Sequel System Data Release: Iso-Seq Results for Hummingbird/Zebra Finch Brain Tissue

[pacbio]

We’ve just released data from Iso-Seq interrogations of brain tissue from two avian models of vocal learning, Anna’s hummingbird (Calypte anna) and zebra finch (Taeniopygia guttata), sequenced in collaboration with the Erich Jarvis and Olivier Fedrigo labs at the Rockefeller University.

For this data set, we used the Iso-Seq method to characterize the transcriptomes of two birds, with brain total RNA. The two species’ brain samples were barcoded, pooled, and sequenced using 4 SMRT Cells on the Sequel System. An average of ~460,000 reads was generated per SMRT Cell; total sequencing data yields ranged from 6.1 to 7.7 Gb per SMRT Cell. More than 15,000 isoforms were identified in each species, including thousands that had not been previously annotated in each bird and 400 to 500 new genes.

Download the data set & read blog post

[nucacidhunter]

I wonder if you could post screenshot of the Run QC for each SMRT Cell detailing productivity, read length, total bases and movie time from SMRT Link as it can be very useful benchmark for sequencing centers and their clients.

[BioKiwi]

Could anyone share their Sequel Run QC of a Iso-seq library. PacBio does not seem keen to share details of what lies behinde their polished data.

[rhall]

See the attached PDF.

[weigrc]

Dear all,

The attached is what we recently done for Iso-Seq application.

We are first time to spike-in Sequel SMRT Cell Control Complex 2.0, anyone knows how to interpret its sequencing data?

Thanks,
Wei

[rhall]

The run looks reasonably good. There are 1593 control reads i.e. its only a very small portion of your data (as it should be), and the polymerase read length and concordance for the control look good, 15105 and 0.85 respectively.
The distribution of polymerase read lengths for your sample is a little lower than for the control, which may indicate the sample isn't of extremely high quality, but it isn't a red flag, given you have 6.56 Gb of raw data.

[nucacidhunter]

Hi Wei, thanks for posting your run results. I wonder what chemistry and software version you have used. Some versions recommends Magbead loading for Iso-seq libraries.

[weigrc]

Originally Posted by nucacidhunter

Hi Wei, thanks for posting your run results. I wonder what chemistry and software version you have used. Some versions recommends Magbead loading for Iso-seq libraries.

We used diffusion loading to keep every size of transcript for sequencing. And refer to below for the versions.

Sequel Sequencing Kit: V2
ICS SW: V5.0.0
SMRT Link: V5.0.1

Thanks,
Wei