Next UltraExpress™ RNA 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 below and in our Data Supplement
 
This new kit meets the need for substantially faster, more streamlined RNA library prep workflows that deliver high-quality directional libraries from a variety of sample types in a single day from total RNA. 

The kit allows the use of a single protocol for all RNA inputs (25 -250 ng total RNA) and incorporates master mixed reagents, reduced incubation times and fewer cleanup steps, reducing total time and consumables used.  

  • 3-hour library prep enables single day total RNA to library, in conjunction with mRNA enrichment or rRNA depletion kits
  • Fewer cleanups and pipetting steps reduce hands-on time
  • A robust single protocol for all input amounts increases ease of use
  • Flexibility is enabled with simple guidelines for customized protocols, if desired
  • Automation-friendly, enabling scalability
Figure 1: NEBNext UltraExpress™RNA Library Prep Kit workflow

Next UltraExpress™ RNA Library Prep Kit |

Figure 2: The NEBNext UltraExpress™ RNA Library Prep Kit produces high library yields across a range of inputs

Next UltraExpress™ RNA Library Prep Kit |

A. Poly(A)-containing mRNA was isolated from Universal Human Reference RNA (UHRR) (Agilent®), using the NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB #E7490). Libraries were prepared using the NEBNext UltraExpress™ RNA Library Prep Kit, Kapa mRNA HyperPrep® Kit or Illumina® Stranded mRNA Kit. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 12 PCR cycles for all inputs, or with customized adaptor dilutions (20X for 76–250ng, 100X for 25–75 ng) and PCR cycle numbers (listed in figure).

B. Ribosomal RNA (rRNA) was depleted from UHRR, and libraries were prepared using the UltraExpress RNA Library Prep Kit (preceded by the NEBNext rRNA Depletion Kit (Human/Mouse/Rat – NEB #E7400), Kapa HyperPrep Kit with RiboErase, or Illumina Stranded Total RNA Library Prep Kit with Ribo-Zero® Plus. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 11 PCR cycles for all inputs, or with customized adaptor dilutions (20X for 76–250 ng, 100X for 25–75 ng) and PCR cycle numbers (listed in figure).

The total RNA input amount and number of PCR cycles are indicated. Library yields calculated from an average of three replicates are shown with error bars indicating the standard deviation between replicates.    

Figure 3: The NEBNext UltraExpress™ RNA Library Prep Kit provides consistent transcript coverage

Next UltraExpress™ RNA Library Prep Kit |

A. Poly(A)-containing mRNA was isolated from Universal Human Reference RNA (UHRR) (Agilent®), using the NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB #E7490). Libraries were prepared using the NEBNext UltraExpress™ RNA Library Prep Kit, Kapa mRNA HyperPrep® Kit or Illumina® Stranded mRNA Kit. Poly(A)-containing mRNA was isolated from Universal Human Reference RNA (UHRR) (Agilent), and libraries were made using the UltraExpress RNA Library Prep Kit, Kapa mRNA HyperPrep Kit and Illumina Stranded mRNA Kit. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 12 PCR cycles for all inputs, or with customized adaptor dilutions and PCR cycle numbers (20X adaptor dilution with 10 PCR cycles for 250 ng input, 100X adaptor dilution with 12 PCR cycles (75 ng) and 14 PCR cycles (25 ng)).

B. Ribosomal RNA (rRNA) was depleted from UHRR, and libraries were prepared using the UltraExpress RNA Library Prep Kit (preceded by the NEBNext rRNA Depletion Kit (Human/Mouse/Rat – NEB #E7400), KAPA HyperPrep Kit with RiboErase, or Illumina Stranded Total RNA Library Prep Kit with Ribo-Zero® Plus. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 12 PCR cycles for all inputs, or with customized adaptor dilutions and PCR cycles (20X adaptor dilution with 9 PCR cycles for 250 ng input, 100X adaptor dilution with 11 PCR cycles (75 ng) and 13 PCR cycles (25 ng)).

Libraries were sequenced on an Illumina NextSeq® 500 (2 x 75 bases). 10 million reads were sampled and mapped to the hg38 reference genome using RNA STAR v2.7.8a and 5´ to 3´ Transcript coverage was calculated from the top 1,000 transcripts using the CollectRnaSeqMetrics (Picard) tool v2.18.2.2.

