The Genome Analysis Core

National Institute Of Allergy And Infectious Diseases

Investigators

Linked publications, trials & patents

Abstract

The Genome Analysis Core (TGAC), within the Human Immunology Section, was set up in 2014 to fulfill many of the specialist sequencing needs of the Vaccine Research Center (VRC). Currently, the VRC is designing, producing and testing candidate immunogens aimed at eliciting broadly neutralizing antibodies against HIV and other infectious agents. At the core of this effort lies the requirement to elicit antibodies that use particular IgH and IgL chain gene pairs which undergo a sequential series of somatic hypermutations (SHM) to achieve effective function. For evaluation of vaccine candidates, the evolution of such antibodies, and thus the effectiveness of the vaccine, needs to be monitored using a variety of molecular techniques that culminate in single-cell NGS of specific B cell Ig genes. In addition, passive antibody applications necessarily depend on the identification and sequencing of broadly neutralizing Env-specific antibodies in HIV-infected people. TGAC has developed a number of next-generation sequencing methodologies. These include: bulk and single cell 5RACE for unbiased TCR and Ig sequencing, genomic baiting for targeted sequencing of Ig loci, bulk mRNAseq and single cell mRNAseq for human, mouse and NHP. In addition, a number of approaches for sequencing HIV have been developed which we have used to characterize the HIV reservoir in infected people, as part of cure studies, and also to monitor the selection of escape variants in clinical studies of passive administration of Env-specific bnAbs. We have purchased two 10x Genomic Chromium platforms and we are currently using these to increase our throughput of single cell 5 and 3 transcriptome sequencing. We have successfully developed and implemented a custom primer library prep approach so that we may adapt the standard 10x 5 immunoglobulin and T cell receptor sequencing protocol for use with samples from rhesus and cynomolgus macaques, and Guinea pigs. In addition, we have increased the sensitivity of the library prep by 100-fold. We have implemented CITE-seq applications using custom conjugated antibodies to perform high parameter cell surface proteomics. Furthermore, many other ongoing research projects at the VRC such as those for influenza, RSV, malaria and TB vaccines require Ig, TCR and transcriptome sequencing as described above. Finally, we have a robust microbiome sequencing program.