Store at -20 °C.
Guaranteed for 12 months at -20°C in a constant temperature freezer.
ThruPLEX DNA-seq 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
||Input sample amount (50 – 500 bp ds DNA)
||Recommended input amount
|WGS, WES, SNV, indels, SNP, STR
||10 – 50 ng
||20 – 50 ng
||1 – 50 ng
||plasma, gDNA or FFPE DNA
||50 pg – 50 ng*
||50 pg – 50 ng*
||50 pg – 50 ng*
*Use of less than 50 pg has been cited in scientific publications.
Click here to read all FAQ’s on ThruPLEX DNA-seq Kit
1. What are the major differences between ThruPLEX®-FD Prep Kit and the new ThruPLEX DNA-seq Kit?
- Increased multiplexing capability from 12 6nt single indexes to up to 96 8nt dual indexes (Illumina TruSeq HT) or up to 48 8nt single indexes (Sanger, see Appendix 1 of the ThruPLEX DNA-seq Kit Instruction Manual)
- Total reaction volume lowered from 75 µL to 50 µL to accommodate more thermal cyclers
- Protocol modified to account for reduced volume
- Fewer PCR amplification cycles during Stage 5 of the Library Amplification Step
- Indexing reagents for kits containing 48 or 96 indexes are pre-dispensed into a 96-well plate
2. Is ThruPLEX DNA-seq Kit compatible with any target enrichment systems?
Yes, ThruPLEX DNA-seq Kit is compatible with the major exome and target enrichment products, including Agilent SureSelect®, Roche NimbleGen® SeqCap® EZ and custom panels. ThruPLEX DNA-seq Kit target enrichment protocols and application notes can be accessed at: http://rubicongenomics.com/applications/enrichment/.
3. What DNA fragment size do you recommend?
ThruPLEX DNA-seq Kit can accommodate a range of 50 – 1,000 bp DNA input. Fragment DNA to the size specified for the Illumina platform you are using, generally between 200 and 500 bp.
4. What is the recommended elution/resuspension buffer for input DNA sample(s)? Are there any buffer concentration considerations?
The DNA sample(s) should be eluted/resuspended in a low-salt and low-EDTA, buffered solution. The preferred buffer is TE with ≤ 10 mM Tris and ≤ 0.1 mM EDTA. Avoid phosphate containing buffers.
5. Do I need to perform cleanup on my DNA sample(s) after shearing?
No, simply proceed to the Template Preparation Step (Section D.I.) in the ThruPLEX DNA-seq Kit Instruction Manual in order to preserve your total DNA and the overall diversity of the library.
6. What types of DNA samples can be used with ThruPLEX DNA-seq Kit?
- Fragmented, double-stranded DNA, such as that from:
- Mechanically-sheared DNA
- Enzymatically-fragmented DNA
- Low molecular weight cell-free DNA from plasma, urine, and other biofluids
- Double-stranded cDNA
- Chromatin Immunoprecipitated (ChIP) DNA
- Sonicated DNA from formalin-fixed material (FFPE)
7. Can RNA be used with ThruPLEX DNA-seq Kit?
Yes, but RNA needs to be converted into double-stranded cDNA and fragmented to proper size prior to preparation with ThruPLEX DNA-seq Kit.
8. Can plasma, urine, or other biofluids be directly used with ThruPLEX DNA-seq Kit?
No, the template DNA must be first extracted with a DNA extraction/purification kit. Since DNA from these sources is normally degraded/fragmented, additional fragmentation is unnecessary. The volume of the sample used in the
ThruPLEX DNA-seq Kit must be no more than 10 µL.
9. Can I prepare samples for both single- and paired-end NGS sequencing?
Yes, ThruPLEX DNA-seq Kit is compatible with both single- and paired-end sequencing. The final amplified libraries will contain Illumina-compatible index sequences.
10. Is there a required DNA sample input volume for ThruPLEX DNA-seq Kit?
Yes, the required DNA sample volume is 10 µL.
ThruPLEX DNA-seq contains everything needed, including indexes in a barcoded 96-well plate.
CAT. NO. R400406 ThruPLEX DNA-seq 48D Kit (48 reactions, 48 dual indexes)
CAT. NO. R400407 ThruPLEX DNA-seq 96D Kit (96 reactions, 96 dual indexes)
CAT. NO. R400427 ThruPLEX DNA-seq 48S Kit (48 reactions, 48 single indexes)
CAT. NO. R400428 ThruPLEX DNA-seq 12S (48) Kit (48 reactions, 12 single indexes)
CAT. NO. R400429 ThruPLEX DNA-seq 12S Kit (12 reactions, 12 single indexes)