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Encore® Rapid Library Systems
Note: Encore Rapid Library Systems replace Encore NGS Library System I (PN 0300-08), Encore NGS Multiplex System I (PN 0301-32) and Encore NGS Multiplex System IB (0302-32)
The Encore Rapid Library Systems provide a simple, fast and scalable solution for producing libraries used in next-generation sequencing. These systems enable library construction starting with as little as 100 ng of double-stranded DNA, without PCR amplification. The library construction workflow is suitable for a wide range of sequencing applications including RNA-Seq, Digital Gene Expression (DGE), genomic DNA sequencing, amplicon sequencing and more.
As shown in Figure 1, the streamlined workflow consists of four main steps: fragmentation of either genomic DNA or double-stranded cDNA, end repair to generate blunt ends, adaptor ligation for multiplexing or no multiplexing and final repair to produce the library. Unlike libraries used in many other NGS protocols, the library workflow does not require PCR amplification. The entire workflow requires only one bead purification step and no gel purification. The protocol can be completed in fewer than two hours and yields libraries ready for cluster formation and either single read or paired-end sequencing.
The Encore Rapid Library Systems are designed for seamless integration with NuGEN's Ovation® RNA-Seq System V2 and Ovation RNA-Seq FFPE System (Part Nos. 7102 and 7150, respectively). Importantly, for DNA sequencing applications, low abundance samples (100 ng or higher) can be input directly to the library construction workflow without pre-amplification. The absence of PCR steps in the library construction workflow makes this method particularly well suited to either high or low GC genomes that may be subject to PCR artifacts (Figure 2).
The Encore Rapid Library System (Part No. 0316) contains reagents for production of non-barcoded libraries, while the Encore Rapid IL Multiplex System 1-8 (Part No. 0317) and IL Multiplex System 9-16 (Part No. 0318) each provide eight unique inline barcoded adaptors for multiplex sequencing. The Encore Rapid DR Multiplex System 1-8 (Part No. 0319) and DR Multiplex System 9-16 (Part No. 0320) provide eight unique barcodes using a dedicated read (DR) design with a second sequencing primer. In combination, each of these multiplex kit pairs enables up to 16-plex sequencing using either inline or dedicated read barcode designs. Lastly, for higher levels of multiplexing the Encore Rapid DR Multiplex System 1-96 (Part No. 0328) provides 96 independent DR barcoded adaptors.
Figure 1. The Encore Rapid Library System Workflow
Figure 2. Sequencing Coverage with High and Low GC Content
Sequencing libraries were constructed using the Encore Rapid IL Multiplex System 1-8 (Part No. 0317) starting with either 100 ng or 500 ng of genomic DNA obtained from S. aureus (low GC - 32.8%) or R. sphaeroides (high GC - 68.5%). Sequencing was performed on the Illumina Genome Analyzer IIx using 40 base-pair single end reads. The distribution of reads from each sample is plotted to determine the depth of coverage across the genome. The blue track represents the Gaussian distribution with randomly chosen 36 base-pair sequences. Experimental coverage plots show comparable representation for 100 ng and 500 ng gDNA input relative to the theoretical in silico generated library.