Across the globe researchers have been deciphering the genetic code to identify the sequences of nucleotides in the human genome that code for proteins. In many cases, they are attempting to identify and understand small nucleotide variants (SNVs) that lead to mutations and can cause disease. Although human DNA is comprised of over 3 billion base pairs, only 1% (30 million) of these base pairs make up protein-coding regions. These regions are referred to as exons, of which there are approximately 180,000 exons that comprise the human exome.

What are the benefits of Whole Exome Sequencing over other sequencing methods?

Unlike whole genome sequencing (WGS), which sequences the entire genomic content of a DNA sample, WES sequences the DNA that make up the exonic regions of the genome. This is often favorable for researchers who are only interested in the protein-coding regions of the genome because it is cheaper to sequence this much smaller region. Moreover, most known mutations that cause disease occur in exons and therefore it is thought to be more efficient to identify these disease-causing mutations by sequencing the exome, rather than the whole genome.

Researchers and clinicians are more likely to outsource their WES needs for the following reasons.

• Outsourcing can be cheaper than doing it in-house
• Access to greater expertise
• Ability to scale research at speed

Here we break down these three reasons:

Outsourcing WES can be cheaper than doing it in-house

The first step to completing any kind of WES research involves acquiring the right tools, including expensive machinery, such as Illumina’s NovaSeq 6000 platform. This will incur a large cost to researchers as such platforms can cost millions of dollars and require a lot of money to run and service annually. Not to mention the costs of consumables which can cost more than you might think, depending on the number of samples you need to sequence and how frequently the platform is used. Outsourcing WES to Sampled will cost a fraction of the cost of purchasing the equipment and allows researchers to leverage cutting edge sequencing platforms such as the NovaSeq 6000 and if necessary, enquire about the best experimental designs for their investigations by working with our experts.

Greater expertise

Working with sequencing platforms and completing WES investigations can be daunting if you are new to sequencing or have never performed WES before. Even if you have the right sequencing platform in-house, you may not have the right team of experts to sequence your samples, as just one mistake in sequencing can result in costly errors that could lose your team time and resources. This is especially important if the sample you’re looking to sequence is finite or irreplaceable. This is another reason why researchers are outsourcing their work to Sampled, as our highly trained experts are able to help you with experimental design and reduce turn-around-times.

The ability to scale research and decrease turn-around times

In clinical settings, there may be a team dedicated to sequencing patient samples using a sequencing platform. Likewise larger research groups may have a similar team and platform in place to study or identify SNVs which lead to a particular disease. However, the ability to scale up research due to shifting demands is crucial for both groups to be successful. This is where outsourcing WES to Sampled can help both clinicians and researchers. We can take excess samples that arrive at the lab and sequence them for you in order to ensure that a patient won’t have to wait longer than is necessary to receive their diagnosis. Likewise, for non-clinical sequencing results we can provide 100x coverage from as little as 1 business week.

Sampled can help with your whole exome sequencing needs by:

• Reducing your overheads by sparing you the costs of purchasing and maintaining sequencing platforms.
• Not having to hire new team members to run WES investigations by providing you with highly trained experts who can do the heavy lifting for you.
• Scale up or down as you see fit, allowing you to meet deadlines and manage your workload

To talk to an expert about how we can help you with your whole exome sequencing needs, click here.

What is Single Cell RNA Sequencing and how does it work?

Single cell RNA sequencing gives researchers the ability to peer into the unique molecular activity of individual cells, transforming our understanding of cellular biology. It has numerous applications in several research areas, including the study of cancer and immune-mediated diseases.

So how does it work? The first step in scRNA-seq involves isolating single cells and capturing the transcriptome before RNA depletion or degradation occurs. Several methods are available for isolating individual cells, such as micromanipulation, fluorescence-activated cell sorting (FACS), and microfluidics-based techniques. Isolated cells are lysed before RNA is extracted (or fixed), reverse-transcribed into cDNA, and used as a template for high-throughput sequencing on platforms such as those from PacBio and Illumina.

What are the Applications of Single Cell RNA Sequencing?

