BluePippin DNA Size Selection

Posted on by

Elizabeth Zepeda and Katie Carter

Our BluePippin instrument is used for DNA size selection. Several genomic applications benefit from collecting only DNA fragments within a specified size range from a pool of DNA. This service is often used before Illumina sequencing to remove undesired PCR peaks or before long read sequencing to increase the proportion of fragments greater than a given length in a library.

Using pulsed-field electrophoresis on pre-cast gel cassettes, a DNA sample is separated and fragments within the target range are eluted into buffer. Each cassette can run up to 5 samples at once. Up to 5 ug of DNA can be loaded into each well.

The following cassettes are available at CQLS:

  • 3% agarose, 100-250bp
  • 2% agarose, 100-600bp
  • 1.5% agarose, 250bp – 1.5kb
  • 0.75% agarose, 1-50kb

The desired size range must lie within the total size range capability of the cassette. For example, between 200-350 bp on a 2% agarose cassette would be an acceptable range.

The expected collection yield of target sizes is approximately 50-80% based on product validation studies. Yield can vary greatly depending on the desired range and input fragment sizes.

Before and after size selection of Illumina sequencing library to remove undesired peak at approx. 300 bp

Contact Katie Carter for inquiries about this service.

Source: sagescience.com

Research Highlight: Identification of rare microbial colonizers of plastic materials incubated in a coral reef environment

Posted on by

Ed Davis, CQLS

In a collaboration between the Tom Sharpton and Steve Giovannoni labs in the microbiology department, graduate student Seb Singleton designed and performed a study to examine degradation of, and the communities that form biofilms on, plastics in the ocean. Plastic waste accumulation in marine environments is a growing problem that has global effects on the macro and micro scales. In order to understand the ecology surrounding plastic-colonizing bacteria in marine environments, Seb designed a 3 month-long study to examine the changes in biofilm communities as well as structural and chemical changes in the polymer surfaces on high density polyethylene (HDPE), low density polyethylene (LDPE), and polypropylene (PP). An overview of the study design is shown below in Figure 1 from the paper.

Figure 1. Summarized experimental workflow: sample collection (biweekly over 3 months) to downstream analysis [cultivation, 16S (V4) sequence analysis, ATR-FTIR spectral analysis and HIM imaging].

The CQLS, including sequencing using the MiSeq platform in the core lab, as well as bioinformatics consulting done by senior bioinformatics scientist Ed Davis, was integral to the successful outcome of this study. The study encountered several technical roadblocks that were overcome using novel analytical techniques that leveraged the CQLS compute infrastructure. Here is a brief summary of the findings and difficulties overcome:

  • Initial Dominance: Common marine microbial families such as Alteromonadaceae, Marinomonadaceae, and Vibrionaceae were initially prevalent.
  • Community Shift: A significant transition in microbial composition occurred between days 42 and 56, with Hyphomonadaceae and Rhodobacteraceae becoming more dominant. These community shifts also coincided with the passing of Tropical Storm Henri!
  • Rare Taxa: 8,641 colonizing taxa (Amplicon Sequence Variants; ASVs) were identified in total, with 594 overall ASVs enriched on one or more polymer types vs. the glass control, and only 25 ASVs, including known hydrocarbon degraders, significantly enriched on specific plastics.
    • Plastic types differ in the ‘rare’ taxa they recruit: Five were specifically enriched on HDPE, nine on LDPE, and eleven on PP.
  • Taxonomic Assignment Difficulties: Of the 594 significant ASVs, many were unable to be classified to lower taxonomic levels using a classifier trained on the Silva database (i.e. Family and/or Genus level). An alternative classification scheme, called Cladal Taxonomic Annotation (CTA), provided additional taxonomic assignments to 171 (29%) of the significantly enriched ASVs. Most importantly, 8 of the 25 plastic-specific significantly enriched ASVs were better assigned after the CTA.

