SEQXI

SEQPLEX-I WGA Kit

Whole Genome Amplification, DNA Amplification

• Synonym(s): SeqX-i DNA Amplification kit, WGA Kit, Whole Genome Amplification kit

Properties

• technique(s): DNA amplification: suitable; PCR: suitable; whole genome amplification: suitable
• compatibility: Illumina Next Generation Sequencing
• shipped in: wet ice
• storage temp.: −20°C

Description

General description

The SeqPlex^™-I WTA kit allows amplification of smallquantities of reverse transcribed RNA or degraded RNA for direct input ontoIllumina^® next-generation sequencing (NGS) flow cells. TheSeqPlex-i process is comprised of three steps: Pre-amplification/LibrarySynthesis, Amplification 1 and Amplification 2

Step 1: In the Pre-amplification/Library Synthesis stepusing the (Library Preparation Reagents), the template RNA is reversetranscribed using primers composed of a semi-degenerate 3′- and universal5′-ends. As polymerization proceeds, displaced and RNaseH generated singlestrands serve as new templates for additional primer annealing and extensionproducing random, overlapping cDNAs flanked by a universal primer (5′) andprimer complement (3′) sequence.

Step 2: In the Amplified Library Synthesis step (using theAmplification 1 Reagents), products from pre-amplification/library synthesisare amplified by single primer PCR via the universal end sequence. Theseamplification products typically range from 200 to 500+ base pairs. Note:Amplicons from degraded RNA, such as Formalin Fixed Paraffin Embedded (FFPE),are typically shorter and dependent upon the length of the starting RNA.

Step 3: In the Sequencing Library Synthesis step (usingAmplification 2 Reagents), single primer amplicons from amplification 1 areconverted to dual Illumina^® primer PCR products ready forpurification, quantification, and Illumina^® NGS.

To order SeqPlexI adapters in tubes, please download NGSO Adapters SeqPlexI tubes (XLSX File). To order arrayed adapters in plates, download NGSO Plate SeqPlexI arrayed plates (XLSX File). Details on our custom adapters product, Next-Gen Sequencing Oligos (NGSO), can be found at SigmaAldrich.com/nextgenoligos. From this page, you can directly access an online ordering configurator by clicking on the "Order Now" option. Both spreadsheets are directly uploadable after selecting the quantity and/or format from the dropdown menus. For adapters in solution in tubes or in plates, NGSO-Silver is the only option available, which has a length cutoff of 60 bases (several of the adapters are longer than 60 bases). In addition, adapters in plates cannot be duplexed. If you would like a feasibility assessment of ordering NGSO-Silver longer 60 bases or duplexed in plates, then please send a request to dnaoligos@milliporesigma.com. All of the adapters are readily available for online ordering if dry and in tubes as either NGSO-Bronze or Gold (recommended purification for any adapter).

Application

SeqPlex^™-I WGA Kit has been used for whole genome amplification.

Packaging

24, 384, 96 reactions

Features and Benefits

• Amplifies fragmented/extremely small quantities of DNA: Yields from ChIP or FFPE, which are 200-500 bp are easily amplified by Random priming technology.
• Facilitates sequencing from as little as 100 pg of DNA
• Enhanced primers for complete genome coverage, minimal sequence bias, and amplicon size ideal for next generation sequencing (NGS)
• Cost-effective: No longer requires an additional NGS library prep step
• Compatible with Illumina^® next-generation sequencing

Other Notes

• A 20 μL Amplification 2 reaction will produce >100 ng of amplified double-stranded cDNA when starting with 100 pg to 5 ng of high-quality DNA. Higher input quantities and higher quality DNA template generally result in increased yields. For damaged DNA, such as from FFPE, 1–50 ng input DNA is recommended.
• The dual index adapter primers (AP100) provided in this kit will only work for one sample. If pooling of samples for sequencing is required, the user must provide additional index primer sets. See example index primer sequences on page 2 of the technical bulletin.

Legal Information

Illumina is a registered trademark of Illumina, Inc.

SeqPlex is a trademark of Sigma-Aldrich Co. LLC

Disclaimer

The SeqPlex-I DNA Amplification Kit for whole genome amplification (WGA) is for R&D use only. Not for drug, household, or other uses.

Safety Information

• Storage Class Code: 10 - Combustible liquids
• WGK: WGK 3