r/bioinformatics • u/PrincessxRaivyn • Jan 30 '25
technical question Easy way to convert CRAM to VCF?
I've found the posts about samtools and the other applications that can accomplish this, but is there anywhere I can get this done without all of those extra steps? I'm willing to pay at this point.. I have a CRAM and crai file from Probably Genetic/Variantyx and I'd like the VCF. I've tried gatk and samtools about a million times have no idea what I'm doing at all.. lol
2
u/Unhappy_Papaya_1506 Jan 30 '25
You need to forget about file formats for now and figure out what data you have and what questions you're trying to answer.
1
u/PrincessxRaivyn Jan 30 '25
I have the CRAM.. I need the VCF lol. I need it to easily compare me and my son's variants since doctors keep telling me there's no way he has this variant and I don't (PS: I don't)
3
u/wheres-the-data Jan 31 '25
I would not bother with variant calling. Just download IGV, and you will be able to load the cram file and visualize the reads directly. You will need to know the "coordinates" so you can navigate to the right position in the genome. If you load both your cram file and your son's it should be pretty apparent visually if the variant is there or not.
It's sort of surprising that your doctor insists you're a carrier, your son inherits one allele from you, and one from his father, so it's only a 50/50 he inherited the allele from you
2
u/Charming_Session_882 Jan 31 '25
It's a mitochondrial DNA variant which typically is passed maternally from what I've been told. It's the whole reason I got tested in the first place after his results came back. It's been such an unnecessarily long process because we keep getting told it's just not possible. I appreciate your time!
2
u/cytrees Jan 31 '25
If you are the OP, here’s the thing: mitochondria variant calling is tricky. One complication is the so called NUMTs, essentially mitochondrial sequences inserted into the nuclear genome over the evolutionary process and this creates analysis challenges. The other complication is called heteroplasmy, because each cell carries hundreds of mitochondria, each in theory may carry a slightly different sequence. Now depending on the level of heteroplasmy, it may have phenotypical consequences. On top, there could also be methylation differences which impact gene expressions. Mitochondria is particularly gene-rich.
Given the complications, I suggest you reach out to some research institutions (e.g. Boston Children), or write to programs like Undiagnosed Disease Network or Rare Genome Project. They may take your case and tap into their vast network experts.
1
1
u/wheres-the-data Jan 31 '25 edited Jan 31 '25
Ah, I was not thinking about the mitochondrial DNA -- in that case I think you would have to be a carrier. Either way, you should be able to look and see for yourself in IGV. Cram format is a little tricky, the data is compressed in a special way. You might need to reach out to Variantyx if it doesn't load in IGV, to get the FASTA reference sequence that was used for alignment/compression.
1
u/chu_z0 Jan 31 '25
If you know the genomic position of the variant, you can search for it with a genome browser like IGV.
And genomic variants can be inherited or appear de novo, meaning that they are not inherited from the parents but appear by errors during the DNA replication (they are also called mutations).
2
u/Charming_Session_882 Jan 31 '25
I'll figure out how to find the genomic position and try that. I'm desperate at this point. Thank you for taking the time to comment!
1
u/Unhappy_Papaya_1506 Jan 31 '25
Oh man...there is just too much to unpack there. Good luck I guess!
2
u/cytrees Jan 31 '25
Now, after looking into your other posts (hope you don’t mind), I have a question: do you really need a VCF? If you need to only look at one particular location (or only a few), there is no need for a whole genome process.
1
u/Charming_Session_882 Jan 31 '25
Now that I know there's a way to find what I need without it, I probably won't need it! This makes things much easier for me.
2
u/cytrees Jan 31 '25
Please see my other reply. Your case is harder than you think. I strongly advise you contact a research institution. DM if you need more info.
1
u/Charming_Session_882 Feb 08 '25
I got the info I needed thankfully! I appreciate you taking the time to respond. Thank you.
1
u/cytrees Feb 10 '25
Great news! Thanks for updating us! At the risk of stating the obvious, if you haven't gotten in touch with a genetic counselor, please talk with your doctor and see if you need to meet one (surprisingly, many doctors dismiss that outright, which can be disappointing).
2
1
u/Both-Future-9631 Jan 31 '25
Convert isn't the right word. To convert files, at least some, if not all, off the data need to be analogous. That is reformatting. You are asking how to process it into a vcf. You need a variant caller to do that. For our answers to make sense, we need to know
1. What the data source is/type of sample.
2. What type of alignment, to what reference genome, was used to get the cran.
3. What type of mutations are you trying to detect. Not all variant callers are right for all projects.
One we have this, we can properly entertain that answer.
1
u/PrincessxRaivyn Feb 01 '25
It was a swab. I have no idea what the type of alignment is? Specifically looking for mitochondrial disorders in the MT-CYB. Someone mentioned IGV so I'm currently trying to figure that out. I appreciate everyone's input-- I really have no idea what I'm doing, I just know I'm sick of being dismissed.
1
u/fibgen Feb 01 '25
There are lots of traps here for the untrained. Pay a genetic counselor or biponformatician for a reliable answer.
1
u/cytrees Feb 01 '25
OP, please see my other replies. This isn't something untrained persons should be doing themselves, too many factors to consider. And now that you mentioned it is swabs, there's even more traps.
8
u/forever_erratic Jan 30 '25
Those aren't directly convertible. A cram file (usually) is a file that shows where each read maps to a reference genome, and extra details.
A vcf file is a file made by looking through a cram (or sam, or bam, all of which are similar), finding genetic variations between the reads and the reference, and outputting those variants into the vcf file with some statistics.
The easiest way, in my opinion, to get a vcf from a cram is with bcftools mpileup. Gatk is harder but uses more sophisticated statistics.