Figure 4: The NEBNext UltraExpress™ RNA Library Prep Kit provides greater sensitivity of transcript detection

Next UltraExpress™ RNA Library Prep Kit |

A. Poly(A)-containing mRNA was isolated from Universal Human Reference RNA (UHRR) (Agilent®), using the NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB #E7490). Libraries were prepared using the NEBNext UltraExpress™ RNA Library Prep Kit, Kapa mRNA HyperPrep® Kit or Illumina® Stranded mRNA Kit. Poly(A)-containing mRNA was isolated from Universal Human Reference RNA (UHRR) (Agilent), and libraries were made using the NEBNext UltraExpress RNA Library Prep Kit, Kapa mRNA HyperPrep Kit and Illumina Stranded mRNA Kit. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 12 PCR cycles for all inputs, or with customized adaptor dilutions and PCR cycle numbers (20X adaptor dilution with 10 PCR cycles for 250 ng input, 100X adaptor dilution with 12 PCR cycles (75 ng) and 14 PCR cycles (25 ng)).

B. Ribosomal RNA (rRNA) was depleted from UHRR, and libraries were prepared using the UltraExpress RNA Library Prep Kit (preceded by the NEBNext rRNA Depletion Kit (Human/Mouse/Rat – NEB #E7400), Kapa HyperPrep Kit with RiboErase, or Illumina Stranded Total RNA Library Prep Kit with Ribo-Zero® Plus. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 11 PCR cycles for all inputs, or with customized adaptor dilutions and PCR cycle numbers (20X adaptor dilution with 9 PCR cycles for 250 ng input, 100X adaptor dilution with 11 PCR cycles (75 ng) and 13 PCR cycles (25 ng)).

Libraries were sequenced on an Illumina NextSeq® 500 (2 x 75 bases). 10 million reads were sampled and average transcript abundance was assessed for transcripts detected with ≤ 10 reads and ≤ 500 reads, respectively. Error bars indicate standard deviation for 3 replicates. Salmon v1.5.1 was used for mapping and quantification of all gencode v38 transcripts and ERCCs.

Figure 5: The NEBNext UltraExpress™ RNA Library Prep Kit provides excellent transcript correlation between inputs and replicates
 
Next UltraExpress™ RNA Library Prep Kit |
A. Poly(A)-containing mRNA was isolated from Universal Human Reference RNA (UHRR) (Agilent®), using the NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB #E7490). Libraries were prepared using the NEBNext UltraExpress RNA Library Prep with a single adaptor dilution (50X) and 12 PCR cycles for all inputs, or with customized adaptor dilutions and PCR cycle numbers (20X adaptor dilution with 10 PCR cycles for 250 ng input, 100X adaptor dilution with 12 PCR cycles (75 ng) and 14 PCR cycles (25 ng)).

B. Ribosomal RNA (rRNA) was depleted from UHRR using the NEBNext rRNA Depletion Kit v2 (Human/Mouse/Rat – NEB #E7400) and libraries. The NEBNext UltraExpress RNA Library Prep Kit was used with a single adaptor dilution (50X) and 11 PCR cycles for all inputs, or with customized adaptor dilutions and PCR cycle numbers (20X adaptor dilution with 9 PCR cycles for 250 ng input, 100X adaptor dilution with 11 PCR cycles (75 ng) and 13 PCR cycles (25 ng)).

Libraries were sequenced on an Illumina NextSeq® 500 (2 x 75 bases). The data was sampled from 10 million reads and libraries were correlated for transcript expression levels across inputs using Salmon v1.5.1 quantification of all gencode v38 transcripts and ERCCs. Each data point represents a transcript, with the log10 abundance in Number of Reads with 250 ng total RNA input on the y-axis compared to a replicate at 250 ng followed by 75 and 25 ng on the x-axis.

