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Rochester Genomics Center

URMC / Research / Rochester Genomics Center / Services / High Throughput Sequencing

 

High Throughput Sequencing Library Preparations

IlluminaKit+Sciclone

The GRC uses several kit-based workflows supplied by the following:

Illumina Logo, courtesy of Illumina, Inc.   

  • TruSeq
  • Illumina Stranded 
  • Nextera 
     

Takara/Clontech Logo, Courtesy of Takara

  • SMARTer Ultra-Low Solutions
  • SMARTer Human/Mouse TCR Profiling 
     

New England BioLabs, Inc

  • NEBNext Small RNA 
  • NEBNext Ultra II DNA
     

Agilent Technologies Logo, Courtesy of Agilent Technologies

  • SureSelect
     

Perkin Elmer Logo

  • Bisulfite-Seq
    Methyl-Seq
     

We also offer custom library preparations.
Please contact us for more information.

Transcriptomics

Library Preparation Total RNA Input Target Capture Application
Illumina TruSeq Stranded mRNA-seq (Legacy Version*) 200ng Poly-A RNA mRNA detection and differential expression, splice variants, rare transcripts complementary SNPs, captures both coding and non-coding transcripts that are poly adenylated, provides strand information, gene fusions 
Illumina Stranded mRNA  25-1000 ng  Poly-A RNA New features that build upon the TruSeq mRNA Technology:
- Broad range of input down to 25 ng
- Multiplex up to 384 samples in a single sequencing run
Illumina TruSeq Stranded Total RNA-seq (Legacy Version*) 200ng Ribosomal RNA Depletion (RiboZero) coding and multiple forms of non-coding RNAs, mRNA detection and differential expression, splice variants, rare transcripts, complementary SNPs, gene fusions, lincRNAs, works well for low quality RNA samples and FFPE samples
Illumina Stranded Total RNA Prep with Ribo-Zero Plus 1-1000ng Enzymatic rRNA Depletion (RiboZero Plus)  New features that build upon the TruSeq Total Technology:
- Broad range of input down to 1 ng
- Multiplex up to 384 samples in a single sequencing run
- New RiboZero Plus - removes abundant RNA from human, mouse, rat, and bacteria
Ultra-Low RNA Input (Clontech SMART-seq v4) 1 ng Poly A RNA Alternative to TruSeq/Illumina Stranded mRNA Prep for low input samples: 

Low input preparation for high quality RNA. Full length transcriptome analysis.
mRNA detection and differential expression, splice variants, rare transcripts, complementary SNPs
SMARTer Stranded Total RNA Kit - Pico Input 250 pg - 10ng rRNA Depletion Alternative to TruSeq/Illumina Stranded Total RNA Prep for low input samples: 

Low input preparation for high or low quality RNA. Removal of cDNAs from rRNA. Includes UMI (Unique Molecular Identifier) to mitigate PCR bias. 
NEB Small RNA 500ng 3' and 5' adaptor ligation of small RNA small regulatory RNAs, including micro RN
*TruSeq Legacy Version Note: We are in a transition period with respect to our Stranded mRNA and Stranded Total library preps. The newer "Illumina mRNA" and "Illumina Total RNA" library preps will eventually become our standard service for their respective library prep projects, replacing the legacy TruSeq library preps (also manufactured by Illumina). We will continue to offer TruSeq library preparations for those investigators who are comparing new projects with previous datasets generated by TruSeq library preps. For more information on how the kits compare, click here. 

Genomics

Library Preparation DNA Input Target Capture Application
TruSeq DNA PCR-Free 1-2ug DNA Whole genome sequencing, SNP/InDel indentification, high GC-rich regions, promoters, and repetitive content, reduced library bias and gaps
Nextera XT 1ng DNA Whole genome sequencing, small genomes, amplicons larger than 500bp, Enzymatic Tagmentation
Illumina DNA Prep
(Nextera Flex)		 100-500ng DNA Whole genome sequencing, SNP/InDel identification, gene fusions
Nextera Mate-Pair 1-4ug DNA Whole genome sequencing, De Novo sequencing, genome assemblies
Nextera Rapid Whole Exome 50ng DNA Exome sequencing, 37Mb or 62Mb (expanded) target baits, custom targets also available
Agilent SureSelect Whole Exome 4ug DNA Exome sequencing, 48.2mB+UTRs (version 7), other options available, custom targets also available
NEBNext Ultra II FS 100 pg - 500 ng DNA DNA library preparation from NEB that includes a new DNA fragmentation reagent that results in reliable fragmentation regardless of DNA input amount or GC content. 

Amplicon

Library Preparation Application
T-Cell Repertoire Sequencing  (Human and Mouse)  - Takara SMARTer Human and Mouse TCR a/b Profiling using the SMART full-length cDNA synthesis technology
- Capture complete V(D)J variable regions of TRA and TRB genes
B-Cell Repertoire Sequencing (Human and Mouse)  - Takara SMARTer Human and Mouse BCR Profiling using the 5' RACE method to reduce variability and allows for priming from the constant region of BCR heavy or light chains. 
- Gene-specific amplification to capture complete V(D)J vairable regions of BCR transcripts
  • 16S Sequencing 
    • 16S library preparations benefit from the inclusion of phased primers to help increase library diversity and sequencing quality. Click here for a helpful resource on designing heterogeneity spacers (aka Phased Primers).