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Anthocyanin Compound Library (20 Anthocyanins)

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Anthocyanin Compound Library (20 Anthocyanins)

品牌：Tokiwa Phytochemical
CAS No.：
储存条件：-20℃
纯度：–
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PL2101

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Next Ultrashear® FFPE DNA Library Prep Kit |NEB酶试剂 New England Biolabs

发表于由

上海金畔生物科技有限公司代理New England Biolabs（NEB）酶试剂全线产品，欢迎访问官网了解更多产品信息和订购。

产品信息

View or download extensive performance data in our Data Supplement.

FFPE DNA is often compromised in quality and quantity and is challenging for library prep and sequencing, including in target enrichment workflows that require high amounts of library input. The NEBNext UltraShear FFPE DNA Library Prep Kit addresses these challenges and improves ease of workflow and scalability:

  • Repairs FFPE-induced damage using the NEBNext FFPE DNA Repair Mix v2
  • Incorporates specialized enzymatic DNA fragmentation using NEBNext UltraShear
  • Uses NEBNext Ultra II library prep reagents in a protocol optimized for FFPE DNA
  • Robustly amplifies libraries using NEBNext MSTC™ FFPE Master Mix
  • High library yields, sufficient for input into target enrichment workflows
  • User-friendly workflow with minimal hands-on time and multiple safe stopping points

The NEBNext UltraShear FFPE DNA Library Prep Kit reduces damage from deamination and oxidation due to the fixation, storage, and extraction process for FFPE DNA samples, thereby reducing false positive variant calls.

The combination of specialized enzymatic FFPE DNA fragmentation and repair with optimized reagents and protocols provides superior performance with this challenging sample type.

For FFPE DNA library prep without enzymatic fragmentation, the NEBNext FFPE DNA Library Prep Kit (NEB #E6650) is available.

FIGURE 1: NEBNext UltraShear FFPE DNA Library Prep Kit workflow overview

Next Ultrashear® FFPE DNA Library Prep Kit |

The NEBNext UltraShear FFPE DNA Library Prep Kit has a streamlined workflow with minimal hands-on time across a range of inputs from 5-250 ng. The protocol has been optimized for the user to safely store the reaction after any step in the workflow overnight at -20°C without affecting library yield or quality.
FIGURE 2: The NEBNext UltraShear FFPE DNA Library Prep Kit enables robust library preparation from a range of sample inputs and quality

Next Ultrashear® FFPE DNA Library Prep Kit |

Libraries were prepared from 5, 50, or 250 ng of normal tissue FFPE DNA ranging in quality from DNA Integrity Number (DIN) 1.8 to 6.8, with the indicated PCR cycles. Libraries were prepared in triplicate for 5 ng and 50 ng input and 1 replicate for 250 ng. Each bar represents the average of triplicates with error bars indicating standard deviation for the 5 and 50 ng inputs. Robust library yields were obtained across sample qualities and input amounts. Most target enrichment workflows require a 200 ng library for hybrid capture input. Sufficient library yield can be obtained using a minimum of 50 ng FFPE DNA with the NEBNext UltraShear FFPE DNA Library Prep Kit.    

FIGURE 3: The NEBNext UltraShear FFPE DNA Library Prep Kit improves library quality

Next Ultrashear® FFPE DNA Library Prep Kit |    

Libraries were prepared in duplicate from 100 ng of low quality, normal tissue FFPE DNA (DIN 1.8) and 9 PCR cycles, using the NEBNext UltraShear™ FFPE DNA Library Prep Kit. Results were compared to other enzymatic fragmentation-based library prep kits that have been validated for use with FFPE samples, using each vendor’s own recommended adaptors (IDT® xGen® EZ UNI, Kapa EvoPlus® Library Prep Kit, QuantaBio® sparQ DNA Library Prep Kit, and Twist Library Preparation EF 2.0 kit). Libraries were sequenced on the Illumina® NovaSeq®6000 (2 x 100 base reads) and downsampled to 5 million paired-end reads. Reads were mapped using Bowtie2 (version 2.3.2.2) to the GRCh38 reference and duplicates were marked using Picard MarkDuplicates (version 1.56.0). Library quality metrics were assessed using Picard Alignment Summary Metrics (version 1.56.0). The level of foldback reads was calculated using Seq_frag_remap (version 0.2). The NEBNext UltraShear FFPE DNA Library Prep Kit improves library quality by reducing the percentage of unmapped, chimeric, non-properly paired, and foldback reads.
FIGURE 4: The NEBNext UltraShear FFPE DNA Library Prep Kit reduces sequencing artifacts