There are four broad types of investigation using Single Cell RNA sequencing:  

1. Characterization of complex cell populations
2. Discovery of new cell types, subpopulations, or cell states
3. Unravelling complex cellular processes
4. Scaling up functional genomics studies

More specifically, there are many research fields that have already benefited from scRNA-seq including:

1. Neurodegenerative disorders– To try to unravel the interplay between cellular processes in conditions such as Alzheimer’s and Parkinson’s disease.
2. Oncology– In order to investigate the RNA profiles for cell populations within tumors.
3. Immunology– To reveal key regulators of immune function and the cellular interplay between immune cells.
4. Stem cell differentiation in fetus development– In order to understand and define “loss of function” phenotypes in transcription factors and key regulatory factors.
5. Diabetes– To reveal wider gene expression patterns that are involved in Type I Diabetes to predict its development even predating autoantibody production.

As scRNA-seq continues to become more widely adopted, new avenues for investigation are discovered. Advancements in technology have also played a major role making it imperative that researchers choose the most appropriate methods for their research.

Which single cell RNA sequencing platforms should I use?

There are more single cell RNA platforms arriving on the market every year, however this abundance of choice also means it can be hard for researchers who are unfamiliar with scRNA-seq to know which one is best for them. Even within one manufacturer’s series of platforms there are subtle differences that favor certain samples or conditions over others.

For instance, 10X Genomics offers a variety of platforms and chemistries enabling profiling of gene expression and cell surface proteins. With their automated Chromium Connect researchers (or service providers) can go from cells to sequencing-ready libraries with 80% less hands-on time. The instrument enables researchers to generate consistent and reproducible single cell results across experiments users and reduces errors introduced by manual pipetting. However, it may not be ideal for researchers who are looking to conduct high-throughput studies.

The Chromium X series, however, is capable of a range of projects, from pilot studies to the highest-throughput analysis. This platform can generate hundreds of thousands of single cell partitions, each containing an identifying barcode for downstream analysis.

However, these platforms are quite expensive to purchase and maintain which can be off putting to smaller labs or those who are new to scRNA-seq. Furthermore, researchers with a specialist focus may not have staff with the expertise to sequence their samples which forces them to invest more money in hiring staff who can.

Why should you outsource your samples with service provider like Sampled?

Sampled is able to do the heavy lifting, preventing individual researchers and labs from absorbing expensive overhead costs and maintenance of instrumentation. This allows researchers to focus on the work that matters most to them. We pride ourselves on not only purchasing cutting-edge platforms such as the Chromium Connect, Chromium X and HIVE solution apparatus, but also on our ability to work with researchers to give them a tailor-made service. This means working with clients to select the right platform for their needs and work with them on their experimental design before sample processing.

Outsourcing scRNA-seq samples in this manner offers researchers the opportunity to:         

• Save on capital expenditure, as they no longer need to purchase expensive platforms.
• Not have to maintain, troubleshoot, or pay for annual service contracts (saving on Operational expenditure)
• Reduce the need to hire new staff with the right expertise to manage instrumentation and processing.
• Generate high quality data that is repeatable and accurate.

Our industry leading turn-around-times mean that we can go from accepting samples at our lab to having “sequence-ready libraries” in as little as one day. We require just one standard business week to sequence samples and for data to be generated.

We are very proud to be a Certified service Provider (CSP) for 10X Genomics as it gives our clients the knowledge that our trained staff have been taught by 10X Genomics scientists to use their platforms correctly and accurately. Thanks to their training, our experts have a high degree of expertise for sequencing samples with 10X Chromium Connect and 10X Chromium X which is why outsourcing to companies like Sampled is becoming more and more popular.

If you would like to learn more about our technology we provide access to you can find out more here, or if you would like to speak to one of our experts about how outsourcing your single cell work  can accelerate your research, click here.

