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文库定量与质量控制简明指引

测序文库精确的定量与准确的质量检测对于二代测序成功是十分关键的。对于不同测序文库试剂盒,我们推荐使用不同的定量和质量控制的方法。在本篇技术文档中,我们总结了目前所有Illlumina建库试剂盒的文库定量与质量控制的方法。在测序前,利用下表中的方法对最终文库进行鉴定。
DNA试剂盒-仅用于科研
试剂盒
定量
质量控制
TruSeq Nano DNA
荧光方法或qPCR
BioAnalyzer 或 Fragment Analyzer
TruSeq PCR Free
qPCR
BioAnalyzer 或 Fragment Analyzer
Illumina DNA Prep, (M) Tagmentation (formerly Nextera DNA Flex)
荧光方法(Qubit/PicoGreen)
BioAnalyzer 或 Fragment Analyzer
Nextera XT
荧光方法或基于磁珠均一化
BioAnalyzer (当使用荧光方法时需要;基于磁珠进行均一化时为可选)
Nextera Mate Pair
qPCR/BioAnalyzer
BioAnalyzer或琼脂糖凝胶
Illumina DNA Prep with Enrichment, (S) Tagmentation (formerly Nextera DNA Flex for Enrichment)
Qubit
BioAnalyzer
Illumina DNA PCR-Free Prep, Tagmentation
Qubit ssDNA或基于投入量的qPCR, 具体细节可参照 Reference Guide
N/A (最终文库为单链DNA)
TruSeq Exome
荧光方法或qPCR
BioAnalyzer 或 Fragment Analyzer
Nextera DNA Exome
荧光方法或qPCR
BioAnalyzer 或 Fragment Analyzer
Nextera Rapid Capture Custom Enrichment
荧光方法或qPCR
BioAnalyzer
TruSight Inherited Disease
荧光方法或qPCR
BioAnalyzer
TruSight Tumor 15
荧光方法(AccuClear/PicoGreen/Qubit)
BioAnalyzer或琼脂糖凝胶
TruSight Tumor 26 (TruSight Tumor)
BioAnalyzer
琼脂糖凝胶/BioAnalyzer
Ampliseq (DNA Panels)
BioAnalyzer/Fragment Analyzer/qPCR/Qubit/AccuClear/QuantIT
BioAnalyzer 或 Fragment Analyzer
TruSight Myeloid
基于磁珠均一化
BioAnalyzer或琼脂糖凝胶
TruSeq Amplicon Cancer Panel
基于磁珠均一化
BioAnalyzer或琼脂糖凝胶
TruSeq Bovine Parentage Sequencing Panel
基于磁珠均一化
BioAnalyzer/Fragment Analyzer/Agarose Gel
TruSeq ChIP
qPCR
BioAnalyzer
TruSeq Methyl Capture EPIC Library Prep Kit
Qubit
BioAnalyzer
RNA 试剂盒-仅用于科研
试剂盒
定量
质量控制
TruSeq RNA v2
qPCR
BioAnalyzer 或 Fragment Analyzer
TruSeq Stranded mRNA
qPCR
BioAnalyzer 或 Fragment Analyzer
TruSeq Stranded Total RNA
qPCR
BioAnalyzer 或 Fragment Analyzer
TruSeq RNA Exome
qPCR/荧光方法
BioAnalyzer 或 Fragment Analyzer
Illumina Stranded mRNA Prep, Ligation
Qubit
BioAnalyzer
Illumina Stranded Total RNA Prep with Ribo-Zero Plus
Qubit
BioAnalyzer
Illumina RNA Prep with Enrichment, (L) Tagmentation
Qubit
BioAnalyzer
TruSight RNA Pan-Cancer
qPCR
BioAnalyzer 或 Fragment Analyzer
TruSight RNA Fusion
qPCR
BioAnalyzer 或 Fragment Analyzer
TruSeq Targeted RNA Expression
BioAnalyzer 或 Fragment Analyzer
BioAnalyzer 或 Fragment Analyzer
TruSeq Small RNA
BioAnalyzer
BioAnalyzer
Ampiseq (RNA Panels)
BioAnalyzer/Fragment Analyzer/qPCR/Qubit/AccuClear/QuantIT
BioAnalyzer 或 Fragment Analyzer
SureCell WTA 3'
BioAnalyzer
BioAnalyzer
DNA和RNA试剂盒-仅用于科研
试剂盒
定量
质量控制
TruSight Tumor 170
Fluorometric Method/基于磁珠均一化
不需要
TruSight Oncology 500
AccuClear/基于磁珠均一化
不需要
TruSight Oncology 500 High Throughput
AccuClear/基于磁珠均一化
不需要
TruSight Oncology 500 ctDNA
AccuClear/基于磁珠均一化
不需要
分子诊断试剂盒
Kits
定量
质量控制
Cystic Fibrosis 139-Variant
基于磁珠均一化
不需要
Cystic Fibrosis Clinical Sequencing
基于磁珠均一化
不需要
TruSeq Custom Amplicon Dx
基于磁珠均一化
不需要
注意:
· 荧光方法为双链DNA特异荧光染料方法,例如QuBit, PicoGreen和AccuClear。
· qPCR定量方法更多的信息可以参考qPCR guide
· 基于磁珠进行均一化的文库不需要进行质量控制。
For any feedback or questions regarding this article (Illumina Knowledge Article #7296), contact Illumina Technical Support [email protected].
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