Next Ultrashear® FFPE DNA Library Prep Kit |

Libraries were prepared in duplicate from 100 ng of low quality, normal tissue FFPE DNA (DIN 1.8) and 9 PCR cycles, using the NEBNext UltraShear FFPE DNA Library Prep Kit. Results were compared to other enzymatic fragmentation-based library prep kits that have been validated for use with FFPE samples, with each vendor’s own recommended adaptors (IDT xGen EZ UNI, Kapa EvoPlus Library Prep Kit, QuantaBio sparQ DNA Library Prep Kit, and Twist Library Preparation EF 2.0 kit). Libraries were sequenced on the Illumina NovaSeq 6000 (2 x 100 base reads) and downsampled to 5 million paired-end reads. Reads were mapped using Bowtie2 (version 2.3.2.2) to the GRCh38 reference and duplicates marked using Picard MarkDuplicates (version 1.56.0). The average frequency of C→T mutations at each C position (A) and G→T mutations at each G position (B) in Read 1 and 2 was calculated for two technical replicates using Tasmanian (version 1.0.7). C→T mutations arising from cytosine deamination and G→T mutations arising from oxidative damage in low quality FFPE DNA are effectively repaired by the NEBNext FFPE DNA Repair v2 Mix in the NEBNext UltraShear FFPE DNA Library Prep Kit. Other kits show a high level of C→T and G→T artifacts in low quality FFPE DNA due to a lack of DNA damage repair.

FIGURE 5: The NEBNext UltraShear FFPE DNA Library Prep Kit enables superior on-target coverage in hybrid capture sequencing

Next Ultrashear® FFPE DNA Library Prep Kit |

Libraries were prepared in duplicate from 100 ng of low quality, normal tissue FFPE DNA (DIN 1.8) and 9 PCR cycles, using the NEBNext UltraShear™ FFPE DNA Library Prep Kit. Results were compared to other enzymatic fragmentation-based library prep kits that have been validated for use with FFPE samples, with each vendor’s own recommended adaptors (IDT xGen EZ UNI, Kapa EvoPlus Library Prep Kit, QuantaBio sparQ DNA Library Prep Kit, and Twist Library Preparation EF 2.0 kit). The full library yield was used in singleplex target enrichment with a custom cancer panel (Twist Bioscience®) and libraries were sequenced on the Illumina NovaSeq 6000 (2 x 100 base reads). 15 million paired-end reads were trimmed with Fastp (version 0.20.0) and mapped with BWA mem (version 0.7.17) to the T2T reference. Duplicates were marked using Picard MarkDuplicates (version 2.20.6) with UMI. Target enrichment quality metrics were assessed using Picard HS Metrics (version 2.18.29). The improved yield, coverage, and fraction of usable reads observed in NEBNext UltraShear FFPE DNA Library Prep Kit whole genome sequencing (WGS) libraries correlates to improved coverage, on-target rate, and coverage uniformity in target enrichment libraries.

