A paper that came out December in the journal Electrophoresis compares Pippin Prep size selection with Caliper’s Labchip XT and finds that Pippin gives “very tight size distributions” not seen with other sizing methods.
“Evaluation and optimisation of preparative semi-automated electrophoresis systems for Illumina library preparation” from lead author Michael Quail and colleagues at the Wellcome Trust Sanger Institute, demonstrates several types of comparisons between the systems. The authors note in their premise that better sizing techniques are needed since manual gels are time-intensive and lack reproducibility while SPRI beads often give ranges “too broad for de novo assembly applications.” The authors assessed size selection methods for general library preparation, PCR bias, noPCR use, and more.
The importance of accurate size selection goes beyond removal of small fragments, including adaptors and adaptor-dimers, to improve de novo sequence assembly and allow for chromosomal rearrangement prediction, the authors note.
In their first test for whether the instruments performed according to their specifications, “both platforms collected the specified size fractions but with quite different results,” Quail et al. write. “The Pippin Prep gave very tight size distributions whereas those from the Labchip XT were much broader. … The distribution of recovered fragment sizes was much more reproducible with the Pippin Prep.” Such tight sizing can lead to lower yield, the authors note, adding that “this can partially be overcome by setting the instrument to collect a wider size range.”
The study also looked at the effects of performing size selection prior to PCR amplification, finding that the tightest possible size fraction was advantageous before amplification. “Whilst Pippin Prep eluted DNA, size selected using the ‘tight’ setting, gave sharp and representative size distributions following PCR, we regularly observe smaller fragment ‘shoulders’ … following amplification of Labchip XT fractionated ligation products,” the scientists write.
While this paper focused on sample prep for the Illumina sequencing workflow, the authors note that Pippin works well with other next-gen technologies too. “We have found the Pippin Prep to be useful for both size fractionation of 100bp and 200bp fragments for the Ion Torrent PGM, and for size fractionation of 3kb fragments for Pacific Bioscience sequencing,” they write.