TECHNOLOGY INTRO: QuantSeq 3’ mRNA-Seq Library Prep Kit

TECHNOLOGY INTRO: QuantSeq 3’ mRNA-Seq Library Prep Kit

ABSTRACT:
Reading the transcriptome gives an insight how the DNA code - a relatively static state of the cell - is being executed - revealing dynamics behind it. While DNA, or genome sequencing is a term that most of the general population will recognize nowadays, not many might know that one of the earliest methods reading nucleotide sequence was addressing RNA. While at that time sequencing of RNA was technically easier to perform than that for DNA, modern next generation RNA sequencing is, on opposite, more demanding due to the diversity of the transcriptome.

Despite that, RNAseq has seen big advances and is already able to offer a more powerful alternative to, for example, microarrays in both research and diagnostic areas. Furthermore, the potential of RNAseq lies not only within detecting gene expression. Even without good prior knowledge of an organisms genome reading the sequence of RNA reveals structural and functional features of the transcriptome.

Diversity of RNA sequencing applications requires well-tailored approaches to the experimental setup, and a whole transcriptome sequencing is not necessarily the one and only solution here.

For setting up an RNA-Seq pipeline and to plan experiments, it is very important to decide which exact data will be needed to answer the questions arising in your project.

If you are interested in splicing variants or mutations, it would be necessary to cover the entire transcriptome with a complete sequence of each transcript. This requires a lot of reads, sequencing space and money.

However, if you only want to look at expression profiles, it is not necessary to sequence that much. It is sufficient to identify the transcript and to get an estimation the relative amount.
So we thought that RNA-Seq should be defined as two different types - Whole transcriptome sequencing and expression profiling sequencing. For expression profiling sequencing, dedicated library preparation technologies that match your needs can help to save sequencing space, make a high degree of multiplexing possible, and to collect the required data in a very cost efficient way. Also, it can facilitate easier analysis of your data – so it is always important to decide on whether whole transcriptome profiling is necessary or whether expression profiling fits your needs.

QuantSeq is ideal for gene expression profiling with NGS, 3’UTR studies and targeted RNA-Seq. It is unique in a way that library is produced from just one fragment per transcript. QuantSeq allows to save your resources – time wise and cost wise. The workflow takes just 4.5 hours including hands-on and machine time. It is the most cost effective library prep for RNA-Seq because of the price for library prep itself and the possibility to multiplex large number of samples in one sequencing run. Besides that, for QuantSeq bioinformatics analysis is simplified significantly. You can get very accurate gene expression values without need for normalization and FPKM values calculation.

QuantSeq actually became one of my favorite projects. Here we were approached by customer who wanted us to develop a fast, easy and affordable NGS method for gene expression detection focusing primarily on the 3’ end. By having only 1 fragment per transcript we wanted to provide accurate gene expression measurements and also save on sequencing space hence also enabling a higher degree of multiplexing.

We originally pursued two completely different strategies to achieve this goal but in the end what is now known as the QuantSeq protocol made the race, because of its ease of use and the cost factor, which ultimately our customers benefit from.

I’m very happy with the way this project and the QuantSeq product turned out and I would love to develop more kits like this – requested by customers, but also of great interest for the scientific community – something that people really want and need.

QuantSeq is an expression profiling library preparation protocol of a new generation. You do not need anymore to compromise with time, costs and quality of your RNA-Seq.

SPEAKERS:
Dalia Daujotyte, Lexogen GmbH
Birgit Steinmetz, Lexogen GmbH
Jekaterina Aleksejeva, Lexogen GmbH
Pamela Moll, Lexogen GmbH

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[Biomeducated]

As a PhD student, this was highly informative! Thanks!

[solibozorgmehr]

I appreciate the video but why are the speakers creepingly reading from a teleprompter?