried to run the read mapping metric analysis (with example data or my assemblies), no (INFO) lines appear below my command line, Additionally Salmon PID seems to run in loop with really memory usage in TOP (99-100% USAGE / 532K memory) and the Global CPU usage is not more than 5% in the activiy monitor. I've tried to move the transrate folder in other locations with no success. Could you please help me on that.

Thanks.

Muskaann

@Muskaann

Hi All, I am getting a really low assembly score (0.13) for my seagrass transcriptome. I was wondering how many plant transcriptomes were assessed using TransRate and if we have information on scores from maybe the "model" plants for me to compare my scores to? I know that a score closest to 1 is the best but do we have something that is an ok score? Thanks.

Matt MacManes

@macmanes

hi @Muskaann, you can see the ORP protocol (https://www.biorxiv.org/content/early/2017/11/22/177253), and corresponding data (https://github.com/macmanes-lab/Oyster_River_Protocol/blob/master/manuscript/orp.csv) for several plant species. In the csv, the score columns show the scores for the different assemblers, and the opt columns are the optimal scores.

Muskaann

@Muskaann

@macmanes Thank you so much for the links. I will also try to use ORP protocol and see if it improves my assembly.

Matt MacManes

@macmanes

@Muskaann let me know if you have difficulties with the ORP.

PauloFlores

@PauloFlores

Hi blahah. I performed a mega RNA-Seq assembly (>800 samples, 16 billion of reads, mapped to my plant genome produced near 180,000 contigs) and now I'm trying to filter the results based on Transrate. I'm doing a few tests to understand the resources/running time required (mapping reads to the contigs and inspecting the alignments), and just testing with 10% of the reads apparently I'll need more than 1 week (32 threads) to get the results. "Calculating read diagnostics..." was the last message. Any advice how to increase the speed or how to run transrate with this amount of data?

Anton Kulaga

@antonkulaga

I hope you will manage to run it. I tried to run it many times on my data and failed

PauloFlores

@PauloFlores

Hi Anton. The size of the dataset maybe is not a problem, perhaps splitting the contigs in smaller datasets solves the situation. However, Salmon is not working (TPMs are zero) and I have no idea why...and finally - how to use the scores to filter the results??

Jochen Bick

@bimbam23

Hi, is there any limit to the size of the fastq files? I'm running two 70GB files versus a big assembly. Somehow it looks like the program got stuck. I can see any movements in the output folder. it looks like the snap-aligner ran into some trouble

Danielle DeLeo

@DeLeoDM_twitter

Hi- I am trying to assess the quality of a reference transcriptome assembly that was assembled with trinity using a sample.tsv file containing paths to replicate (n=5) read files (paired-end). I am hoping to get information about the assembly score with transrate, but am unsure of how to run the read metrics mode (--left --right) with multiple sample files. Is there a way to use this sample.tsv file or list multiple files?

Jochen Bick

@bimbam23

the error message is: [ERROR] 2018-03-12 16:47:36 : Snap failed
Welcome to SNAP version 1.0dev.96.

@DeLeoDM_twitter --left=<s> Left reads file(s) in FASTQ format, comma-separated
--right=<s> Right reads file(s) in FASTQ format, comma-separated

comma-separated should work

Grant Batzel

@grant_batzel_twitter

how long does transrate take to run? I have 90M reads in my forward file and 90 million reads in my reverse. I'm currently running transrate on a computer with 40 CPUs and 160G memory

TepoltC

@TepoltC

@svanscarfo, I think I'm having the same issue. On a Mac Pro, transrate's read mapping analysis doesn't appear to multithread even when I specify multiple threads in the command. Instead, it sits at the salmon stage, using a single thread and taking FOREVER (with no info printout). Did you figure out a solution? Does anyone else have any suggestions? Would love to use transrate but I think it's untenable on a single thread.

Grant Batzel

@grant_batzel_twitter

update: took me 90 minute to run transrate on my assembly

PauloFlores

@PauloFlores

--threads worked fine (linux) with me. Snap is slower than Salmon but 90 million reads shouldnt take very long - couple of hours?

Urwah Nawaz

@urwahnawaz_twitter

Hi all, I've generated a de novo assembly using Trinity and used TransRate to evaluate the accuracy. My transRate score turned out to be very low (0.03). I used other metrics as well (BUSCO, back mapping, ExN50), and so far they all look good. I wanted to know if TransRate would be good for my species (I'm sequencing a cephalopod and they are known to have a highly repetitive content). Any advice on this would be great :)

jereBrand

@Jeremias-Brand

Hi, all. I am running transrate on multiple assemblies simultaneously as part of a large snakemake pipeline. For this I limit threads for each run to a fraction of total cores used. Transrate seems to then only use the global limit. If I run 3 runs with 10 cpus assigned each then all 3 transrate runs will be executed at the same time but only using 10 cores. This is really slowing down the whole pipeline. Any suggestions on how to fix that?=

@blahah

Stuart Willis

@stuartwillis

@blahah is there a 'simple' reason why scores would be different between 1.0.1 (w/ salmon 0.4.2) and 1.0.3 (w/ salmon 0.6.0) for the same assembly? The fragments mapped etc. are much lower for 1.0.3 (e.g. 0.91 vs. 0.29). I tried to swap in the most recent salmon (0.9.?) to see what effect that had but transrate returned 'missing 0.6.0'. Any thoughts?

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