For PCR-free, single-cell whole genome libraries with comprehensive coverage and high sequence fidelity

Features

Whole genome libraries from single cells, in under 4 hours

Analyze sequence and copy-number variations anywhere in the genome

Delivers PCR-free NGS libraries and amplified gDNA for follow-up testing

REPLI-g DNA polymerase MDA delivers high-fidelity amplification and reduces false-positives

Includes unique dual indices (UDI) to minimize sequencing artifacts due to “index hopping” on Illumina NGS instruments

Product Details

The QIAseq Single Cell DNA Library Kit UDI provides a complete solution for whole genome sequencing from isolated single cells or low amounts of genomic DNA. The kit includes all reagents required for cell lysis, whole genome amplification using REPLI-g DNA polymerase, enzymatic DNA fragmentation and PCR-free NGS library preparation compatible with Illumina NovaSeq and other NGS instruments. This kit provides comprehensive genome coverage and exceptional sequence fidelity, reducing false positives and minimizing dropouts. Due to the PCR-free workflow, this kit is highly suited for the analysis of aneuploidy, copy number and sequence variation in single cells from rare samples. The kit contains all reagents necessary for DNA library construction, including QIAseq Unique Dual-Index Y-Adapters and QIAseq Beads for reaction cleanup. The QIAseq Single Cell DNA Library Kit UDI can support up to 384 samples in a single flow-cell lane, increasing sample throughput and lowering sequencing costs.

The QIAseq Single Cell DNA Library Kit UDI includes protocols for unbiased whole genome amplification and for target enrichment of small genomes, such as viral DNA or mitochondrial genomes.

Performance

The kit delivers exceptional genome coverage from single cells, populations of cells and low quantities of genomic DNA, including regions with high GC-content. In comparison to PCR-based methods of whole genome amplification, the Multiple Displacement Amplification (MDA) technology included in the QIAseq Single Cell DNA Library Kit UDI eliminates PCR duplicates and minimizes GC-bias, ensuring high library diversity and maximum genome coverage. Additionally, the high-fidelity REPLI-g DNA polymerase introduces fewer sequencing errors than high-fidelity PCR enzymes, reducing false positives and enabling more sensitive mutation detection. Low background and comprehensive coverage make this kit highly suited for the analysis of aneuploidy, copy number variations and mutations from single cells and low-input samples.

Principle

This kit relies on three key technologies to deliver high-diversity whole genome libraries from single cells. After cell lysis, a high-fidelity MDA reaction is used to amplify microgram amounts of gDNA using the high-fidelity REPLI-g DNA Polymerase. The amplified gDNA is then fragmented into short inserts using QIAGEN’s highly random enzymatic FX DNA fragmentation reaction. The fragmented DNA is then end-repaired and an “A” base is added to the 3’ ends to prepare the insert for ligation. Lastly, a ligation reaction adds the unique dual index to both ends of the insert. The entire process, beginning with an isolated single cell, is PCR-free and requires only 3.5 hours to complete.

Procedure

The QIAseq Single Cell DNA Library Kit UDI is a complete cell-to-library solution that includes all the reagents necessary to generate high-diversity whole genome libraries from single cells or limited quantities of intact gDNA. The kit contains reagents for the effective cell lysis of isolated single cells or small clusters of cells, for example from laser-capture microdissection platforms. After lysis, genomic DNA is amplified using the proof-reading, ultra-high fidelity REPLI-g DNA polymerase. This amplified gDNA is subjected to an efficient, random enzymatic fragmentation method, which cleaves the DNA into shorter fragments compatible with any Illumina NGS instrument. These fragments are then end-polished and undergo a highly efficient ligation reaction with the included QIAseq Unique Dual-Index Y-Adapters. This effectively captures the fragmented DNA into a library, which can be purified using the included QIAseq Beads. Additional PCR amplification of the library is unnecessary, ensuring the entire procedure is PCR-free, thus eliminating the possibility of introducing PCR errors or duplicates, and ensuring maximum library diversity.

Applications

Target-specific enrichment of small DNA genomes

Analysis of inter-cellular genome heterogeneity

Mutation detection

Copy number variation analysis

Aneuploidy analysis