GenNext™ NGS Library Prep Kit comprises the enzymes and buffers for preparing libraries for illumina® sequencing from fragmented double-stranded DNA and PCR products. With this system, it is possible to conveniently and quickly convert a broad range (1ng - 1μg) of input amounts of DNA into libraries for illumina® sequencing. Terminal repair and 3' end adenylation of the fragmented DNA can be conducted in the end repair and A-tailing step. Platform-specific adapters are then ligated to both ends of the DNA fragments. If required, a high-fidelity amplification step can be performed using the reagents included in the GenNext™ NGS Library Prep Kit. Library Amplification Master Mix uses a highly-accurate PCR enzyme developed using genetically-modified KOD DNA polymerase. This minimizes the influence of GC bias on amplification and can amplify various regions evenly.
- Simple and quick operation flow
The steps from terminal repair and 3' end adenylation to adapter ligation can be conducted in the same container. Terminal repair and adenylation at the 3' end can be performed in 15 minutes. Adapter ligation can be done in 15 minutes. Library amplification can be performed in cycles of 10 seconds’ annealing and 15 seconds’ extension.
- A wide range of input amount
GenNext™ NGS Library Prep Kit is compatible with various inputs from 1 ng to 1 μg.
- Low bias library amplification
Library Amplification Master Mix uses a highly-accurate PCR enzyme developed using genetically-modified KOD DNA polymerase. It minimizes the influence on GC-bias-induced amplification, and it is possible to amplify various regions evenly.
Store at-20 ℃
The kits include the following reagents, which can be used for 8 (LPK-101T), 24 (LPK-101) and 96 (LPK-101L) reactions. All reagents should be stored at -20°C.
|GenNext™ NGS Library Prep Kit||LPK-101T||LPK-101||LPK-101L|
|Size||8 Rxns||24 Rxns||96 Rxns|
|End Repair and A-tailing Buffer*||80 μL||240 μL||960 μL|
|End Repair and A-tailing Enzyme*||20 μL||60 μL||240 μL|
|Ligation Solution||400 μL||1,200 μL||1,600 μl × 3|
|Library Amplification Master Mix||230 μL||690 μL||1,380 μl × 2|
|Library Amplification Primer Mix||46 μL||138 μL||552 μL|
*Do not store mixed solution.
Example 1.Comparison data of library quantification and quality control
Libraries were prepared from fragmented human genomic DNA, E. coli genomic DNA, or 100 bp DNA ladder with the GenNext™ NGS Library Prep Kit or another company’s Kit (Company A). Libraries were amplified using 0-11 cycles of PCR and the size distribution checked using a MultiNA (Shimadzu Corporation). Library quantifications were performed using a GenNext™ NGS Library Quantification Kit (Code No. NLQ-101). There was no difference in the distribution of libraries between the GenNext™ NGS Library Prep Kit and the other company’s kit (Company A). For most illumina® sequencing platforms, 2 - 4 nM for each library is the preferred starting concentration for denaturation and dilution guidelines. These data illustrate that the GenNext™ NGS Library Prep Kit achieved sufficient adapter-ligated library yields, even with low input amounts of DNA.
Example 2.Comparison data of next generation sequencing results
Libraries were prepared from 1μg or 1ng of E. coli genomic DNA with the GenNext™ NGS library prep kit or another company’s library construction kit (Company A). Libraries prepared from 1ng DNA were amplified using 12 PCR cycles. Sequencing was performed on an illumina® MiSeq® and data analyzed using CLC Genomics Workbench (QIAGEN / CLC bio). Sequencing reads were down-sampled to 1 million per library prior to analysis. These data illustrate that GenNext™ NGS library prep kit enables high quality sequence data.