The shift in taxa over the study period are shown below in Figure 5 from the study:

Figure 5. Gradual temporal shift in a/b diversity shared among material colonizing communities. The Shannon alpha diversity plot (A), Bray-Curtis PCoA (MDS) ordination (B), and Relative abundance stacked bar chart (C) showcase the transition in community complexity and inter-, intra-group similarity over time. In plot (A), alpha diversity measures of the substrate attached communities sharply increases following the mid-experimental transition (between days 42 and 56). Plot (B) explores the compositional dissimilarity of the microbial communities (9,069 unique ASVs) present on the plastics, glass and seawater over the incubation period based on a Bray-Curtis distance matrix. Plot (C) shows the community composition of the top 5% taxa present in each substrate type throughout the incubation period.

Taxa enriched on one or more plastics throughout the study are shown below in Figure 7 from
the paper:

Figure 7. Polymer enriched marine taxa. The Log2foldchange plot showcases NBC classified ASVs that were significantly enriched (adjusted p-value ≤ 0.05) on either one or more polymer types throughout the incubation. The color, size and shape of the data points are associated with the enriched taxon’s class, mean abundance, and substrate preference, respectively. Mean abundance is the average of the sequence depth normalized count values for all included samples, whereas Log2FoldChange is the effect size estimate. All ASVs listed possess >3 log fold differences in abundance compared to glass. Day 42 (and 56 for HDPE) Log2FoldChange data were not included due to loss of sample replicates at the time point, similar rationale was used for Day 14 for all three polymers in respect to the loss of glass control biological replicates.

Degradation of plastics was confirmed using high resolution helium ion microscropy (HIM), and
relevant examples are shown below from Figure 4 of the paper:

Figure 4. Post incubation biodegradation artifacts. HIM images of 77-day incubated polyolefins with biofilm removed in contrast to unexposed controls to exhibit artifacts of biodegradation by colonizing taxa. Marine-incubated LDPE (A) (1–4), HDPE (B) (1–2) and PP (C) (1–2). Unexposed polyolefins: LDPE (A), HDPE (B) and PP (C).

This research highlights the complex interactions between microbes and plastic surfaces in
marine environments, offering insights into the ecological impact of plastic pollution.


Citation: Singleton SL, Davis EW, Arnold HK, Daniels AMY, Brander SM, Parsons RJ, Sharpton
TJ and Giovannoni SJ (2023) Identification of rare microbial colonizers of plastic materials
incubated in a coral reef environment. Front. Microbiol. 14:1259014. doi:
10.3389/fmicb.2023.1259014

COVID Wastewater Surveillance Sequencing

Posted on by

Tyler Radniecki, CBEE

Born at the beginning of the COVID-19 pandemic, OSU’s wastewater surveillance efforts, led by Drs. Christine Kelly and Tyler Radniecki (both professors in the School of Chemical, Biological and Environmental Engineering), are still going strong.  On-going collaborative efforts include researching how wastewater surveillance can contribute to pandemic resilient cities (National Science Foundation), creating a national wastewater surveillance network for tracking antibiotic resistance genes, bacteria and pharmaceuticals (US Environmental Protection Agency), as well as monitoring state-wide community disease dynamics for SARS-CoV-2, influenza and RSV (Oregon Health Authority).  Additional current pilot-scale wastewater surveillance projects include monitoring for Candida auris and antibiotic resistance genes at health care facilities and identifying the presence of the markers for H5N1 influenza strain in Oregon communities. 

Throughout it all, Oregon State University’s Center of Quantitative Life Sciences (CQLS) has been a critical partner in these efforts.  CQLS wet lab staff assist with nucleic acid extractions from wastewater, library preparation and sequencing of wastewater samples.  Additionally, CQLS bioinformatics staff have helped develop and implement bioinformatic pipelines to identify wastewater surveillance targets and report relevant results to OHA and the Center for Disease Control and Prevention.  I can honestly say that every member of the CQLS staff has played a hand in our wastewater surveillance efforts. 

It is due to our collaborative efforts with CQLS that a lot of exciting advancements in wastewater surveillance have been made.  For instance, in collaboration with the OSU TRACE team, we demonstrated that wastewater surveillance is less biased than clinical surveillance at estimating COVID-19 prevalence in a community and that wastewater surveillance can identify COVID-19 hotspots and variant compositions of a community.  We have used wastewater sequence surveillance to identify COVID-19 variants in a community before they were identified in clinical samples.  Additionally, we demonstrated that wastewater sequence surveillance could accurately identify the COVID-19 variant relative abundances in the state, a critical finding as clinical COVID-19 sequencing has declined substantially from its peak.  Finally, we have used wastewater surveillance data to help evaluate the effectiveness of COVID-19 policies implemented by Oregon State University during the first two years for the pandemic.