Figure 6: The NEBNext UltraExpress™ RNA Library Prep Kit provides excellent transcript correlation across sample types

Next UltraExpress™ RNA Library Prep Kit |

RNA from human whole blood (HWB) was depleted of globin mRNA and rRNA (A) using the NEBNext® Globin & rRNA Depletion Kit (Human/Mouse/Rat – NEB #E7750) or rRNA was depleted from a pool of 20 different bacterial organisms (ATCC® #MSA-2002) (B) using the NEBNext rRNA Depletion Kit (Bacteria – NEB #E7850). Libraries were prepared using the NEBNext UltraExpress™ RNA Library Prep Kit, with customized adaptor dilution and PCR cycles per input (20X adaptor dilution with 9 PCR cycles for 250 ng input, 100X adaptor dilution with 11 PCR cycles (75 ng) and 13 PCR cycles (25 ng)). Libraries were sequenced on an Illumina® NextSeq® 500 (2 x 75 bases). The data was sampled from 10 million reads and libraries were correlated for transcript expression levels across inputs using Salmon v1.5.1 quantification of all gencode v38 transcripts (HWB) and a composite genome (Bacterial). Each data point represents a transcript, with the log10 abundance in number of Reads with 250 ng total RNA input on the y-axis compared to a replicate at 250 ng followed by 75 and 25 ng on the x-axis.
产品类别:
Library Preparation for Illumina Products,
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
    • E3330S     -20    
        NEBNext UltraExpress™ RNA Fragmentation Mix E3329AVIAL -20 1 x 0.096 ml 2 X
        NEBNext UltraExpress™ First Strand Enzyme Mix E3331AVIAL -20 1 x 0.024 ml Not Applicable
        NEBNext UltraExpress™ Strand Specificity Reagent E3332AVIAL -20 1 x 0.096 ml Not Applicable
        NEBNext UltraExpress™ Second Strand Master Mix E3333AVIAL -20 1 x 0.72 ml Not Applicable
        NEBNext UltraExpress™ End Prep Enzyme Mix E3334AVIAL -20 1 x 0.036 ml Not Applicable
        NEBNext UltraExpress™ End Prep Reaction Buffer E3335AVIAL -20 1 x 0.06 ml Not Applicable
        NEBNext UltraExpress™ Ligation Master Mix E3336AVIAL -20 1 x 0.288 ml Not Applicable
        NEBNext UltraExpress™ USER Enzyme E3337AVIAL -20 1 x 0.048 ml Not Applicable
        NEBNext® MSTC™ High Yield Master Mix E3338AVIAL -20 1 x 1.2 ml Not Applicable
        NEBNext® Adaptor Dilution Buffer E7762AVIAL -20 1 x 2.4 ml Not Applicable
        0.1X TE E3341AVIAL -20 1 x 2.4 ml Not Applicable
        Nuclease-free Water E3342AVIAL -20 1 x 0.096 ml Not Applicable
        NEBNext® Bead Reconstitution Buffer E3339AVIAL -20 1 x 0.096 ml Not Applicable
    • E3330L     -20    
        NEBNext UltraExpress™ RNA Fragmentation Mix E3329AAVIAL -20 1 x 0.384 ml 2 X
        NEBNext UltraExpress™ First Strand Enzyme Mix E3331AAVIAL -20 1 x 0.096 ml Not Applicable
        NEBNext UltraExpress™ Strand Specificity Reagent E3332AAVIAL -20 1 x 0.384 ml Not Applicable
        NEBNext UltraExpress™ Second Strand Master Mix E3333AAVIAL -20 3 x 0.96 ml Not Applicable
        NEBNext UltraExpress™ End Prep Enzyme Mix E3334AAVIAL -20 1 x 0.144 ml Not Applicable
        NEBNext UltraExpress™ End Prep Reaction Buffer E3335AAVIAL -20 1 x 0.24 ml Not Applicable
        NEBNext UltraExpress™ Ligation Master Mix E3336AAVIAL -20 1 x 1.16 ml Not Applicable
        NEBNext UltraExpress™ USER Enzyme E3337AAVIAL -20 1 x 0.192 ml Not Applicable
        NEBNext® MSTC™ High Yield Master Mix E3338AAVIAL -20 1 x 4.8 ml Not Applicable
        NEBNext® Adaptor Dilution Buffer E7762AAVIAL -20 1 x 9.6 ml Not Applicable
        0.1X TE E3341AAVIAL -20 1 x 9.6 ml Not Applicable
        Nuclease-free Water E3342AAVIAL -20 1 x 0.384 ml Not Applicable
        NEBNext® Bead Reconstitution Buffer E3339AAVIAL -20 1 x 3.84 ml Not Applicable

