Validation of the DNA Fluid+ Kit on viscous samples is shown below. HMW DNA was extracted from GM12878 cells using the Circulomics Nanobind CBB Big DNA Kit. First, the sample was pre-sheared using the DNA Fluid+ Kit (speed 59) and then diluted to 50 ng/uL and sheared using the Long Hydropore (speed 31) on the Megaruptor 3 system. PacBio HiFi sequencing was performed on the sheared DNA using a 30 hr movie, SMRTbell Express Template Prep Kit 2.0, Binding Kit 2.0, and Sequel II Sequencing Kit 2.0, and Circulomics SRE XS size selection. These results show that Circulomics SRE XS is a good substitute for gel-based size selection and that the DNAFluid+ works well to pre-condition DNA for Long Hydropore shearing without clogging.
Fig.1. The Femto Pulse traces show fragment size distributions of: 1) native HMW DNA isolated from GM12878 cells using the Circulomics Nanobind CBB Big DNA Kit (black curve), 2) after pre-shearing with the Diagenode DNA Fluid+ Kit (blue curve) and 3) after shearing to ~18 kb with the Diagenode Long Hydropore using Megaruptor3 (red curve). Pre-shearing the HMW DNA enables more consistent shearing performance across samples of varying viscosity.
Fig. 2. After shearing the gDNA that was pre-conditioned with the Diagenode DNA Fluid+ Kit, HiFi read length distribution of GM12878 SMRTbell library shows excellent read length distribution. Sequencing was performed on the PacBio Sequel II system.