1. After agreeing to how the samples should be sequenced, the client sends them to the service provider
2. The service provider will then sequence the samples as per the clients’ requested sequencing parameters
3. The service provider sends the raw data and (if they have the ability) the analysis report to the client

Unfortunately, not all companies who provide such services are equal in their offering and not having the right considerations in place before you work with them can result in a loss of time and resources. In this article, we outline four key considerations before outsourcing your samples for RNA-Sequencing.

1. Level of expertise and offerings

First, it should be noted that every service provider who offers RNA-Seq services will have different Next Generation Sequencing (NGS) technology platforms to process your samples on. Moreover, some service providers are offered an opportunity to become “Certified Service Providers” for a specific vendor, but only after they have met their rigorous analytical standards. Having a service provider who is also a CSP ensures that the scientists who are handling your samples are highly trained, capable and have the highest level of expertise. To learn more about certified service providers click here.

Another aspect of expertise that varies between service providers is the ability to provide data analysis and assist in experimental design. Sampled prides itself on aiding researchers who are either new, or unsure of, the best approach to take as it pertains to RNA-Seq. This level of assurance is crucial for investigators who are scaling up their research because it allows them to obtain their data efficiently, without wasting time or resources.

In addition to finding certified service providers, expertise is important for when you require specific types of data analysis. Examples include single cell RNA sequencing (scRNA-Seq), analysis or whole-genome analysis (structural variants, copy number variation, etc.). Not all companies have a bioinformatics team or platform to assist with data analysis which is a part of the service which would be crucial for investigators who are inexperienced with sequencing.  

When considering single cell RNA sequencing designs, one of the most common challenges is the ability to capture the gene expression profile of individual cells so that the sequencing data appropriately reflects the activity of the cells at their time of collection. This is critical when performing time-lapse experiments that require cells collected at different time points, yet require a singular sequencing run. Our experts can work with researchers to select the right transportation and sequencing platforms to ensure that gene expression is preserved for scRNA-seq.

2. Turnaround times for RNA-Sequencing projects

Research is moving faster than ever, and it is imperative that researchers meet deadlines and deliver on time. Depending on the level of work to be undertaken, some service providers may not be able to sequence samples as quickly as others. The need for nucleic acid extraction, processing, number of samples/batches, and downstream analysis can affect overall turnaround time. Typically, 100-200 samples that undergo routine sequencing can take 3-4 weeks and if there are changes to the project design affecting the number of samples and require additional work, more time is required.

At Sampled, we account for these unexpected occurrences and have been an infrastructure with redundant capabilities including multiple short-read sequencers, duplicate PacBio long-read sequencers, and multiple liquid handlers for library preparations. In addition, we also have an automated workflow for 10X single-cell genomics processing. This allows for numerous samples to be added into a production queue in a timely manner. By having a wide array of platforms in duplicate or even triplicate, we can provide turnaround times of as short as 2 weeks for certain projects should they need to be expedited.

3. CLIA approved services

The Clinical Laboratory Improvement Amendments (CLIA) regulate laboratory testing and require clinical laboratories to be certified by the Center for Medicare and Medicaid Services (CMS) before they can accept human samples for diagnostic testing. Laboratories can obtain multiple types of CLIA certificates, based on the kinds of diagnostic tests they conduct. Labs that are involved in clinical trials are able to outsource their research to service providers but require them to be CLIA certified. Sampled has numerous CLIA approved assays which allows researchers to be compliant with federal regulations when working with samples that are involved in clinical trials.

4. Expenditure

While pricing for different services will inevitably vary, there are ways of potentially reducing costs and saving overall on the budget allocated for operational costs. Sampled offers a wide array of services beyond NGS, including data analysis and long-term storage capabilities (including cell lines and DNA/RNA samples). Rather than storing the samples yourself after they have been processed, it could be cheaper to store them with the lab that ran the sequencing instead. Sampled offers a wide range of biorepository solutions as well as downstream processing services that can be bundled to potentially save expenditure to labs that may require these services. To learn more about our wide range of storage solutions, click here.

Selecting the right sequencing partner can be tricky, which is why at Sampled our experts work with clients to ensure that the right platforms and experimental design are selected based on their needs. If you would like to talk to a member of our team to learn more, click here to get in touch with us.