I am a senior staff scientist at Washington University School of Medicine (Division of Oncology) with 10 years of experience in the fields of cancer genomics, bioinformatics, and computational biology. My research focuses on the development of software to aid in the interpretation of genomic mechanisms as they relate to cancer, as well as the utilization of these software in analyzing large cohorts across a breadth of cancer types. In addition to my primary research focus, I am involved in the creation of educational resources in the areas of bioinformatics, genomic data visualization, cancer genomics and transcriptomics, and precision medicine.
Zachary Skidmorea, Hans Rindtb, Shirley Chub, Bryan Fiska, Catrina Fronicka, Robert Fultona, Mingyi Zhoub, Nathan Bivensb, Carol Reinerob, Malachi Griffitha, Jeffrey Bryanb, Obi L. Griffitha
aWashington University, St. Louis, MO, USA; bUniversity of MO, Columbia, MO, USA
Canine models of cancers are increasingly being recognized as valuable tools in the exploration of disease. Further it is becoming increasingly apparent that a deeper understanding of the interaction between the immune system and tumorigenesis may yield promising new insights and therapies. To date however, tools and methods to investigate the interaction between canine cancers and their immune systems are wanting.
Here we demonstrate the use of custom dog-specific primers in conjunction with 10x cDNA libraries to produce scTCRseq and scRNAseq data. Briefly two independent canine samples responding to an autologous deglycosylated melanoma vaccine were selected to demonstrate the new methodology. Ficoll-separated peripheral blood mononuclear cells were obtained and custom primers for canine samples were designed for use with the Chromium Single Cell V(D)J Reagent kits. Samples were enriched for the alpha/beta V(D)J T-cell receptor region and libraries constructed for scTCRseq. GEX enriched libraries were also constructed according to kit protocols. Libraries were sequenced on the NovaSeq-6000 platform.
We observed a diverse profiling of possible V(D)J combinations for TCR alpha and beta exhibiting a majority of combinations. Further we observe the expansion of two dominant clonotypes for alpha and beta chains that mapped to a population of CD8+ T-cells. Gene expression for GEX enriched libraries also successfully identify large populations of CD8+ and CD4+ T cells overlapping with V(D)J-expressing cells. Together these data indicate the 10x kits were successfully adapted for canine samples.