  • 特性和用法

    需要但不提供的材料

    • NEBNext Multiplex Oligos for Illumina®
    • NEBNext Multiplex Oligos options can be found at www.neb.com/oligos. Alternatively, customer supplied adaptor and primers can be used; please see information in link: https://www.neb.com/faqs/2019/03/08/can-i-use-this-nebnext-kit-with-adaptors-and-primers-from-other-vendors-than-neb
    • SPRIselect™ Reagent Kit (Beckman Coulter®, Inc. #B23317) or AMPure® XP Beads (Beckman Coulter, Inc. #A63881)
    • Magnetic Rack (NEB S1515S, Alpaqua® cat. #A001322, or equivalent)
    • 80% Ethanol (freshly prepared)
    • Thermal cycler
    • DNase-, RNase-free PCR strip tubes, for example TempAssure® PCR flex-free 8-tube strips (USA Scientific® #1402-4708)
    • Bioanalyzer® or TapeStation® (Agilent® Technologies, Inc.) and associated reagents and consumables

     

    For use with NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB #E7490):

    • 1.5 ml Microcentrifuge tube and NEB #S1506 Magnet stand or equivalent (for washing beads only)

     

    For use with NEBNext rRNA Depletion Kit v2 (Human/Mouse/Rat) (NEB #E7400) and other NEBNext RNA depletion kits that do not include beads (NEB #E7750, E7850, E7865):

    • Agencourt® RNAClean® XP Beads (Beckman Coulter, Inc. #A63987)

  • 相关产品

    相关产品

    • e7490-nebnext-polya-mrna-magnetic-isolation-module
    • E7400 NEBNext rRNA Depletion Kit v2 Human Mouse Rat
    • NEBNext® 多样本接头引物试剂盒 5（96 种 Unique 双端 Index 引物）
    • NEBNext® 文库定量试剂盒（Illumina®）
    • NEBNext® 磁性分离架
    • T2030 Monarch RNA Cleanup Kit 10 ug
    • Monarch® 总 RNA 小量提取试剂盒

操作说明、说明书 & 用法

  • 操作说明

    1. Where can I find protocols for using the NEBNext UltraExpress™ RNA Library Prep Kit (NEB #E3330)?

  • 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。

    • manualE3330

工具 & 资源

  • Web 工具

    • NEBNext Selector

FAQs & 问题解决指南

  • FAQs

    1. Is the NEBNext UltraExpress™ RNA library prep kit a directional (stranded) library kit?
    2. What is the difference between the NEBNext UltraExpress™ RNA Library Prep Kit and the NEBNext® Ultra™ II Directional RNA Library Prep Kit? How do I choose the right one?
    3. Is it possible to use total RNA input amounts outside the suggested range for the NEBNext UltraExpress™ RNA library Kit?
    4. Does the NEBNext UltraExpress™ RNA Library Prep Kit include poly(A) mRNA isolation or rRNA Depletion reagents?
    5. Do the NEBNext UltraExpress™ Library Prep kits come with beads?
    6. Can I perform poly(A) mRNA isolation or rRNA depletion and then freeze my samples at -80°C before beginning library preparation with the NEBNext UltraExpress™ RNA Library Prep Kit?
    7. Which NEBNext® Multiplex Oligos (Adaptors & Primers) can be used with the NEBNext UltraExpress Library Prep Kits?
    8. What should I do if I see adaptor dimer in my NEBNext UltraExpress™ libraries?
    9. Can this kit be used with adaptors and primers from suppliers other than NEB?
    10. Are the USER enzyme included in the NEBNext UltraExpress™ RNA Library Prep Kit (blue cap) and the USER enzyme included with NEBNext Index Primer sets (red cap) the same?
    11. What sample types and quality are compatible with the NEBNext UltraExpress™ RNA Library Prep Kit?
    12. What do I do if I see a precipitate in the Ultra II End Prep Reaction Buffer?