Optical DNA Mapping of the Saccharomyces cerevisiae Genome
| dc.contributor.author | Zachrisson, Hanna | |
| dc.contributor.department | Chalmers tekniska högskola / Institutionen för life sciences | sv |
| dc.contributor.department | Chalmers University of Technology / Department of Life Sciences | en |
| dc.contributor.examiner | Westerlund, Fredrik | |
| dc.contributor.supervisor | Leal Garza, Luis Mario | |
| dc.date.accessioned | 2023-06-30T11:50:47Z | |
| dc.date.available | 2023-06-30T11:50:47Z | |
| dc.date.issued | 2023 | |
| dc.date.submitted | 2023 | |
| dc.description.abstract | DNA molecules are present in every living organism and contain all the genetic information necessary for an organism’s existence. It is of great interest to extract this information from the DNA molecules and one common approach to doing so is through sequencing, in which the sequence of the DNA molecules is obtained with base pair resolution. However, structural variations (SVs), events affecting the chromosome organisation, can be challenging to detect with sequencing techniques as relatively short reads are generated which are hard to localise in the genome. SVs are however important to detect as they could be associated with diseases. Optical DNA mapping (ODM) is a complementing technique for obtaining genomic information, including SVs. In this master thesis project, ODM was performed on the genome of the budding yeast Saccharomyces cerevisiae (strain BY4742), with the aim to perform reference-based assembly and explore first steps towards a de novo assembly of its genome. This was performed by culturing the yeast strain, extracting the DNA from the yeast cells, and performing ODM based on competitive binding (CB). ODM was performed by labelling the DNA molecules in a sequencespecific manner using YOYO-1 and netropsin, stretching the DNA molecules in nanochannels in a nanofluidic chip, and imaging the molecules using fluorescence microscopy. Kymographs and corresponding barcodes were generated from each DNA molecule and were used for the data analysis. With the generated barcodes, reference-based alignment was performed as well as de novo assembly experiments. The results from the reference-based alignment indicate good quality data which could be used to perform the de novo assembly. However, especially chromosome I and XII showed unexpected results which could indicate complex or currently unmapped regions within these chromosomes. A complete de novo assembly of the genome was not obtained in this project. However, high quality ‘bargroups’ were generated from the assembly experiments, which indicates that the assembly pipeline could potentially assemble the whole genome. To this date, this project is the first time CB-based ODM using a nanofluidic chip is applied on the genome of S. cerevisiae, and is a first step towards a de novo assembly of its genome using ODM. By further improving the assembly pipeline, there is a high probability that the data collected in this project is able to generate a reliable de novo optical genome assembly of the yeast Saccharomyces cerevisiae, which could be used for detecting SVs. A future objective is to expand and apply this technique on the human genome to detect SVs associated with diseases, and thereby utilising it as a diagnostic tool. | |
| dc.identifier.coursecode | BBTX03 | |
| dc.identifier.uri | http://hdl.handle.net/20.500.12380/306519 | |
| dc.setspec.uppsok | LifeEarthScience | |
| dc.subject | DNA | |
| dc.subject | Structural variations | |
| dc.subject | Optical DNA Mapping | |
| dc.subject | Saccharomyces cerevisiae | |
| dc.subject | Competitive binding | |
| dc.subject | YOYO-1 | |
| dc.subject | Netropsin | |
| dc.subject | Reference-based alignment | |
| dc.subject | De novo assembly | |
| dc.title | Optical DNA Mapping of the Saccharomyces cerevisiae Genome | |
| dc.type.degree | Examensarbete för masterexamen | sv |
| dc.type.degree | Master's Thesis | en |
| dc.type.uppsok | H | |
| local.programme | Biotechnology (MPBIO), MSc |