I am working on getting expressed, paired IGH and IGL (immunoglobin) sequences from data we acquired by designing custom primers for IGH and IGL in horse and then preparing the samples using a 10X kit as we normally do with mouse, human, etc. Sadly, the horse genome has terrible annotations for IG genes, and I have been having to create my own custom reference manually. I am wondering if anyone else has had to do this, design a custom VDJ reference, with what I'll call a "second-tier" genome such as cow, horse, cat, dog, ferret, etc.
After lots of trial and error, my reference is working but not very well, with only a small portion of the cells being called as containing IG genes and very few of these genes having pairing information.
The VDJ gene annotations from Ensembl were basically trash, but the tools and sequences from IGMT were actually quite good.
I have the IGL part of the reference working well but the IGH gene references still have problems. I'd love to speak with anyone who has gone through a similar process. I'd also love to hear from anyone with experience with tools that predict the V, D, J, and C regions of IG genes. The IMGT/V-QUEST tool has helpful for the IGH sequences but does not work at all for the IGL sequences.
10x tech support can be a great resource. They are generally responsive and can help you out.
Thanks, I have contacted them and they have been helpful. Unfortunately they really don't have experience outside of human and mouse, so problems with bad genome annotations aren't something they encounter.