VR真人彩票

Introduction

A decades long fascination with RNA led Professor Jo Vandesompele, PhD to found Biogazelle in 2007. The company provides transcriptome analysis and experimental design services to pharmaceutical, biotechnology, and clinical researchers. Its breadth of services include: RNA biomarker discoveries and validation through customized RNA quantification studies; high-throughput antisense oligonucleotide screenings; the identification, measurement, and silencing of long-coding RNA (lncRNA) targets; and assisting customers with lead drug discovery and preclinical studies.

Noncoding RNA studies are at the heart of much of Biogazelle鈥檚 oncology work. Some cancer cells appear to be addicted to the presence of certain lncRNAs and die when the transcript is silenced.¹ Working with formalin-fixed paraffin-embedded (FFPE) and liquid biopsy samples can be difficult because of the low input amounts and highly fragmented nature of these cancer samples. Biogazelle uses the NextSeq 500 System and TruSeq RNA Exome kit to perform RNA-Seq, obtaining the most information from these samples.

Jo Vandesompele, PhD, is a Professor at Ghent University, and Co-Founder and Chief Scientific Officer at Biogazelle.

Making the Most of Highly Fragmented Samples

According to Professor Vandesompele, there isn鈥檛 a better instrument available for RNA-Seq than the NextSeq 500 System. 鈥淚t has the capacity, flexibility, and scale that we need to perform total RNA-Seq,鈥� Professor Vandesompele said. 鈥淚t is also very forgiving in terms of supporting diverse library types.鈥�

These cancer specimens are extremely valuable. 鈥淥ur customers and partners don鈥檛 want to sacrifice biomaterial for a method that requires a microgram or even 100 nanograms of RNA,鈥� Professor Vandesompele added.

Biogazelle works with VR真人彩票 to reduce the sample amount needed for various RNA-Seq applications. 鈥淲e鈥檙e seeking answers about how small the sample can be to perform RNA-Seq successfully,鈥� Professor Vandesompele said. 鈥淲ill 10 nanograms work? How about 1 nanogram? Can we sequence a liquid biopsy sample where we cannot even reliably measure the RNA concentration? Surprisingly, it鈥檚 not difficult with a few simple, clever modifications. We can perform accurate and sensitive RNA measurements with as little as a few nanograms of total RNA or only 200 碌l of a biofluid.² We are grateful for the VR真人彩票 team鈥檚 support in enabling us to achieve this.鈥�

The NextSeq 500 System "has the capacity, flexibility, and scale that we need to perform total RNA-Seq. It is also very forgiving in terms of supporting diverse library types."

Overcoming the Challenge

The TruSeq RNA Exome Library Prep Kit offers a 鈥済reat solution鈥� to address FFPE and liquid biopsy samples that combine the challenges of ultra-low input and highly fragmented RNA. It converts total RNA into template molecules of known strand origin, followed by sequence-capture of exons of interest. 鈥淥ther total RNA prep methods consume many reads on introns that are not meaningful and generate lower quality data,鈥� Professor Vandesompele stated. 鈥淭he RNA Exome probe-based capture method is superior and works like a charm.鈥�

The one limitation of RNA Exome is that the probes target only protein-coding genes. 鈥淲hen we performed total RNA-Seq, the lncRNAs were almost at the noise level, with 95% of the reads going to highly abundant coding protein genes,鈥� Professor Vandesompele explained. 鈥淎s a solution, VR真人彩票 used the comprehensive human LNCipedia database that we created at Ghent University to develop custom probes against all of them.³ Supplementing the kit with custom probes targeting these low-abundant, challenging lncRNAs enables us to perform fantastic experiments. It is also opening up numerous possibilities for our customers to study things that they couldn鈥檛 before, which is great.鈥�

鈥淭he RNA Exome probe-based capture method is superior and works like a charm.鈥�

Modifying Workflows for Low Input Samples

While TruSeq RNA Exome is a good technology for handling low input amounts, Biogazelle has made some workflow modifications depending on the RNA molecule. 鈥淔or low RNA input amounts, we apply several different tricks, such as increasing the input volume by concentrating the sample,鈥� Professor Vandesompele said.

The Biogazelle team found that multiplexing prior to capture with TruSeq RNA Exome is a problem with liquid biopsies. 鈥淏ecause of the varying RNA content, our libraries are never equimolar,鈥� Professor Vandesompele added. 鈥淚nstead of pre-capture multiplexing, we perform separate captures for each sample. It is more work, but the data are significantly better and the libraries are equimolar.鈥�

鈥渓ncRNAs offer promise in the fields of oncology, neurology, and other disease types. We鈥檙e at the beginning of a completely new type of medicine.鈥�

High Value RNA-Seq Oncology Studies

Professor Vandesompele believes that RNA-Seq data provides the same, if not more, informative data as DNA sequencing. The biomarker potential is significantly greater with 49,000 high-confidence lncRNA genes versus 21,000 protein-coding genes. However, the value might lie in looking at RNA and DNA signatures together to assess cancer or other diseases. 鈥渓ncRNAs offer promise in the fields of oncology, neurology, and other disease types,鈥� Professor Vandesompele said. 鈥淲e鈥檙e at the beginning of a completely new type of medicine.鈥�

Read more about RNA-Seq studies at Biogazelle:

Biogazelle iCommunity article

Learn more about the products and systems mentioned in this article:

NextSeq 500 System

References

1. Huarte M and Rinn JL. Hum Mol Genet. 2010;19:152-161.
2. Biogazelle. Expertise/Liquid biopsies.听. Accessed. June 17, 2019
3. LNCipedia. A comprehensive compendium of human long non-coding RNAs. . Accessed May 28, 2019.