Library quantification and quality control quick reference guide

03/06/20


Accurate quantification and proper quality check of next-generation sequencing libraries is key to a successful sequencing run. There are different methods for quantification and quality control depending on the sequencing library kit being used. In this bulletin, we summarize the library quantification and quality control methods for libraries prepared with all current Illumina kits. Use the methods listed in the tables below to validate the final library before sequencing.

DNA Kits – Research Use Only

Kits Quantification Quality Control
TruSeq Nano DNA Fluorometric Method or qPCR BioAnalyzer or Fragment Analyzer
TruSeq PCR Free qPCR BioAnalyzer or Fragment Analyzer
Nextera DNA Flex Fluorometric Method (QuBit/PicoGreen) BioAnalyzer or Fragment Analyzer
Nextera XT Fluorometric Method or Bead-based Normalization BioAnalyzer (Needed when using Fluorometric Method; Optional when using Bead-based Normalization)
Nextera Mate Pair qPCR/BioAnalyzer BioAnalyzer or Agarose Gel
Nextera DNA Flex for Enrichment QuBit BioAnalyzer
TruSeq Exome Fluorometric Method or qPCR BioAnalyzer or Fragment Analyzer
Nextera DNA Exome Fluorometric Method or qPCR BioAnalyzer or Fragment Analyzer
Nextera Rapid Capture Custom Enrichment Fluorometric Method or qPCR BioAnalyzer
TruSight Inherited Disease Fluorometric Method or qPCR BioAnalyzer
TruSight Tumor 15 Fluorometric Method (AccuClear/PicoGreen/QuBit) BioAnalyzer or Agarose Gel
TruSight Tumor 26 (TruSight Tumor) BioAnalyzer Gel/BioAnalyzer
Ampliseq (DNA Panels) BioAnalyzer/Fragment Analyzer/qPCR/QuBit/AccuClear/QuantIT BioAnalyzer or Fragment Analyzer
TruSight Myeloid Bead-based Normalization BioAnalyzer or Agarose Gel
TruSeq Amplicon Cancer Panel Bead-based Normalization BioAnalyzer or Agarose Gel
TruSeq Bovine Parentage Sequencing Panel Bead-based Normalization BioAnalyzer/Fragment Analyzer/Agarose Gel
TruSeq ChIP qPCR BioAnalyzer
TruSeq Methyl Capture EPIC Library Prep Kit QuBit BioAnalyzer

 

RNA Kits – Research Use Only

Kits Quantification Quality Control
TruSeq RNA v2 qPCR BioAnalyzer or Fragment Analyzer
TruSeq Stranded mRNA qPCR BioAnalyzer or Fragment Analyzer
TruSeq Stranded Total RNA qPCR BioAnalyzer or Fragment Analyzer
TruSeq RNA Exome qPCR/Fluorometric Method BioAnalyzer or Fragment Analyzer
TruSight RNA Pan-Cancer qPCR BioAnalyzer or Fragment Analyzer
TruSight RNA Fusion qPCR BioAnalyzer or Fragment Analyzer
TruSeq Targeted RNA Expression BioAnalyzer or Fragment Analyzer BioAnalyzer or Fragment Analyzer
TruSeq Small RNA BioAnalyzer BioAnalyzer
Ampiseq (RNA Panels) BioAnalyzer/Fragment Analyzer/qPCR/QuBit/AccuClear/QuantIT BioAnalyzer or Fragment Analyzer
SureCell WTA 3' BioAnalyzer BioAnalyzer

 

DNA and RNA kits - Research Use Only

Kits Quantification Quality Control
TruSight Tumor 170 Fluorometric Method/Bead-based Normalization Not Needed

 

Molecular Diagnostics Kits

Kits Quantification Quality Control
Cystic Fibrosis 139-Variant Bead-based Normalization Not Needed
Cystic Fibrosis Clinical Sequencing Bead-based Normalization Not Needed

 

Notes:

  • Fluorometric method = a double strand DNA-specific fluorescent dye method, such as Qubit, PicoGreen, or AccuClear
  • Additional information on qPCR quantification can be found in the Sequencing Library qPCR Quantification guide
  • Not Needed = quality control is not needed with bead-based normalized libraries