As we continue to move forward with our wastewater surveillance work, the CQLS will remain a critical collaboration.  Together we are developing novel bioinformatic tools and pipelines to identify strains of RSV, norovirus and influenza.  Additionally, we are moving forward with new wastewater surveillance projects that will explore links between the environment and human pathogens as well as use our national wastewater surveillance network to monitor the spread climate sensitive diseases in the US.  While these endeavors remain challenging, I am grateful to have access to the CQLS to advance our goals.

Dr. Brent Kronmiller appointed as Interim Director of the Center for Quantitative Life Sciences (CQLS)

Posted on by

Colleagues,

It is my pleasure to announce that Dr. Brent Kronmiller has been selected to serve as the Interim Director of the Center for Quantitative Life Sciences (CQLS) effective March 1.

Dr. Kronmiller has been at OSU and affiliated with CQLS for 11 years. Most recently, he has served as the CQLS Assistant Director of the Bioinformatics and Data Science Group that helps consult and collaborate on bioinformatics and data science research across the university. He is also an Assistant Professor (Senior Research) in the Department of Botany and Plant Pathology in the College of Agricultural Sciences.

In the CQLS, he has provided technical and scientific advising to staff and the OSU community and provided guidance and direction to both the CQLS Biocomputing group and the CQLS Core Laboratory group. He also manages the CQLS teaching effort, courses and workshops for students and employees to learn high-performance computing (HPC), bioinformatics and data science skills. Dr. Kronmiller also provides scientific leadership for designing experiments, providing laboratory quotes and troubleshooting errors and issues.

Dr. Kronmiller succeeds Dr. Kathy Higley, who has served as CQLS interim director since December 2021. My special thanks to Kathy for leading the CQLS during this transition period for the center. While Kathy will remain as OSU faculty, she will also be assuming the duties as the President of the National Council on Radiation Protection and Measurements.

Please welcome Dr. Brent Kronmiller to this new role as we take the CQLS to the next stage in support of Prosperity Widely Shared.

Best,

Irem

Irem Y. Tumer, Ph.D., ASME Fellow
Vice President for Research and Innovation
Professor, School of Mechanical, Industrial, and Manufacturing Engineering 

Division of Research and Innovation
Oregon State University | 541-737-0664

An Update to the CQLS Community

Posted on by

Dear CQLS Community,

As the fall term is underway, I recognize that it is long-past time to provide an update on CQLS activities.  Just a reminder, the Center has four distinct competencies, including: laboratory services; bioinformatics and data science; high performance computing (HPC); and health data and informatics.  Each of our four functional groups has had substantial undertakings over the last year. A very brief overview of key activities is offered below

o   Laboratory services:

  • We’ve expanded and reorganized our core lab footprint; this now includes ALS 3012 (A-B), 3020, 3131, 3137 (A-F), 3139 and 2070.  Multi-user and drop-off are now through ALS 3139.
  • Our Core lab is working on proposals to bring updated microscopy instrumentation to OSU.
  • We have a new QuantStudio Absolute Q digital PCR for multi-users.
  • We’ve updated our Genotyping Software.
  • Amplicon sequencing and longer 300 base-pair runs are now available on the NextSeq2000.
  • We have access to Pacific Biosystem’s Revio long read sequencing.
  • Our workflows have improved with the purchases of hoods and minor equipment.
  • Most significantly, the core lab has developed and posted a new position: Assistant Director for the Core Laboratory.  This role will take the lead on maintaining and expanding our core laboratory services, including working with our oversight boards, coordinating support among our constituencies, large scale project management, and coauthoring grants for new equipment which aligns with the evolving science. This open position closes at the end of the month.

o   High Performance Computing (HPC):

