Genome Analysis Technology Core

 

To continue to receive funding from the Cancer Center and School Of Medicine, we are encouraged to demonstrate the core's impact on scientific output in the form of a citation in publications and/or presentations generated using the core services.  PLease let us know what services were used by include the core name and core ID  on your slides and peered review publications  

eg.: This work was supported by the UVA Genome Analysis and Technology Core, RRID:SCR_018883. 

When  appropriate thank the core staff by name or include them as co-Authors.  We routinely provide Method sections write up and proof reading.  

 

Mission Statement

The GATC continuing mission is to serve as a scientific resource and biotechnology hub enhancing the scope and quality of basic and translational research at UVA, and to provide customer satisfaction, collaboration and quality control.The goal of the core staff is to enable high throughput (Next Generation) sequencing projects for both novice and experienced users alike, by providing timely assistance in all facets of experimentation from project design/execution to data analysis/reporting. 

Brief Overview of Services

COMING SOON    Revio PacBio Long Read library prep and sequencing services -- Full sevice or  "a la carte"

• Kinnex Full-Length RNA Kit for Isoform Sequencing full service includes:  

Total RNA QC:  Using the Agilent Tape Station and Qubit
cDNA synthesis and cleanup using Iso-seq Express 2,0 
cDNA amplification and cleanup: generates up to 12 barcoded cDNAs
Kinnex PCR: Set-up  from cDNA pool, 8 parallel Kinnex PCR reactions to generate orientation specific Kinnex segmentation fragments.
Kinnex Arrary formation: Pool PCR and array assembly via ligation.  each Array includes a kiinnex SMRT Bell.  Kit supports  4-plex Kinnex SMRT Bell
Nuclease treatment and final cleanup: Includes Full Array QC and sizing using the Agilent Femto PulseNA 
Preparation of the SMRT Bell library for Sequencning on the Revio (24 to 30 hours)

• Kinnex Single Cell RNA Kit:  Starts from single cell indexed cDNAs

cDNA QC:  Using the Agilent Tape Station and Qubit.  
TSO PCR and clean up: remove cDNA artifacts
Kinnex PCR: Set-up  16 parallel Kinnex PCR reactions to generate orientation specific Kinnex segmentation fragments.
Kinnex Arrary formation: Damage repair, A-tailing, and adapter ligation.  Support up to 4-plex Kinnex SMRT Bell adapters
Nuclease treatment and final cleanup: Includes Full Array QC and sizing using the Agilent Femto Pulse
Preparation of the SMRT Bell library for Sequencning on the Revio (24 to 30 hours) 

• Whole Genome SMRTBell Prep Kit 3.0

gDNA QC:  Uses the Femto pulse on Native DNA  
DNA Shearing optimization utilize the Femto Pulse for QC

Sheared DNA repair and tailing Femto Pulse QC and Qubit
Adapter ligation and clean up
Nuclease treatment and final cleanup: Includes sizing using the Agilent Femto Pulse
Preparation of the SMRT Bell library for Sequencning on the Revio (30 hours) 

 

10x Genomics library prep services

 Single cell or nuclei preps are submitted to the core for inclusion into any of the protocols below. 

Overall, emulsions are generated on the Chromium X to enable production of cell specific barcoded cDNAs as well as identify through Feature barcodes expression at the surface of each cell (proteogenomics).  The cell specific cDNAs can also be used as substrate for the generation of long read isoform sequencing (see  Kinnex single cell service above).  In case of ATAC seq single nuclei are subjected to transposase to label open chromatin structures prior to emulsion. Multiple indexed libraries are routinrly pooled for seqeucning on the Illumina NextSeq 2000.   

• GEM-X - 5' and 3' RNA seq Library prep
• GEM-X VDJ - T or B cell with 5' RNAseq
• ATACseq and MultiOmics (ATAC-seq and GEX)
• Cell-Plex and Feature barcode with 3' or 5' RNAseq
• Fixed Cell (probe based)

Illumina Single cell Library prep- PIP-Seq (under development)

• T2 prep: 5K input cells and 100M reads per sample.
• T10 prep: 17K input cells and 340M reads per sample.
• T20 prep: 40K input cells and 800M reads per sample.
• T100 prep:200K input cells and 4B reads per sample.

 

Illumina NextSeq2000 sequencer (NSQ2K)

 

 

 

Indexed Illumina short read libraires are routinely pooled and sequenced on the NextSeq 2000 sequencer using one of the flow cell as indicated below.  Short read sequencing  is routinely used for differential gene expression of mRNA, whole transcriptome analysis, genomic variant detection, metogenomics studies and other genomic applications.  

• Flow cell reads per run output:
• 100 Million (P1)
• 400 Million (P2)
• 1.2 Billion (P3)
• 1.8  Billion (P4) 
• Sequencing  single or paired end (PE) read legth:
• 50 bp (PE 2X25)
• 100bp (PE 2X50)
• 200bp (PE 2X100)
• 300bp (PE 2X150)

Illumina MiSeq

The instrument is soon to be discontinued by the vendor, we will continue to maintain our services until time when Kits are no longer available.  

Note for Single cell our QCruns have now been moved from the Miseq nano and Micro kit to the NextSeq2000 P1-100 flow cell.  

•  Nano 1.2 millions or  Micro 3 million reads 300 cycle kits
• V2 (12-14 Million reads) and V3 (24million reads) chemistries offered

 

Library Prep for Illumina's NextSeq and MiSeq

 

Leadership

 

Katia Sol-Church, Ph.D, Director

Location and hours of operation

Hours	Location
Monday - Friday              9AM - 5PM	Pinn Hall Rm 1076 and 1044

Links and Resources

UVA RESEARCH CORES HOME PAGE

 

GATC Home Page