One of the Chaperon lab members describes his experience after attending a summer school on single cell techniques as one of the trainees.
In September 2020, a single-cell training course with a dense programme was organized by Dr. Monika Holubová from a neighboring lab in the Biomedical Center. The course was open to applicants from all European countries but due to the SARS-CoV-2 ongoing pandemics it was a challenge to be organized safely. A number of applicants and tutors had to cancel their travel to Czechia in a last moment. Hence, one of the empty slots was grabbed Dr. Martin Mokrejš, our bioinformaticist.
The course started with introductory lectures about isolation of individual cells, approaches to perform PCR reactions in a large scale using microfluidics devices, how to prepare sequencing libraries (Illumina) and perform their quality control. Also the tips and tricks lecture was very helpful to everybody.
On the next day, hands-on lab work started. Trainees learned how to isolate individual cells and afterwards how to isolate their RNA content. Later, each of them prepared several Illumina paired-end sequencing libraries. But, the libraries were not from millions of cells (for common bulk RNAseq) but from a few or a few dozens of cells. Some were selected using the flow cytometer and sorter (commonly abbreviated as FACS), some using other methods.
The key device which enabled us to perform the microfluidics reactions was a tiny and fancy box called 10X Chromium controller (from 10X Genomics company). In the end, we obtained copies of portions of cell’s mRNA molecules barcoded per cell and per sample. The barcodes are molecular identifier tags and enable researchers to assign traces of gene expression to not only a sample but to an individual cell within the sample.
Additional days were dedicated to data analysis. Most of the work was handled by quite complicated and very advanced tools written in R programming language and heavily reliant on networked databases, which were necessary for annotation of the genes.
Website of the course is available at: https://singlecell-pilsen.zcu.cz/program.html