产品类别:
FFPE DNA Products,
Next Generation Sequencing Library Preparation Products

  • 试剂盒组成

    本产品提供以下试剂或组分：

    NEB # 名称 组分货号 储存温度 数量 浓度
    • E6655S     Multi-temperature    
        NEBNext Ultrashear® FFPE DNA Library Prep Kit E6655-2 -20    
        NEBNext® FFPE DNA Repair Mix v2 E7361AVIAL -20 1 x 0.048 ml Not Applicable
        NEBNext® Thermolabile Proteinase K E7362AVIAL -20 1 x 0.048 ml Not Applicable
        NEBNext UltraShear™ Reaction Buffer E6657AVIAL -20 1 x 0.168 ml Not Applicable
        NEBNext UltraShear™ E6656AVIAL -20 1 x 0.096 ml Not Applicable
        500mM DTT E6658AVIAL -20 1 x 0.048 ml Not Applicable
        NEBNext® Ultra II End Prep Enzyme Mix E7646AVIAL -20 1 x 0.072 ml Not Applicable
        NEBNext® Ultra™ II Ligation Master Mix E7648AVIAL -20 1 x 0.72 ml Not Applicable
        NEBNext® Ligation Enhancer E7374AVIAL -20 1 x 0.024 ml Not Applicable
        NEBNext® MSTC FFPE Master Mix E6651AVIAL -20 1 x 0.6 ml 2 X
        NEBNext® Sample Purification Beads E6652S 25    
        NEBNext® Sample Purification Beads E6652AVIAL 25 1 x 4.3 ml Not Applicable
    • E6655L     Multi-temperature    
        NEBNext Ultrashear® FFPE DNA Library Prep Kit E6655-3 -20    
        NEBNext® FFPE DNA Repair Mix v2 E7361AAVIAL -20 1 x 0.192 ml Not Applicable
        NEBNext® Thermolabile Proteinase K E7362AAVIAL -20 1 x 0.192 ml Not Applicable
        NEBNext UltraShear™ Reaction Buffer E6657AAVIAL -20 1 x 0.672 ml Not Applicable
        NEBNext UltraShear™ E6656AAVIAL -20 1 x 0.384 ml Not Applicable
        500mM DTT E6658AAVIAL -20 1 x 0.192 ml Not Applicable
        NEBNext® Ultra II End Prep Enzyme Mix E7646AAVIAL -20 1 x 0.288 ml Not Applicable
        NEBNext® Ultra™ II Ligation Master Mix E7648AAVIAL -20 3 x 0.96 ml Not Applicable
        NEBNext® Ligation Enhancer E7374AAVIAL -20 1 x 0.096 ml Not Applicable
        NEBNext® MSTC FFPE Master Mix E6651AAVIAL -20 2 x 1.2 ml 2 X
        NEBNext® Sample Purification Beads E6652L 25    
        NEBNext® Sample Purification Beads E6652AAVIAL 25 1 x 17 ml Not Applicable

  • 特性和用法

    需要但不提供的材料

    • 80% Ethanol
    • Nuclease-free Water
    • 0.1X TE (1 mM Tris-HCl, pH 8.0, 0.1 mM EDTA)
    • DNase-, RNase-free PCR strip tubes
    • NEBNext Multiplex Oligos for Illumina® (www.neb.com/oligos)
    • Magnetic rack/stand (NEB #S1515S, Alpaqua® cat. #A001322, or equivalent)
    • Thermal cycler
    • Agilent® Bioanalyzer® or TapeStation® and associated reagents and consumables
    • Adaptor Dilution Buffer NEB #B1430S or NEBNext Unique Dual Index UMI Adaptor Dilution Buffer supplied with NEBNext Unique Dual Index UMI Adaptor DNA Sets (NEB #E7395/E7874/E7876/E7878)

  • 相关产品

    相关产品

    • e6650nebnext-ffpe-dna-library-prep-kit
    • NEBNext® 多样本接头引物试剂盒 1（96 种 Unique 双端 Index 引物）
    • NEBNext® 多样本接头引物试剂盒 2（96 种 Unique 双端 Index 引物）
    • NEBNext® Multiplex Oligos for Illumina® (96 Unique Dual Index Primer Pairs Set 3)

操作说明、说明书 & 用法

  • 操作说明

    1. Where can I find guidelines and protocols for use of the NEBNext UltaShear FFPE DNA Library Prep Kit (NEB #E6655)?

  • 说明书

    产品说明书包含产品使用的详细信息、产品配方和质控分析。

    • manualE6655

工具 & 资源

  • 选择指南

    • NEBNext® Multiplex Oligos Selection Chart

FAQs & 问题解决指南

  • FAQs

    1. What sample types can I use with the NEBNext UltraShear FFPE DNA Library Prep Kit?
    2. How much starting material is required for the NEBNext UltraShear FFPE DNA Library Prep Kit?
    3. When preparing samples using the NEBNext UltraShear FFPE DNA Library Prep Kit, can my input DNA be in EDTA-containing solutions?
    4. Do I really need to vortex the NEBNext UltraShear enzyme mix before use?
    5. What fragmentation time should be used with the NEBNext UltraShear FFPE DNA Library Prep Kit for FFPE DNA samples?
    6. Is size selection recommended?
    7. What are the safe stopping points in this protocol?
    8. Which NEBNext Multiplex Oligos can be used with the NEBNext UltraShear FFPE DNA Library Prep Kit?
    9. What input amount (nanograms) is recommended for standard versus target enrichment library prep with FFPE DNA samples?
    10. Can I use this NEBNext kit with adaptors and primers from vendors other than NEB?
    11. Will treating my DNA with the FFPE DNA Repair Mix v2 as part of the NEBNext UltraShear FFPE DNA Library Prep Kit hurt my downstream reaction?
    12. Does the FFPE DNA Repair v2 Mix repair DNA-protein crosslinks?
    13. Does the FFPE DNA Repair v2 Mix fix blocked 3′ ends?
    14. Can the FFPE DNA Repair v2 Mix repair damage in both single- and double-stranded DNA? Or, does it require double stranded DNA as a template?
    15. If I had a DNA template with mutation sites (i.e. 8-oxoguanine or deaminated cytosines) that are directly adjacent to each other on opposite strands would treatment with the FFPE DNA Repair v2 Mix cause a double strand nick/break?
    16. What gap lengths can be repaired with the FFPE DNA Repair v2 Mix?