  • The CQLS HPC represents nearly 50% of the research computing capabilities at OSU (both for processing and storage).  Over the years CQLS (largely through the efforts of our past Assistant Director Chris Sullivan and current HPC manager Ken Lett) has developed an HPC model that provides a robust, resilient, cost-effective, cutting edge, and accessible computing resource for our users.  
  • The CQLS is highly engaged with campus-wide discussions on the future of OSU’s HPC needs for research computing.  Through this re-imagining the University intends to develop the next generation of HPC users and create a widely available HPC resource while still maintaining expert computing support for individual research domains.
  • We continue to upgrade the CQLS HPC.  Earlier this year our zfs storage server nfs0 was retired as users moved to newer servers.  We continue to upgrade servers and transition users to new hardware.

o   Bioinformatics and Data Science

  • CQLS instructors continue to train OSU researchers on HPC and research computing. Six CQLS workshops were taught last year including “Introduction to Linux”, “Introduction to Python Programming”, “RNAseq Analysis”, “Environmental Sequence Analysis”, and “Genotyping by Sequencing”.  Several courses were also taught in conjunction with the Oregon Data Science Collaborative. 
  • The CQLS maintains an isolated HPC for instruction, the ACTF.  This HPC provides a sandbox learning experience for HPC instruction.
  • Six CQLS Bioinformatics and Data Science consultants are available for collaboration on grant proposals, project discussion, experimental design, pipeline development, and data analysis on projects in bioinformatics and data science.

o   Health Data and Informatics

  • Working with Protected Health Data (PHI) and Personally Identifiable Information (PII), requires incredible attention to the safety and security of the information. Denise Hynes and Matthew Peterson, along with UIT and the Office of Information Security (OIS), coordinate the development of secure cloud resources within Microsoft Azure to facilitate the collection and processing of sensitive data for OSU researchers.
  • In particular, Denise and Matthew have worked to secure Oregon Health Authority (OHA claims data) and deployed Vanderbilt’s Research Electronic Data Capture (REDCap) software in production for multiple NIH grant-funded projects. 
  • The Health and Data Informatics group also maintains a ‘Community of Learning and Practice’ that serves an educational mission for the University, empowered by the CQLS’ newest part-time hire, Joe Spring, who has a wealth of REDCap training and administrative experience.
  • At the Center Level and Beyond:
    • Konnie Handschuch has joined us to provide administrative support to the Center.  Konnie had provided interim support and has now joined CQLS as a permanent member. She divides her time between CQLS and the Research Office.
    • As part of an overall effort to streamline our activities and achieve efficiency, we have handed off much of our budget and expense as well as HR activities to the Research Office.
    • Liz Zepeda has taken on the job of Building manager, stepping into the role after long-time manager, Chris Sullivan moved to CEOAS.
    • While Chris Sullivan moved to CEOAS he left his heart in CQLS.  He remains an incredible advocate for our Center and has worked with Ken Lett (appointed as Infrastructure Manager) to provide seamless support for researchers in CEOAS that are prime users of the CQLS infrastructure.
    • Chris Sullivan, Ken Lett, Chris Thompson (of the College of Engineering) and DRI (Mark Keever) have made tremendous strides in creating single logins, shared storage, and unified job scheduling for consistent access across OSU’s HPC infrastructure, while staying aligned with the security policy of the university.  This is incredibly important for ease of access of users from across and outside of OSU.
    • The next item on our to-do list is the zero-based budget effort, which will kick off soon. Don’t be surprised if some of you are asked to participate in that effort.

It’s been nearly two years since I assumed the role of Interim Director, with my focus on making changes to operations that will position CQLS for long-term success, and its next Director.  We are nearly there.

Sincerely,

Kathryn A. Higley, Ph.D., CHP, HPS Fellow
Interim Director CQLS – Center for Quantitative Life Sciences,
Professor, School of Nuclear Science and Engineering

The Year in Review, and What’s Next

Posted on by | Leave a reply

Dear members of the CQLS Community,

I wanted to reach out and give you an update on the status of the Center.  It has been, as they say, an interesting year.  We have seen an easing of COVID restrictions, and with that return of staff to the Center.  We have had several personnel changes: Bo (Finance Manager), Dana (Bioinformatics Analyst), Matt (Bioinformatics Analyst), and Lynn (Admin. Assistant) have left for new opportunities.  Max Schmidt (Computational Scientist) and Steven Carrell (Bioinformatics Scientist) have joined the Center.  Konnie Handschuch has been serving as our part time Admin Assistant.

