Guida veloce per quantificazione e controllo qualità (QC) delle librerie NGS

La corretta quantificazione delle librerie ed un appropriato controllo della loro qualità sono fondamentali per la buona riuscita di un sequenziamento. In funzione del kit utilizzato per la preparazione delle librerie esistono differenti metodi di quantificazione e QC. Il presente bollettino fornisce un elenco di metodi compatibili con le attuali librerie NGS preparate con kit Illumina. Utilizzare i metodi riportati in tabella per validare correttamente la concentrazione finale della libreria prima di procedere al sequenziamento.

Kits DNA – Solo a scopo di ricerca

Kits	Quantificazione	Controllo Qualità (QC)
TruSeq Nano DNA	Metodo Fluorimetrico o qPCR	BioAnalyzer o Fragment Analyzer
TruSeq PCR Free	qPCR	BioAnalyzer o Fragment Analyzer
Nextera DNA Flex	Metodo Fluorimetrico (QuBit/PicoGreen)	BioAnalyzer o Fragment Analyzer
Nextera XT	Metodo Fluorimetrico o Normalizzazione Bead-based	BioAnalyzer (necessario con Metodo Fluorimetrico; opzionale con Normalizzazione Bead-based)
Nextera Mate Pair	qPCR/BioAnalyzer	BioAnalyzer o Agarose Gel
Nextera DNA Flex for Enrichment	QuBit	BioAnalyzer
TruSeq Exome	Metodo Fluorimetrico o qPCR	BioAnalyzer o Fragment Analyzer
Nextera DNA Exome	Metodo Fluorimetrico o qPCR	BioAnalyzer o Fragment Analyzer
Nextera Rapid Capture Custom Enrichment	Metodo Fluorimetrico o qPCR	BioAnalyzer
TruSight Inherited Disease	Metodo Fluorimetrico o qPCR	BioAnalyzer
TruSight Tumor 15	Metodo Fluorimetrico (AccuClear/PicoGreen/QuBit)	BioAnalyzer o Agarose Gel
TruSight Tumor 26 (TruSight Tumor)	BioAnalyzer	Gel/BioAnalyzer
Ampliseq (DNA Panels)	BioAnalyzer/Fragment Analyzer/qPCR/QuBit/AccuClear/QuantIT	BioAnalyzer o Fragment Analyzer
TruSight Myeloid	Normalizzazione Bead-based	BioAnalyzer o Agarose Gel
TruSeq Amplicon Cancer Panel	Normalizzazione Bead-based	BioAnalyzer or Agarose Gel
TruSeq Bovine Parentage Sequencing Panel	Normalizzazione Bead-based	BioAnalyzer/Fragment Analyzer/Agarose Gel
TruSeq ChIP	qPCR	BioAnalyzer
TruSeq Methyl Capture EPIC Library Prep Kit	QuBit	BioAnalyzer

Kits RNA – Solo a scopo di ricerca

Kits	Quantificazione	Controllo Qualità (QC)
TruSeq RNA v2	qPCR	BioAnalyzer o Fragment Analyzer
TruSeq Stranded mRNA	qPCR	BioAnalyzer o Fragment Analyzer
TruSeq Stranded Total RNA	qPCR	BioAnalyzer o Fragment Analyzer
TruSeq RNA Exome	qPCR/Metodo Fluorimetrico	BioAnalyzer o Fragment Analyzer
TruSight RNA Pan-Cancer	qPCR	BioAnalyzer o Fragment Analyzer
TruSight RNA Fusion	qPCR	BioAnalyzer o Fragment Analyzer
TruSeq Targeted RNA Expression	BioAnalyzer o Fragment Analyzer	BioAnalyzer o Fragment Analyzer
TruSeq Small RNA	BioAnalyzer	BioAnalyzer
Ampiseq (RNA Panels)	BioAnalyzer/Fragment Analyzer/qPCR/QuBit/AccuClear/QuantIT	BioAnalyzer o Fragment Analyzer
SureCell WTA 3'	BioAnalyzer	BioAnalyzer

Kits DNA e RNA - Solo a scopo di ricerca

Kits	Quantificazione	Controllo Qualità (QC)
TruSight Tumor 170	Metodo Fluorimetrico o Normalizzazione Bead-based	Non Necessario

Kits per Diagnostica Molecolare

Kits	Quantificazione	Controllo Qualità (QC)
Cystic Fibrosis 139-Variant	Normalizzazione Bead-based	Non Necessario
Cystic Fibrosis Clinical Sequencing	Normalizzazione Bead-based	Non Necessario

Notes:

Metodo Fluorimetrico = metodo basato sull'utilizzo di un fluoroforo specifico per molecole di DNA double strand, e.g. Qubit, PicoGreen o AccuClear
Ulteriori informazioni relative alla quantificazione via qPCR si possono trovare al seguente link: Sequencing Library qPCR Quantification guide
Non Necessario = in caso di normalizzazione Bead-based non è necessario effettuare alcun QC

Guida veloce per quantificazione e controllo qualità (QC) delle librerie NGS

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Guida veloce per quantificazione e controllo qualità (QC) delle librerie NGS

Contact Us

Technical Support

Share With Tech Support

Other Support

Contact Us

Technical Support

techsupport@illumina.com

Other Support