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英文名称： ThruPLEX-FD kit for Illumina NGS Platforms
型号:null    产品货号： R40048
价格:请致电：010-57128832,18610462672
品牌: 美国

Overview

The ThruPLEX-FD Prep Kit* represents a breakthrough in library preparation that streamlines the Illumina Next Generation Sequencing protocol starting with fragmented DNA or cDNA. ThruPlex employs patented technology* based on stem loop adaptors and couples this with a high fidelity polymerase to deliver a highly efficient and user-friendly process. The entire reaction takes place in a single tube in less than 2 hours and eliminates purifications/clean-up steps. Efficiencies, sensitivities and reaction rates are all improved. Regardless of the NGS application– DNA-seq, RNA-seq, ChiP–seq-the ThruPLEX-FD Prep Kit provides consistent and reliable results.

*Protected by US patent 7,803,550 and EP1924704

Workflow

thruplex_workflow_illumina_tube

Learn more about ThruPLEX technology.

Shortest and Easiest Workflow

ThruPLEX-FD Prep Kits simple one-tube workflow requires less hands-on and total time than any kit on the market.

Library Diversity & Sensitivity

Covaris-fragmented human DNA was prepared and sequenced by a university core lab using an NEBNext prep and ThruPLEX-FD prep. Library complexity as measured by DNAnexus “bottleneck score” or diversity calculations shows ThruPLEX has ~10X higher diversity (at 10 ng input) or 30X higher sensitivity (at 10 ng diversity). These metrics can be evaluated at <300K reads.

Uniform Coverage Over a Wide Dynamic Range of Input DNA

Data at low resolution across the entire human chr 1 demonstrates the difference in coverage with different products and input amounts. ThruPlex preps performed well with fragmented DNA at 0.2 ng and 20 ng. This metric can be qualitatively evaluated with less than 300K reads. (Data courtesy of Washington University and the Broad Institute)

Uniform Coverage in CpG Islands

When compared to the Illumina PCR-free prep, the ThruPLEX-FD prep has strong representation across 4 RASSF1 CpG islands. The TruSeq prep shows significant systematic dips in representation across the same CpG island. Note the difference in input DNA levels. All data was analyzed at low coverage.

 
Shortest and Easiest Workflow
Library Diversity & Sensitivity
Uniform Coverage Over a Wide Dynamic Range of Input DNA
Uniform Coverage in CpG Islands
Low PCR Background
Low Resolution Coverage is Insensitive to Number of PCR Cycles
Library Diversity & Unmapped Reads are Insensitive to PCR Cycles
GC Representation is Insensitive to PCR Cycles
ChIP Results Show Concordance with TruSeq Standard Prep
Workflow Advantage
 
 

Store at -20 °C. Guaranteed for 9 months at -20◦C in a constant temperature freezer.

ThruPLEX-FD Prep Kit is designed to work on a wide variety of DNA samples with amounts varying between 50 pg and 50 ng.  To ensure that your experiment delivers quality data, the table below provides a range of recommended input amounts of  human DNA based on (1) your application and (2) the source material.  The actual amount that you chose to use may vary from those suggested below.  Note that as the amount of input DNA decreases, the diversity of the library may also decrease

Application Input sample amount (50 – 500 bp ds DNA) Recommended input amount
WGS, WES, SNV, indels, SNP, STR gDNA 10 – 50 ng
FFPE DNA 20 – 50 ng
plasma DNA 1 – 50 ng
CNV plasma, gDNA or FFPE DNA 50 pg – 50 ng*
ChIP-seq ChIP DNA 50 pg – 50 ng*
RNA-seq cDNA 50 pg – 50 ng*
     

*Use of  less than 50 pg has been cited in scientific publications.

FAQs

Read all FAQ’s on ThruPLEX-FD Prep Kit

1. Is ThruPLEX®-FD Prep Kit compatible with SureSelect (Agilent Technologies) enrichment?
Yes, it is compatible with SureSelect enrichment, although ThruPLEX-FD libraries must first be amplified before undergoing the enrichment. Also, it might be necessary to optimize the amount of amplified ThruPLEX-FD library to be used as input to SureSelect, and/or optimize the concentration of bait.

2. Can components of the ThruPLEX-FD Kit be used in the ThruPLEX DNA-seq Kit?

NO! The kits have different volume requirements and are NOT interchangeable. Please refer to each kit’s respective manual for detailed instructions.

3. Are the barcodes in the ThruPLEX-FD kit and  ThruPLEX DNA-seq interchangable?

NO! The barcodes for the ThruPLEX DNA-seq kit were redesigned specifically for that kit; the ThruPLEX-FD barcodes will perform only with the ThruPLEX-FD Kit.

4. Can ThruPLEX-FD Kit be run without a real time PCR machine? If so, how do I know when to stop my run?
Yes, the kit can be used even if a lab does not have access to a real time PCR machine. Use the table on page 6 of the ThruPLEX-FD Prep Kit manual as an approximation guide to the number of PCR cycles that should be run for one’s particular DNA input amount.

5. What DNA fragmentation size do you recommend?
Average fragment size of 300 bp is recommended; although this kit can accommodate a range of 50 – 1000 bp DNA input. Additionally, depending on the desired sequencing read length, the DNA needs to be fragmented to a specific average size.

6. An unexpected fragment is seen in the Bioanalyzer.  What is the cause?
Unexpected fragment sizes post library prep (fragments were expected to be larger, but the end-result seemed to be as if only 60 bp were added onto their fragments and not the expected 120 bp), may be a result of incorrrect use of thermal cyclers  during the library prep.  In instruments that do not  support the required 75-µl volume, the reaction is not cycled properly. Ensure that the instrumentation used during the library prep  can support 75-µl volume reactions.  Monitoring amplification in real time will also ensure proper library preparation and amplification.

7. Is a high-fidelity enzyme used in the amplification reaction?
Yes, a high-fidelity, high-processivity, low-bias DNA polymerase is used in the amplification reaction.

8. What is the recommended elution or resuspension buffer for DNA sample(s) that will be used in Template Preparation step (step A.1 of the ThruPLEX-FD Prep Kit manual)? Are there any DNA purity considerations?
The DNA sample(s) used  must be eluted or resuspened in a low-salt, low-EDTA, buffered solution, such as TE buffer, or TE with reduced EDTA content.  Water is also acceptable.

9. Is a DNA sample cleanup after shearing recommended?
No. In order to preserve your total DNA and the overall diversity of the library, remove a 10 μl aliquot of the sheared DNA sample and proceed directly to the  Template Preparation step (Step A.1 of the ThruPLEX-FD Prep Kit manual).

10. What is the recommended fluorescent dye for real time PCR monitoring?
EvaGreen® fluorescent dye (Biotium, Catalog No: 31000) is recommended, due to its high sensitivity and low interference with the amplification chemistry.

 

 

ThruPLEX®-FD Prep Kit (48 rxn, 12 indexes for Illumina® HiSeq/GA/MiSeq)   CAT. NO. R40048.

ThruPLEX®-FD Prep Kit (12 rxn, 12 indexes for Illumina® HiSeq/GA/MiSeq)    CAT. NO. R40012

Each kit has everything required for library preparation when starting with fragmented DNA or cDNA including 12 Illumina indices.