SGN-53807 Methods in Single Cell Biology, 3 cr
Additional information
This course replaces the old course SGN-53806 Techniques in Molecular Biology and Applications to Gene Expression
Only 18 students can attend the course every year, due to space limitations in the laboratory. The following criteria will be used to select which students will attend the course. From those enrolled and attending the first lecture, we will select according to the order of enrolment in the course. If you do not attend the first lecture, the next person in the list will be given priority. The order of enrolment is visible in POP.
Course webpage:https://www.cs.tut.fi/~sanchesr/SGN-53807/index.htm
Suitable for postgraduate studies.
Person responsible
Andre Sanches Ribeiro
Lessons
| Implementation | Period | Person responsible | Requirements |
| SGN-53807 2016-01 | 4 |
Andre Sanches Ribeiro |
To pass the course, the student is required to: a) Pass in all three grading requisites. b) Attend at least 50% of the lessons. c) Complete at least 50% of the take home exercises. There is no final exam in this course. |
Learning Outcomes
The student will be introduced to a variety of experimental techniques and analysis tools in single cell, quantitative biology studies. The aim is to provide understanding of the techniques of single cell observation and on how to interpret the results. The focus of this course will be the manipulation and analysis of gene expression in prokaryotic systems along with live cell imaging using state-of-the-art multi-modal microscopy systems. Students will also become familiar with wet-lab methods commonly used to validate results from microscopy measurements. Finally, students will gain understanding of the objectives, applicability and limitations of the methods and techniques in quantitative single cell biology studies. After the course, the student will be able to: 1) Identify, list and define common techniques used in genetic engineering and gene expression analysis, 2) Interpret data generated from the techniques discussed, classify strengths and weaknesses of the methodologies, summarize results and methods, infer consequences of results, and explain procedures. 3) Apply the acquired knowledge to better interpret experimental results. 4) Analyze results of gene expression measurements. Compare methodologies for verifying a hypothesis or measuring a variable. 5) Compare and appraise related methods in single-cell gene expression studies, and interpret and comment conclusions in such studies. 6) Create and develop an experimental procedure in single-cell biology studies using the methods learned, that can address simple questions regarding gene expression in single cells. 7) Have basic knowledge of microscopy techniques.
Content
| Content | Core content | Complementary knowledge | Specialist knowledge |
| 1. | Genetic manipulation in bacteria I | This lecture will introduce the structure, design and applicability of bacterial plasmid vectors to clone a gene of interest and express an RNA and protein. Methods used in sub-cloning, such as PCR, transformation, and sequencing will be reviewed. | |
| 2. | Genetic manipulation in bacteria II | This lecture will continue to review the techniques introduced in the previous week, focusing on genomic insertion by homologous recombination. Methods to evaluate proper gene insertion, such as Northern and Southern blots, and introduction of mutations by site directed mutagenesis will be reviewed. | |
| 3. | Measurements of mean RNA levels at the population level. | This lecture will introduce studies of bacterial gene expression at the single cell level. A number of methods used to detect and quantify changes in RNA levels due to, e.g., stress will be reviewed. Students will be given a summary of multi-cell observation techniques, including real-time PCR and microarray analysis. | |
| 4. | Measurement of RNA levels at the single-cell level | This lecture will introduce studies of the cell-to-cell variability in bacterial gene expression. Techniques capable of counting individual mRNAs will be reviewed, including fluorescence in situ hybridization (FISH), real-time measurements and single-cell RT-PCR. | |
| 5. | Cell culturing techniques | In this lecture, students will be introduced to bacterial cell culturing and will culture their own cells, which will be imaged in the next lecture. | |
| 6. | qPCR, plate reader and gel electrophoresis | Students will learn gel electrophoresis, qPCR, and plate reader techniques. | |
| 7. | Microscopy | Students will learn the basics of how to operate a multi-modal microscope that includes brightfield, phase contrast, epifluorescence and confocal channels. |
Study material
| Type | Name | Author | ISBN | URL | Additional information | Examination material |
| Book | Biochemistry | Berg JM, Tymoczko JL, Stryer L. | No | |||
| Book | Molecular Biology of the Cell. 4th edition. | Alberts B, Johnson A, Lewis J, et al. | No | |||
| Journal | Techniques: recombinogenic engineering ¿ new options for cloning and manipulating DNA | Muyrers, J. P. P., Zhang, Youming, and Stewart, A. F., | No | |||
| Journal | Visualization of single RNA transcripts in situ | Fermino, A. M., Fay, F. S., Fogarty, K. and Singer, R. H., | No | |||
| Journal | mRNA quantitation techniques: considerations for experimental design and application | Reue, K. | No |
Additional information about prerequisites
Basic knowledge of biology/systems biology.
Correspondence of content
| Course | Corresponds course | Description |
| SGN-53807 Methods in Single Cell Biology, 3 cr | SGN-53806 Techniques in Molecular Biology and Applications to Gene Expression, 3 cr | |
| SGN-53807 Methods in Single Cell Biology, 3 cr | BMT-53807 Methods in Single Cell Biology, 3 cr |