There have been changes within our laboratory space as well.  With closing of the previous Director’s lab, we’ve began the process of expanding our core laboratory into that space. A consultant has reviewed laboratory workflows for spatial optimization, and we are excited that this effort will help streamline sample processing and provide better staff space.

As part of functional realignment efforts in the Research Office, we have transferred several administrative duties to them.  Parts of CQLS billing, ordering, and HR efforts now take place in the Research Office.  This change will provide substantial capacity and personnel overlap and minimize bottlenecks.

The biocomputing side of CQLS has continued its efforts to replace and upgrade our computational infrastructure.  That has required us to retire some systems and move researchers on to newer (and improved!) components.  We have also engaged with University Information and Technology (UIT) to aggressively maintain the security posture of our systems.

Chris Sullivan has recently accepted an exciting position within the College of Earth, Oceans and Atmospheric Sciences (CEOAS) but, for the next two months, will work half time with the CQLS to enable a smooth transition. The CQLS will hire an Acting Assistant Director for Biocomputing, and we anticipate a search for a permanent position.

CQLS Bioinformatics and Data Science group continues to teach CQLS workshops and offer research consulting services.  The CQLS trainers teach introductory level workshops for learning high performance computing and Python programming, and bioinformatics workshops to learn analysis of RNAseq, genotyping, and environmental sequencing.  Bioinformatics research consulting analyzes weekly samples for COVID wastewater testing as well as many custom bioinformatics projects across the University.  The newly formed Oregon Data Science Collaborative (ODSC), in conjunction with UO and PSU, brings data science research into the CQLS.  The ODSC teaches workshops and offers research consulting on data analytics, data integration, and machine learning.  The ODSC just hosted its first state-wide data science symposium.

The coming year we will undertake several processes that will help shape the future of the CQLS. First, we will reengage with our stakeholders and steering committees, revisiting the components of the strategic plan.  Secondly, we will undertake a budget analysis in cooperation with the OSU Research Office.  This will help ensure the continued success of the Center.  Third, our Core Laboratory will work with the RO and help them construct CQLS-centered components for the new Research Equipment Laboratory Management System (RELMS).  This will, in turn, help CQLS better serve its customers.

When I accepted the position of Interim Director, I knew that there were challenges that needed to be addressed. Some involved our emergence from the restrictions imposed by the pandemic.  Others were more structural.  One thing that was certain, however, was that the Research Office is very committed to the success of CQLS.  They have been unwavering in their support and continue to help us navigate these challenging times.  CQLS will continue to be here, to provide support, guidance, and a place to grow your research.

Looking forward to this year,

Kathy

Kathryn A. Higley, Ph.D., CHP, HPS Fellow
Interim Director CQLS – Center for Quantitative Life Sciences,
Professor, School of Nuclear Science and Engineering

Chris Sullivan’s Next Steps

Posted on by
Christopher Sullivan at CQLS

Dear CQLS Community,

Chris Sullivan, the CQLS Assistant Director of Biocomputing, has accepted a new position within OSU’s College of Earth Ocean and Atmospheric Sciences.  While we are sad to see him leave, his new position provides exciting new opportunities for him, and continues a long relationship he has had with the computing activities of CEOAS.  Please join us in congratulating Chris in his new position within OSU. 

Chris has been an integral part of CQLS (originally CGRB) since 1999.  Under his guidance the high-performance computing resources in the CQLS have grown from a few computing servers to greater than 7000 processors with more than 7 petabytes of storage and serving more than 20 departments across the University.  Chris has had great success developing bleeding edge computing research for the CQLS and the University.  He has also been phenomenal in his success at obtaining equipment grants from the likes of IBM and OpenPOWER.

Chris will remain with the CQLS at half time for the next three months to enable a smooth transition. He remains committed to the ongoing success of CQLS and to OSU. To ensure there are no interruptions in service the CQLS will fill an interim position to manage this transition before launching a full Assistant Director search in the New Year.

Again, please join us in congratulating Chris.  We will miss him, but we know that he’s just across campus.

Sincerely, Kathryn A. Higley, Ph.D., CHP, HPS Fellow
Interim Director CQLS – Center for Quantitative Life Sciences,
Professor, School of Nuclear Science and Engineering