Need help with STAR aligner
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16 months ago

Hi guys, I hope your are fine, I am new to bioinformatics and I am trying to use STAR aligner I already installed it but I am feeling lost, I need help how to start if you have any advice or useful links it will be appreciated thanks in advance.

RNA-Seq • 1.0k views
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Please google for the STAR manual and then try to apply it. You are welcome to ask for help if you get stuck but please either try something first or post a specific question.

The extensive manual: https://github.com/alexdobin/STAR/blob/master/doc/STARmanual.pdf

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thanks for your help, I already read it but I feel that I don't understand it , do you know any way to use it on labtop? with a limited RAM size

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STAR is not intended to be used in a Laptop, maybe Salmon or Kallisto is better for this

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what organism do you work with? for human size genome you would need 32GB of RAM, for organisms with smaller genomes (like yeasts with 10-12 Mb), STAR alignment is doable with a modest laptop.

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you need at least 25 or 30 GB of RAM for STAR if I'm not mistaken. Do you maybe have access to an HPC? You could check out the online platform galaxy, which holds a set of bioinformatic tools for usage in a cloud. STAR is called RNA STAR in there.

https://usegalaxy.org/

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thank you soooooo much I'll give it a try

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It will be tricky on a laptop. How many samples do you have and what organism are you working with? If you give a bit of context what you are analyzing there are most likely solutions not as hardware-demanding as STAR.

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16 months ago

Hi Zeinab, if your laptop's RAM lower than 32 GB , it will not work, actually you can start alignment but after same time the process going to stop and you will get a massage like 'Process Killed'. I have used STAR for Human data analysis and it's very powerful tool and super fast. As our colleges mentioned above we need more information regarding you analysis to help you ! generally speaking first step is generating Genome Index "assuming it's Human data download Reference data with it's annotation and i highly recommend you to use GeneCod data base 'https://www.gencodegenes.org"

STAR --runThreadN 8 --runMode genomeGenerate --genomeDir 'pathToYourGenome' --genomeFastaFiles 'PathToYourFastaFile' --sjdbGTFfiles 'PathToYourGTFfile' --sjdbOverhang 100

*genomeDir= is where to placed indexed genome *sjdbOverhang= specified the length of the genomics sequenced around annotated junctions for construction the splicing junction database it's correspond to the read length of you RNASeq data (value of 100 will usually 'Not allows' work with most cases) * this process will take around 30 Min. "for me" and it's dependent on machine specifications

second step Mapping: *For PE:

STAR --runThreadN 8 --runMode alignReads --genomeDir 'pathToYourIndexedGenome' --readFilesIn 'pathToYourReadR1&R2' --outSAMtype BAM SortedByCoordinate

*For advance option see STAR manual 'https://physiology.med.cornell.edu/faculty/skrabanek/lab/angsd/lecture_notes/STARmanual.pdf'

Now you are done with STAR and you can proceed with GATK for Alignment post processing and variant calling, i highly recommend you to follow GATK best practices 'https://gatk.broadinstitute.org/hc/en-us/articles/360035531192-RNAseq-short-variant-discovery-SNPs-Indels-'

If you have Limited RAM, then STAR not for you and try to find another tools like: GSNAP, TopHat2, MapSplice < i did not test any one of them myself "Not Sure about system requirements"

Best Regards Abdullah

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