ThruPLEX®-FD Prep Kit

ThruPLEX-FD Prep Kit is no longer available.

 

 ThruPLEX DNA-seq Kit now offers better performance and more indexing options.

 

Contact our sales team for more information.

 

Learn more about ThruPLEX technology.

For a complete list of all scientific publications, use our Resource Search.
ThruPLEX-FD Kit product sheet
ThruPLEX-FD Prep Kit Instruction Manual
ThruPLEX-FD Prep Kit Quick Protocol
MSDS ThruPLEX-FD Kit
ThruPLEX-FD Kit FAQ’s

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 is no longer available.

For RNA-seq work, we recommend the Clontech SMARTer®  Universal Low Input RNA Kit.

Resources

For a complete list of all scientific publications, use our Resource Search.
ThruPLEX-FD Kit product sheet
ThruPLEX-FD Prep Kit Instruction Manual
ThruPLEX-FD Prep Kit Quick Protocol
MSDS ThruPLEX-FD Kit
ThruPLEX-FD Kit FAQ’s

FAQs

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.

 

 

Ordering Information

ThruPLEX®-FD Prep Kit is no longer available.

For RNA-seq work, we recommend the Clontech SMARTer®  Universal Low Input RNA Kit.