Next® FFPE DNA Library Prep Kit |NEB酶试剂 New England Biolabs

发表于由

上海金畔生物科技有限公司代理New England Biolabs（NEB）酶试剂全线产品，欢迎访问官网了解更多产品信息和订购。

产品信息

View or download extensive performance data in our Data Supplement.

FFPE DNA is often compromised in quality and quantity and is a challenging sample type for library prep and sequencing. The NEBNext FFPE DNA Library Prep Kit addresses these challenges in multiple ways:

  • Repairs FFPE-induced damage using the NEBNext FFPE DNA Repair Mix v2
  • Uses NEBNext Ultra II library prep reagents in a protocol optimized for FFPE DNA
  • Robustly amplifies libraries using NEBNext MSTC™ FFPE Master Mix
  • High library yields, sufficient for input into target enrichment workflows
  • User-friendly workflow with minimal hands-on time and multiple safe stopping points

The NEBNext FFPE DNA Library Prep Kit reduces damage from deamination and oxidation due to the fixation, storage, and extraction process for FFPE DNA samples, thereby reducing false positive variant calls.

For further performance improvement, the NEBNext UltraShear FFPE DNA Library Prep Kit (NEB #E6655) also incorporates specialized enzymatic DNA fragmentation.

FIGURE 1: NEBNext FFPE DNA Library Prep Kit workflow

Next® FFPE DNA Library Prep Kit |

The NEBNext FFPE DNA Library Prep kit has a streamlined workflow with minimal hands-on time. The protocol has been optimized for the user to safely store the reaction after any step in the workflow overnight at -20°C without affecting library yield or quality.

FIGURE 2: The NEBNext FFPE DNA Library Prep Kit enables robust library preparation from a range of sample inputs and quality

Next® FFPE DNA Library Prep Kit |

Libraries were prepared from 5, 50 or 250 ng of Covaris®-sheared normal tissue FFPE DNA ranging in quality from DNA Integrity Number (DIN) 1.8 to 6.8 with the indicated PCR cycles. Libraries were made in triplicate for 5 and 50 ng input and 1 replicate for 250 ng. Each bar represents the average of triplicates with error bars indicating standard deviation for the 5 and 50 ng inputs. Robust library yields were obtained across sample qualities and input amounts. Most target enrichment workflows require 200 ng of library for hybrid capture input, and sufficient library yield can be obtained using a minimum of 50 ng FFPE DNA with the NEBNext FFPE DNA Library Prep Kit.

FIGURE 3: NEBNext FFPE DNA Library Prep Kit reduces damage-derived sequencing artifacts

Next® FFPE DNA Library Prep Kit |    

Libraries were prepared from 50 ng of Covaris-sheared normal tissue FFPE DNA of either low (DIN 1.8) or high (DIN 6.8) quality using the NEBNext FFPE DNA Library Prep Kit and other library prep kits as shown. Libraries were sequenced on the Illumina® NextSeq® 500 (2 x 76 bases). Libraries were downsampled to 600,000 paired-end reads, mapped using bowtie2 (version 2.3.2.2) to the GrCh38 reference, and duplicates marked using Picard MarkDuplicate (version 1.56.0). The average frequency of C→T mutations at each C position in Read 2 was calculated for two technical replicates using Tasmanian (version 1.0.7). C→T mutations arising from cytosine deamination damage in low quality FFPE DNA are effectively repaired by the NEBNext FFPE DNA Repair v2 mix included in the NEBNext FFPE DNA Library Prep Kit. Other kits show a high level of C→T artifacts in low quality FFPE DNA (DIN 1.8) due to a lack of DNA damage repair.

