Transcriptome analysis of single cells: a plankton characterization of Arctic waters
The functional and taxonomic diversity of marine protists in the Arctic is large and a scientifically underestimated source of biodiversity. However, this diversity is being masked in terms of environmental bulk sampling, just as it is done within population-averaged samples retrieved from pure cult...
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| Format: | Thesis |
| Language: | unknown |
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University of Applied Sciences Bremerhaven - Faculty 1, Bremerhaven, Germany
2018
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| Online Access: | https://epic.awi.de/id/eprint/48872/ https://hdl.handle.net/10013/epic.fa205a4a-b21e-4bd2-8f8c-d6b2ef6dc87c |
| Summary: | The functional and taxonomic diversity of marine protists in the Arctic is large and a scientifically underestimated source of biodiversity. However, this diversity is being masked in terms of environmental bulk sampling, just as it is done within population-averaged samples retrieved from pure cultures. Yet in recent years, low-input RNA-sequencing methods have been adapted to work with single cells. Thus, increasing the number of unicellular transcriptomes sequenced and deepening the knowledge about the species distribution as well as phylogeny based on functional data analysis pipelines. Within this thesis, the SMART-Seq v4 protocol by Takara Clontech was successfully applied to study single cells during the HE492 field trip in Arctic waters. Further, it greatly enabled the simultaneous processing of multiple protists around Spitsbergen. Altogether, 42 single cells have been processed with the SMART-Seq v4 protocol. Afterwards 21 samples were indexed according to the Illumina Nextera XT library preparation protocol and subsequently 20 individual cDNA libraries pooled for sequencing on the Illumina NextSeq 500 platform. After sequencing each transcriptome has been de novo assembled and annotated via the Trinotate processing pipeline. Overall, the thesis evaluates the work flow establishment regarding single cell transcriptomics as a tool for field sampling and analysis positively, as resulted sequence data could be promisingly functionally annotated. The results demonstrate the possibility to study single cells to quantify inter-population heterogeneity previously masked in bulk measurements and moreover the method bypasses the need of cell cultivation. Ultimately single cell approaches will open new analytical avenues for studying culture independent unicellular plankton species in terms of cell subtypes and gene expression dynamics even in remote areas. |
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