Size Selection Resources for Long-Read Sequencing
Size Selection Chart
This chart summarizes the current Sage Science platforms, applications, and status for PacBio sequencing (click to enlarge):
PacBio’s Iso-Seq application is a great tool for RNA researchers. We’re pleased to provide size selection capabilities for this application with both the BluePippin and the SageELF platforms. Learn more about Iso-Seq from this poster, and this one.
30 Kb – 40 Kb High Pass on the BluePippin
Our new High Pass protocol enables size selection of fragments longer than 30 Kb to 40 Kb. This requires a new kit using Marker U1 (part number PAC30KB). Please note: you must enter 80000 for the bpEND value for this new protocol. Here’s an example of a size-selected shear, analyzed on the Pippin Pulse.
Revised BluePippin High Pass User Guide
This includes all High Pass protocols for the BluePippin.
High-Throughput High Pass with the PippinHT
High Pass protocols are now available on our PippinHT instrument. This can run 2 X 12-lane cassettes at a time. Since the High Pass protocol requires an external marker, up to 22 samples may be size selected in a run. The maximum loading amount for the HT is 1.5 ug (vs. 5 ug on the BluePippin), but leads to ~2-fold greater sample recovery. This PDF compares High Pass recoveries on the BluePippin vs. Pippin HT.
Pippin Pulse for Fragment Analysis
The Pippin Pulse instrument provides researchers with an affordable tool to resolve DNA out to 100 Kb and beyond. Based on the pulsed-field power supply from our BluePippin, the Pippin Pulse is a great alternative to expensive analysis platforms for ultra-long DNA fragments. To access PacBio’s guidelines for using the Pippin Pulse, click here.
Check out these app notes from PacBio users:
High Pass protocol on BluePippin for human genomic DNA
BluePippin for size selection of three bacterial libraries
Several blog posts and publications describe the work of many PacBio users who rely on BluePippin or SageELF for library prep. Check them out here.