FIGURE 4: The NEBNext FFPE DNA Library Prep Kit detects expected variants in formalin-compromised reference standard DNA

Next® FFPE DNA Library Prep Kit |

Libraries were prepared in triplicate from 50, 100, or 250 ng of Covaris-sheared formalin-compromised reference standard DNA (severe) (Horizon Discovery HD803) using either the NEBNext FFPE DNA Library Prep Kit or NEBNext Ultra™ II DNA Library Prep Kit. The full library yield was used in a target enrichment workflow with a custom cancer panel (Twist Bioscience®) as a 6-plex capture reaction. Libraries were sequenced on the Illumina NovaSeq® 6000 (2 x 100 bases). All fastq files were downsampled to 22 million paired-end reads, mapped using BWA mem (version 0.7.17) to the T2T reference, and duplicates marked using Picard MarkDuplicates (version 1.56.0). Variant allele frequencies (VAF) were calculated from Mpileup using Samtools (version 1.16.1) and plotted against the expected VAF in the Horizon reference DNA. A variant with a frequency of 0.88% was missed at 50 ng input for 1/3 replicates, but all variants were detected in all replicates at 100 and 250 ng input. The NEBNext FFPE DNA Library Prep kit improves the correlation of expected to observed VAF in low input libraries (50 ng) compared to standard library prep (NEBNext Ultra II DNA), indicating the benefit of the yield and coverage obtained with the NEBNext FFPE DNA Library Prep Kit including repair with NEBNext FFPE DNA Repair v2.

产品类别:
FFPE DNA Products,
Next Generation Sequencing Library Preparation Products

  • 试剂盒组成

    本产品提供以下试剂或组分：

    NEB # 名称 组分货号 储存温度 数量 浓度
    • E6650S     Multi-temperature    
        NEBNext® FFPE DNA Library Prep Kit E6650-2 -20    
        NEBNext® FFPE DNA Repair Mix v2 E7361AVIAL -20 1 x 0.048 ml Not Applicable
        NEBNext® Thermolabile Proteinase K E7362AVIAL -20 1 x 0.048 ml Not Applicable
        NEBNext® FFPE DNA Repair Buffer v2 E7363AVIAL -20 1 x 0.168 ml Not Applicable
        NEBNext® Ultra II End Prep Enzyme Mix E7646AVIAL -20 1 x 0.072 ml Not Applicable
        NEBNext® Ultra™ II Ligation Master Mix E7648AVIAL -20 1 x 0.72 ml Not Applicable
        NEBNext® Ligation Enhancer E7374AVIAL -20 1 x 0.024 ml Not Applicable
        NEBNext® MSTC FFPE Master Mix E6651AVIAL -20 1 x 0.6 ml 2 X
        NEBNext® Sample Purification Beads E6652S 25    
        NEBNext® Sample Purification Beads E6652AVIAL 25 1 x 4.3 ml Not Applicable
    • E6650L     Multi-temperature    
        NEBNext® FFPE DNA Library Prep Kit E6650-3 -20    
        NEBNext® FFPE DNA Repair Mix v2 E7361AAVIAL -20 1 x 0.192 ml Not Applicable
        NEBNext® Thermolabile Proteinase K E7362AAVIAL -20 1 x 0.192 ml Not Applicable
        NEBNext® FFPE DNA Repair Buffer v2 E7363AAVIAL -20 1 x 0.672 ml Not Applicable
        NEBNext® Ultra II End Prep Enzyme Mix E7646AAVIAL -20 1 x 0.288 ml Not Applicable
        NEBNext® Ultra™ II Ligation Master Mix E7648AAVIAL -20 3 x 0.96 ml Not Applicable
        NEBNext® Ligation Enhancer E7374AAVIAL -20 1 x 0.096 ml Not Applicable
        NEBNext® MSTC FFPE Master Mix E6651AAVIAL -20 2 x 1.2 ml 2 X
        NEBNext® Sample Purification Beads E6652L 25    
        NEBNext® Sample Purification Beads E6652AAVIAL 25 1 x 17 ml Not Applicable

  • 特性和用法

    需要但不提供的材料

    • 80% Ethanol
    • Nuclease-free Water
    • 0.1X TE (1 mM Tris-HCl, pH 8.0, 0.1 mM EDTA)
    • DNase-, RNase-free PCR strip tubes
    • NEBNext Multiplex Oligos for Illumina® (www.neb.com/oligos)
    • Magnetic rack/stand (NEB #S1515S, Alpaqua® cat. #A001322, or equivalent)
    • Thermal cycler
    • Agilent® Bioanalyzer® or TapeStation® and associated reagents and consumables
    • Adaptor Dilution Buffer NEB #B1430S or NEBNext Unique Dual Index UMI Adaptor Dilution Buffer supplied with NEBNext Unique Dual Index UMI Adaptor DNA Sets (NEB #E7395/E7874/E7876/E7878)

