p a c b .c o m / b i o t e c h

Long reads were used to identify exact vector-mediated AAV viral integration sites within a human diploid cell line1.

“WITH THE FLEXIBILITY TO EVALUATE ENGINEERED NUCLEASES AND TARGETING CONSTRUCTS DIRECTLY AT

DESIRED LOCI WITHOUT THE DEVELOPMENT OF REPORTER SYSTEMS, SMRT DNA SEQUENCING CAN STREAMLINE THE

DEVELOPMENT OF GENOME-EDITING PROJECTS...”

— AYAL HENDEL, STANFORD UNIVERSITY2

“[SMRT SEQUENCING] IS STILL UNIQUE IN ITS ABILITY TO RESOLVE HIGHLY-SIMILAR REPETITIVE

SEQUENCES THANKS TO AN AVERAGE READ-LENGTH OF OVER 10 kb AT AN AFFORDABLE

100 FOLD COVERAGE…”

— GOMEZ-ESCRIBANO, JOHN INNES CENTRE4

A PacBio® de novo assembly of Streptomyces putatively detected an intact NRPS gene cluster in a single contig, while a short-read assembly contained two incomplete fragments of the gene cluster3.

VALIDATE TARGETS WITH CONFIDENCE - Unambiguously characterize integration sites

within host genomes - Validate vector population prior to

transformation - Achieve Sanger-like quality with higher

throughput and more efficient workflow

FUEL BIOTECH DISCOVERY WITH CONFIDENT CHARACTERIZATION OF MICROBES AND THEIR COMMUNITIES

Industrial microbiologists rely on comprehensive genomic information to identify and develop complex biological products. Single Molecule, Real-Time (SMRT®) Sequencing delivers a more complete view of individual organisms and microbial communities, fueling research for modern pharmaceutical discovery, environmental remediation, chemical commodity production, and agriculture products.

DISCOVER MORE IN GENOME MINING - Generate complete microbial assemblies - Reconstruct intact genes and elucidate gene clusters - Identify critical members of complex microbial

communities - Sequence intact transcriptomes and

metatranscriptomes to directly define full-length bacterial operons

a

b

KEY REFERENCES1. Hüser, D. et al. (2014) Adeno-associated virus type 2 wild-type and vector-mediated genomic integration profiles of human diploid fibroblasts

analyzed by third-generation PacBio DNA sequencing. Journal of Virology. 88(19), 11253–11263.

2. Hendel, A. et al. (2014) Quantifying genome-editing outcomes at endogenous loci with SMRT sequencing. Cell Reports. 7 (1), 293-305

3. Harrison, J. & Studholme, D. J. (2014) Recently published Streptomyces genome sequences. Microbial Biotechnology. 7 (5), 373–380.

4. Gomez-Escribano, J. P. et al. (2016) Next generation sequencing of Actinobacteria for the discovery of novel natural products. Marine Drugs. 14(4), 1-19.

5. O’Connell Motherway, M. et al. (2014) Identification of Restriction-Modification Systems of Bifidobacterium animalis subsp. lactis CNCM I-2494 by SMRT Sequencing and associated methylome analysis. PLoS ONE. 9(4), e94875.

6. Badran, A. H. et al. (2016) Continuous evolution of Bacillus thuringiensis toxins overcomes insect resistance. Nature. 533(7601), 58-63.

7. Boitano et al. (May, 2015) Full-length cDNA Sequencing of Prokaryotic Transcriptome and Metatranscriptome Samples. American Society for Microbiology. New Orleans, Louisiana.

8. Marsic, D. et al. (2014) Vector design Tour de Force: integrating combinatorial and rational approaches to derive novel adeno-associated virus variants. Molecular Therapy. 22(11), 1900-1909.

9. Monk, I. R. et al. (2015) Complete bypass of restriction systems for major Staphylococcus aureus lineages. mBio. 6(3), e00308-15.

10. Singer, E. et al. (2016) High-resolution phylogenetic microbial community profiling. The ISME Journal. 1-13. doi:10.1038/ismej.2015.249

“…IT IS NOW APPRECIATED THAT STRAIN-SPECIFIC R-M BARRIERS ARE A CONTRIBUTING FACTOR IN THE INABILITY TO GENETICALLY ACCESS SEVERAL BIFIDOBACTERIAL STRAINS.”

— O’CONNELL MOTHERWAY, NATIONAL UNIVERSITY OF IRELAND5

For Research Use Only. Not for use in diagnostic procedures. © Copyright 2016, Pacific Biosciences of California, Inc. All rights reserved. Information in this document is subject to change without notice. Pacific Biosciences assumes no responsibility for any errors or omissions in this document. Certain notices, terms, conditions and/or use restrictions may pertain to your use of Pacific Biosciences products and/or third party products. Please refer to the applicable Pacific Biosciences Terms and Conditions of Sale and to the applicable license terms at http://www.pacificbiosciences.com/licenses.html.

Pacific Biosciences, the Pacific Biosciences logo, PacBio, SMRT, SMRTbell, Iso-Seq, and Sequel are trademarks of Pacific Biosciences. BluePippin and SageELF are trademarks of Sage Science. NGS-go and NGSengine are trademarks of GenDx. All other trademarks are the sole property of their respective owners. PN: VM105-060616

CAPTURE EPIGENETIC INFORMATION TO IMPROVE GENETIC ACCESSIBILITY - Simultaneously capture epigenetic information while collecting genomic sequencing data - Explore how epigenetic regulation affects the efficiency of microbial genetic manipulation - Design plasmid DNA with appropriate methylation to overcome restriction-modification (R-M) barriers

With the knowledge gained from a SMRT Sequencing methylome analysis of the predicted R-M systems in a Bifidobacterium animalis strain, scientists achieved a 22-fold higher transformation efficiency using appropriately methylated plasmid DNA compared with unmethylated DNA5.