Pacific Biosciences (PacBio) Third Generation Sequencing

PacBio Applications:

• De Novo Assembly: To finish genomes in order to fully identify, annotate and decipher genomic structures.
(www.pacb.com/denovo)

• Targeted Sequencing: To comprehensively characterize genetic variations including structural variants, haplotypes, and rare SNPs. (www.pacb.com/target)

• Base Modification Identification: Using real-time kinetic data to characterize genetic regulation and DNA damage. (www.pacb.com/basemod)

 

Library Prep Type	Sample Input Required	~ Average Insert	~ Mbp SMRTCell
10 kb MagBead*	1-10 ug	2,500 bp**	65-140 Mbp
10 kb CLR	5-10ug	2,500 bp	65-140 Mbp
5 kb CLR	2-4 ug	2,000 bp	60-120 Mbp
1kb CCS	500 ng - 1ug	500 bp	40 Mbp
500 bp CCS	250 ng - 1ug	350 bp	20 Mbp
250 bp CCS	250 ng - 1ug	150 bp	7.5 Mbp

 

*CLR: Continuous Long Read
*CCS: Circular Consensus Sequence (Note: CCS Mbp estimates are based on nonredundant coverage.)
**MagBead: PacBio MagBead sequencing provides more reads at longer lengths (comparative to a standard PacBio CLR library) and   reduces the sample input significantly.

Table Note: The standard result estimates outlined in the table above are based on output averages of several different projects and are to be used as a guideline. Individual project results may vary from these estimates. PacBio read-lengths and read quantities per SMRTCell are highly dependent on sample quality.

Quality Control: NCGR is committed to providing the best possible library prep and sequencing results for every sample we process. NCGR QC-tests all samples via Qubit (quantity) and Bioanalyzer and/or Gel (quality). 

PacBio Turnaround Time: Approximately four to eight weeks after receipt of signed quote, completed "NCGR Sample Form" documentation, and experimental samples of adequate sequencing quantity and quality.

XL Chemistry: January 2013 - NCGR will begin offering PacBio sequencing using the new and improved XL polymerase chemistry. PacBio reports that the use of the XL enzyme yields extraordinarily long read lengths that average 5,000 bases, with some reads as long as 20,000 bases and an enhanced sequencing output of up to approximately 250 Mbp per SMRTcell.