  • 相关产品

    相关产品

    • e6655nebnext-ultrashear-ffpe-dna-library-pep-kit
    • NEBNext® 多样本接头引物试剂盒 1（96 种 Unique 双端 Index 引物）
    • NEBNext® 多样本接头引物试剂盒 2（96 种 Unique 双端 Index 引物）
    • NEBNext® Multiplex Oligos for Illumina® (96 Unique Dual Index Primer Pairs Set 3)

操作说明、说明书 & 用法

  • 操作说明

    1. Where can I find guidelines and protocols for use of the NEBNext FFPE DNA Library Prep Kit (NEB #E6650)?

  • 说明书

    产品说明书包含产品使用的详细信息、产品配方和质控分析。

    • manualE6650

工具 & 资源

  • 选择指南

    • NEBNext® Multiplex Oligos Selection Chart

FAQs & 问题解决指南

  • FAQs

    1. What sample types can I use with the NEBNext FFPE DNA Library Prep Kit?
    2. How much starting material is required for the NEBNext FFPE DNA Library Prep Kit?
    3. Is size selection recommended?
    4. What are the safe stopping points in this protocol?
    5. What input amount (nanograms) is recommended for standard versus target enrichment library prep with FFPE DNA samples?
    6. Can I use this NEBNext kit with adaptors and primers from vendors other than NEB?
    7. What do I do if I see a precipitate in the NEBNext FFPE DNA Repair Buffer v2?
    8. Will treating my DNA with the FFPE DNA Repair Mix v2 as part of the NEBNext FFPE DNA Library Prep Kit hurt my downstream reaction?
    9. Which NEBNext Multiplex Oligos can be used with the NEBNext FFPE DNA Library Prep Kit?
    10. Does the FFPE DNA Repair v2 Mix repair DNA-protein crosslinks?
    11. Does the FFPE DNA Repair v2 Mix fix blocked 3′ ends?
    12. Can the FFPE DNA Repair v2 Mix repair damage in both single- and double-stranded DNA? Or, does it require double stranded DNA as a template?
    13. If I had a DNA template with mutation sites (i.e. 8-oxoguanine or deaminated cytosines) that are directly adjacent to each other on opposite strands would treatment with the FFPE DNA Repair v2 Mix cause a double strand nick/break?
    14. What gap lengths can be repaired with the FFPE DNA Repair v2 Mix?

Next® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) |NEB酶试剂 New England Biolabs

发表于由

上海金畔生物科技有限公司代理New England Biolabs（NEB）酶试剂全线产品，欢迎访问官网了解更多产品信息和订购。

产品信息

The ARTIC SARS-CoV-2 protocol from the ARTIC Network is a multiplexed amplicon approach covering the whole viral genome. The NEBNext ARTIC SARS-CoV-2 Library Prep Kit is based on this method and incorporates optimized and novel reagents. 

A single RT protocol is used regardless of input amount (10-10,000 viral genome copies), and no normalization step is required ahead of targeted amplification. The V3 ARTIC primers have been balanced, using methodology developed at NEB based on empirical data from sequencing, to provide uniform amplicon coverage across the viral genome. Library preparation uses the NEBNext Ultra II DNA reagents and workflow, with reagent volumes tailored to the ARTIC application. Note that adaptors and primers for library prep are purchased separately. The novel NEBNext Library PCR Master Mix allows use of the same number of PCR cycles to amplify all libraries, regardless of initial input amounts. 

The NEBNext ARTIC SARS-CoV-2 Library Prep Kit generates libraries with inserts ~400 bp. If shorter inserts are preferred, the NEBNext ARTIC SARS-CoV-2 FS Library Prep Kit (NEB #E7658) produces ~150 bp insert libraries compatible with 2 x 75 sequencing. 

Figure 1: Improved genome coverage with the NEBNext ARTIC SARS-CoV-2 Library Prep Kit (Illumina) 
Next® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) |
Integrative Genome Viewer visualization of read coverage across the SARS-CoV-2 genome. Amplicons were generated from 1,000 copies of SARS-CoV-2 viral gRNA inputs (ATCC VR-1986 and VR-1991) in 100 ng of Universal Human Reference RNA (ThermoFisher® QS0639) using IDT ARTIC nCoV-2019 V3 Panel (“Standard”) or the NEBNext balanced ARTIC SARS-CoV-2 primer pools, with or without NEBNext ARTIC Human Control Primer Pairs. Libraries were constructed using the NEBNext ARTIC SARS-CoV-2 Library Prep Kit (Illumina) and sequenced on a MiSeq® instrument (2×250 bp). Coverage depth per base was determined, reads were down-sampled with seqtk and aligned to SARS-CoV-2 reference genome (NCBI, NC_045512) with Bowtie2.

