QIAseq FX Single Cell DNA Library Kit (96)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
Thaw, prepare, and aliquot the kit reagents	Follow manufacturer’s instructions Carry out an additional 1:1 cleanup step followed by an additional size selection step according to the KAPA Hyperplus Kit protocol to avoid large, undesired products

Upstream tips
Thaw, prepare, and aliquot the kit reagents
Protocol tips
Follow manufacturer’s instructions Carry out an additional 1:1 cleanup step followed by an additional size selection step according to the KAPA Hyperplus Kit protocol to avoid large, undesired products

Publication protocol

Repli-g MDA reagents were thawed, prepared and aliquoted (Qiagen MIDI kit, QIAseq FX single-cell DNA kit) in PCR Hood 1 and transferred to PCR Hood 2. The enzyme mastermix was kept on ice during cell lysing steps. Sorted samples were thawed in PCR Hood 2, spun briefly and the reaction was initiated according to manufacturer’s instructions with the exception of 20 μl of total mastermix added per sample instead of 40 μl for REPLI-g reactions (Text S1). During lysis, tubes were kept outside of the hood on a precooled rack. The reaction proceeded on a thermocycler with heated lid for 4.5 or more hours. We recommend a routine amplification time of 6.5 h. For 24 MAW0 single-cells, the manufacturer’s protocol for QIAseq FX Single-Cell DNA Kit was followed. In both cases, MDA DNA products were recovered from reaction mixtures with Zymo Genomic Cleanup kits and eluted in 55 μl of water. Library preparations (see below) and MDA products are stored long term at -80°C on separate shelves to reduce the potential for contamination.
Library preparation was carried out according to manufacturer’s instructions for QIAseq FX single-cell DNA Kit. The recommended standard protocol yielded large, undesired products, so we carried out an additional 1:1 cleanup step followed by an additional size selection step according to the KAPA Hyperplus Kit protocol. Additional experiments revealed improved product distribution by increasing fragmentation incubation time to 33 min before proceeding with the recommended cleanup and size-selection step in the Qiagen protocol.



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Manufacturer protocol

Download the product protocol from Qiagen for QIAseq FX Single Cell DNA Library Kit (96) below.

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