Genomics

While the interrogatory tools of genome structure and function have become exponentially more precise and efficient in just the past five years, the infrastructure, procedures, and computational analysis required to generate and understand these vast amounts of data remain beyond the reach of most individual scientists. Equally inaccessible is the expertise to conduct genomics research in an ethical way that balances patient-centered innovation with acceptable risks for participants. Frontiers investigators who collaborate with the FGC receive access to teams of experts at the Genomic Medicine Center at Children’s Mercy Kansas City and the Genome Sequencing Facility at the University of Kansas Medical Center, who will consult on study design and ethical considerations to produce the highest-quality data and ensure they are handled appropriately.

Genomic Assays:

• Whole exome sequencing: Human whole exome sequencing performed with the Illumina TruSeq library preparation kit and the IDT v2 Exome baits targeting a 34 Mb region (19,433 genes) of the human genome, CNV backbone, and custom mitochondrial baits.
• Whole genome sequencing: Whole genome sequencing performed with the Illumina TruSeq library preparation kit, either with or without PCR.
• PacBio HiFi long-read whole genome sequencing: Highly accurate long reads detect all types of variants with high precision and recall, allow for haplotype phasing, and simultaneously measure the epigenome by detecting 5-methylcytosine at CpG sites.
• Illumina TruSeq ChIP-Seq: Selectively sequence DNA bound by target proteins.
• PGx panel: Human pharmacogenetics panel performed with the Illumina TruSeq library preparation kit and custom IDT baits targeting a 335 Mb region (~240 genes) of the human genome. Standard coverage of 47 samples plus 1 control per MiSeq run is available. If submitting fewer than 47 samples, the charge per sample will increase. Please contact the FGC Navigator for details on smaller batch costs by completing the form at the bottom of the page.

Transcriptomic Assays:

• RNA-seq: Generate mRNA and non-coding RNA libraries from total RNA that provide strand of origin for sequenced transcripts using Illumina’s Stranded Total RNA with RiboZero gold library preparation kit.
• PacBio Iso-Seq long-read RNA sequencing: Long-read RNA sequencing of intact, full-length transcripts allows unbiased, direct detection of all the isoforms present in a sample. This method can be used to profile whole transcriptomes exhaustively at the isoform level, characterize alternative start and polyadenylation sites as well as exon-skipping events, or detect isoforms as potential biomarkers.
• 10x Genomics Single Cell 3’ Gene Expression: Sequence mRNA at single-cell resolution to discover novel cell types and states, characterize complex cell populations, unravel complex cellular processes, and more.
• 10x Genomics Single Cell 5’ Gene Expression and V(D)J Enrichment: Simultaneously measure 5’ gene expression and profile full-length (5’ UTR to constant region), paired T-cell or B-cell receptor transcripts at single-cell resolution.
• Tecan Universal Plus mRNA-seq: Generate mRNA libraries directly from total RNA that provide strand of origin for sequenced mRNA transcripts. Oligo dT-based capture method.
• Illumina TruSeq RNA Exome: Convert total RNA into template molecules of known strand of origin coupled with sequence-specific probe capture of coding mRNA. Allows interrogation of low-yield / low-quality RNA.
• Illumina TruSeq Small RNA: Generate small RNA / miRNA libraries directly from total RNA.

Epigenomic Assays:

• PacBio HiFi long-read whole genome sequencing: Highly accurate long reads detect all types of variants with high precision and recall, allow for haplotype phasing, and simultaneously measure the epigenome by detecting 5-methylcytosine at CpG sites.
• Whole genome bisulfite sequencing: Detect methylated versus unmethylated DNA with whole genome bisulfite sequencing performed with Kapa’s Hyper Prep library preparation kit and Zymo’s EZ-96 DNA Methylation-Lightning MagPrep kit.
• Tecan Ovation RRBS Methyl-seq: Reduced representation bisulfite sequencing (RRBS) is used to generate single-base resolution DNA methylation information across a genomic sample. By analyzing a reduced representation of the genome, the amount of sequencing required is greatly reduced relative to whole genome bisulfite sequencing. The TrueMethyl oxBS module allows the differential interrogation of both 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC) and provides a method to accurately quantify the true level of cytosine methylation through the subtractive analysis of two parallel library sequence data sets, one with oxidative conversion of 5hmC and one without.
• 10x Genomics Single Cell ATAC: Profile chromatin accessibility at single-cell resolution. Requires optimized nuclei isolation. Please contact the FGC Navigator for more details by completing the form at the bottom of the page.
• 10x Genomics Single Cell Multiome: Simultaneously profile chromatin accessibility and expression of nuclear RNA at single-cell resolution. Requires optimized nuclei isolation. Please contact the FGC Navigator for more details by completing the form at the bottom of the page.

Assists with:

• Sequencing of investigator-provided libraries
• Study design
• Ethical, legal, and social issues
• Sample biobanking
• Access to bioinformatics tools and support

Access to genomic analyses by translational researchers can be limited by the volume of data generated and by requisite access to proprietary tools and big-data genomics resources. The FGC provides access to necessary workflows and applications through federated access to a cloud-based virtual genome compute platform where researchers can access their own data and have access to common workflows such as alignment, variant detection, and variant annotation. The FGC also serves as a point of entry for Frontiers investigators seeking access to public genomic resources, visualization tools, and consoles for interactive data analysis.

The unique nature of genomic data and familial implications of genomic findings require policies and procedures to support balancing innovation and collaboration while maintaining acceptable risks for research participants. The FGC aids Frontiers investigators in navigating a host of ethical, legal, and social issues (ELSI) such as informed consent and assent, enrollment of vulnerable and under-represented populations, and data sharing. Additionally, through consultation or collaboration with an FGC-specific ELSI Advisory Group and Patient and Parent Stakeholder Group, Frontiers investigators can gain support in addressing and studying ethical and social issues that emerge in translational genomics research. The FGC also provides consultation and resources to implement responsible data sharing that protects patient privacy and meets NIH and publisher data sharing requirements.

Frontiers Genomics Core (FGC) is excited to announce the installation of our new PacBio Revio long-read sequencer at Children’s Mercy Kansas City – among the first institutions to deploy this system! The Revio increases the throughput and affordability of generating, with industry-leading accuracy, reads that are approximately 100x longer (16,000 bp) than standard next-generation sequencers. HiFi reads can be used for whole-genome sequencing for de novo assembly, comprehensive variant detection, and epigenetic characterization by 5mC-detection – all in one sequencing test from the same DNA. Also, long-read RNA sequencing capturing full-length cDNA sequences can be performed at increased cost efficiency. Interested in exploring how HiFi sequencing can advance your research? Reach out to the FGC Navigator, Dan Louiselle (dalouiselle@cmh.edu), to connect with one of our experts.