Figure 2: Improved coverage depth per base with the NEBNext ARTIC SARS-CoV-2 Library Prep Kit (Illumina) 

Next® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) |

Amplicons were generated from 10–10,000 copies of SARS-CoV-2 viral gRNA inputs (ATCC VR-1986 and VR-1991) in 100 ng of Universal Human Reference RNA (ThermoFisher QS0639) using IDT ARTIC nCoV-2019 V3 Panel (“Standard”) or the NEBNext balanced ARTIC SARS-CoV-2 primer pools. Libraries were constructed using the NEBNext ARTIC SARS-CoV-2 Library Prep Kit (Illumina) and sequenced on a MiSeq instrument (2×250 bp). Coverage depth per base was determined, reads were down-sampled with seqtk and aligned to SARS-CoV-2 reference genome (NCBI, NC_045512) with Bowtie2.

Figure 3: Including human control primers does not adversely affect genome coverage
Next® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) |
Amplicons were generated from 1,000 copies of SARS-CoV-2 viral gRNA inputs (ATCC VR-1986 and VR-1991) in 100 ng of Universal Human Reference RNA (ThermoFisher QS0639) using NEBNext balanced ARTIC SARS-CoV-2 primer pools, with or without NEBNext ARTIC Human Control Primer Pairs. Libraries were constructed using the NEBNext ARTIC SARS-CoV-2 Library Prep Kit (Illumina) and sequenced on a MiSeq instrument (2×250 bp). The fraction of the genome covered at each depth was determined for a range of inputs and reads down-sampled to 10,000, 100,000, 500,000 and 1,000,000.

Figure 4: Express and Standard workflows for the NEBNext ARTIC SARS-CoV-2 Library Prep Kit for Illumina

Next® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) |

 
 
产品类别:
NEBNext® ARTIC products for SARS-CoV-2 sequencing,
RNA Library Prep for Illumina,
Next Generation Sequencing Library Preparation Products

  • 试剂盒组成

    本产品提供以下试剂或组分：

    NEB # 名称 组分货号 储存温度 数量 浓度
    • E7650S     Multi-temperature    
        NEBNext® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) E7650-1 -20    
        LunaScript® RT SuperMix E7651AVIAL -20 1 x 0.048 ml 5 X
        Q5® Hot Start High-Fidelity 2X Master Mix E7652AVIAL -20 1 x 0.3 ml 2 X
        NEBNext® Ultra II™ End Prep Enzyme Mix E7653AVIAL -20 1 x 0.036 ml Not Applicable
        NEBNext® Ultra II™ End Prep Buffer E7654AVIAL -20 1 x 0.084 ml Not Applicable
        NEBNext® Ultra II™Ligation Master Mix E7655AVIAL -20 1 x 0.36 ml Not Applicable
        NEBNext® Library PCR Master Mix E7656AVIAL -20 1 x 0.3 ml 2 X
        0.1X TE E7657AVIAL -20 1 x 1.3 ml Not Applicable
        Nuclease-free Water E7667AVIAL -20 1 x 1.5 ml Not Applicable
        NEBNext® ARTIC SARS-CoV-2 Primer Mix 1 E7725AVIAL -20 1 x 0.042 ml Not Applicable
        NEBNext® ARTIC SARS-CoV-2 Primer Mix 2 E7726AVIAL -20 1 x 0.042 ml Not Applicable
        NEBNext® ARTIC Human Control Primer Pairs 1 E7727AVIAL -20 1 x 0.007 ml Not Applicable
        NEBNext® ARTIC Human Control Primer Pairs 2 E7728AVIAL -20 1 x 0.007 ml Not Applicable
        NEBNext® Sample Purification Beads E7659S 25    
        NEBNext® Sample Purification Beads E7659SVIAL 25 1 x 2.1 ml Not Applicable
    • E7650L     Multi-temperature    
        NEBNext® ARTIC SARS-CoV-2 Library Prep Kit (Illumina®) E7650-2 -20    
        LunaScript® RT SuperMix E7651AAVIAL -20 1 x 0.192 ml 5 X
        Q5® Hot Start High-Fidelity 2X Master Mix E7652AAVIAL -20 1 x 1.2 ml 2 X
        NEBNext® Ultra II™ End Prep Enzyme Mix E7653AAVIAL -20 1 x 0.144 ml Not Applicable
        NEBNext® Ultra II™ End Prep Buffer E7654AAVIAL -20 1 x 0.336 ml Not Applicable
        NEBNext® Ultra II™Ligation Master Mix E7655AAVIAL -20 2 x 0.72 ml Not Applicable
        NEBNext® Library PCR Master Mix E7656AAVIAL -20 1 x 1.2 ml 2 X
        0.1X TE E7657AAVIAL -20 1 x 5.2 ml Not Applicable
        Nuclease-free Water E7667AVIAL -20 1 x 1.5 ml Not Applicable
        NEBNext® ARTIC SARS-CoV-2 Primer Mix 1 E7725AAVIAL -20 1 x 0.168 ml Not Applicable
        NEBNext® ARTIC SARS-CoV-2 Primer Mix 2 E7726AAVIAL -20 1 x 0.168 ml Not Applicable
        NEBNext® ARTIC Human Control Primer Pairs 1 E7727AVIAL -20 1 x 0.007 ml Not Applicable
        NEBNext® ARTIC Human Control Primer Pairs 2 E7728AVIAL -20 1 x 0.007 ml Not Applicable
        NEBNext® Sample Purification Beads E7659L 25    
        NEBNext® Sample Purification Beads E7659SVIAL 25 4 x 2.1 ml Not Applicable

  • 特性和用法

    需要但不提供的材料

    • NEBNext Singleplex or Multiplex Oligos for Illumina – www.neb.com/oligos
    • 80% Ethanol (freshly prepared)
    • Nuclease-free Water
    • DNA LoBind Tubes (Eppendorf #022431021)
    • Magnetic rack/stand (NEB #S1515, Alpaqua®, cat. #A001322 or equivalent)
    • Thermocycler
    • Vortex Mixer
    • Microcentrifuge
    • Agilent® Bioanalyzer® or similar fragment analyzer and associated consumables
    • DNase RNase free PCR strip tubes (USA Scientific 1402-1708)

  • 优势和特性

    Features

    • Streamlined, high-efficiency protocol
    • Effective with a wide range of viral genome inputs (10-10,000 copies)
    • Improved uniformity of SARS-CoV-2 genome coverage depth with a more balanced primer pool
    • No requirement for amplicon normalization prior to library preparation
    • Library inserts in the 400 bp range
    • Compatible with 2 x 250 Illumina sequencing
    • Includes NEBNext Sample Purification Beads (SPRIselect®)
    • Adaptors and primers available separately; For a 384 unique dual index format, please contact us

  • 相关产品

    相关产品

    • E7658 NEBNext ARTIC SARS-CoV-2 FS Library Prep Kit Illumina
    • E7660 NEBNext ARTIC SARS-CoV-2 Companion Kit Oxford Nanopore Technologies

  • 参考文献

    1. Josh Quick nCoV-2019 sequencing protocol v2 (GunIt). protocols.io.

操作说明、说明书 & 用法

  • 操作说明

    1. Where can I find guidelines and protocols for using the NEBNext ARTIC SARS-CoV-2 Library Prep Kit, including cDNA synthesis, cDNA amplification and library prep?

  • 说明书

    产品说明书包含产品使用的详细信息、产品配方和质控分析。

    • manualE7650

工具 & 资源

  • Web 工具

    • NEBNext Selector

FAQs & 问题解决指南

  • FAQs

    1. Are the sequences of the primers in the NEBNext ARTIC SARS-CoV-2 Primer Mixes the same as the ARTIC Network V3 primers?
    2. How many sequencing reads do I need for sequencing of libraries generated using the  NEBNext ARTIC Kits for Illumina?
    3. What Ct values are recommended for inputs used with the NEBNext ARTIC kits for Illumina?
    4. How do I analyze data from libraries generated using the NEBNext ARTIC kits for Illumina?
    5. Which NEBNext adaptors and primers are recommended for use with the ARTIC kits for Illumina?
    6. Do I need to add PhiX to the sequencing run?
    7. Do I need to normalize library concentration prior to sequencing?
    8. What read length is recommended for sequencing of libraries generated using the  NEBNext ARTIC SARS-CoV-2 Kit (Illumina)?
    9. What should the final loading concentration be for MiSeq® and NextSeq® 500/550 Illumina® sequencing platforms?
    10. Does my total RNA sample need to be DNA-free prior to starting the ARTIC workflow?