What you see here is the average face of the average person currently living on planet earth. By averaging we lose all the little details that make us unique. Our beauty is in our difference and this is true at all levels: We are different at all scales! . From our faces, our minds, our proteins down to individual cells that make up our body even down to single molecules. National Geographic
Our beauty is in our differences Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
Our beauty is in our difference and this is true down to level of individual cells and individual molecules.
School of Electronic and Electrical Engineering Single-Cell Genomics Dr Paolo Actis School of Electronic and Electrical Engineering p.actis@leeds.ac.uk @paoloactis
Single-Cell Genomics Single-cell genome sequencing: current state of the science, Nat Rev Genetics 2016 Single Cell Genomics: Advances and Future Perspectives, PLOS Genetics, 2014
Single-Cell Genomics 1. Cell Isolation
Single-Cell Genomics 1. Cell Isolation 2. Genome Amplification
Single-Cell Genomics 1. Cell Isolation 2. Genome Amplification 3. Genome Sequencing
Single-Cell Genomics 1. Cell Isolation 2. Genome Amplification 3. Genome Sequencing 4. Bioinformatics analysis
1. Cell Isolation
Cell Isolation Manual Micropipetting ✔Visual confirmation, Applicable to low number of cells ✖Low throughput, Operator Bias
Cell Isolation FACS (fluorescence activated cell sorting) ✔High Throughput , Sorting based on phenotype ✖Large amount of cells required, Putative damage to cells, Occasionally more than one cell isolated
Cell Isolation Microfluidics ✔High Throughput , highly standardized, nanoliter reaction volumes, automated with visual confirmation ✖Putative loss of cells
Cell Isolation Single nuclei sorting ✔No need to isolate whole cell, isolation form cryopreserved or fresh tissues ✖Potential loss of micronuclei
Cell Isolation Laser Capture Microdissection ✔Retains topological information of the cell, isolation form cryopreserved or fresh tissues ✖Contamination with other cells, Potential loss of cellular material
Single-Cell Nanobiopsy
Electrowetting in a nanopipette H2O DCE
Electrowetting in a nanopipette
Single-Cell Nanobiopsy
Single-Cell Nanobiopsy Actis et al, ACS Nano, 2013
RNA-Sequencing High read coverage of RNA sequencing of nanobiopsies Full length cDNA from transcripts isolated by nanobiopsy
Organelle Surgery
Single-Cell Nanobiopsy
Single-Cell Nanobiopsy
Sub-Cellular Sequencing
Sub-Cellular Sequencing
Sub-Cellular Sequencing
Ambition Nucleolar trafficking during virus infection (Ade Whitehouse, FBS) Sub-cellular Sequencing (Thierry Voet, Sanger Institute) Omics analysis (You)
2. Genome Amplification
Genome Amplification
Genome Amplification High Uniformity but High error rates
Genome Amplification
Genome Amplification Low Uniformity but Low error rates
Genome Amplification
intermediate error rates Genome Amplification High Uniformity but intermediate error rates
3. Genome Sequencing
Genome Sequencing Easy …just ask Ian Carr
4. Bioinformatics analysis
Amplification artefacts Loss of coverage Amplification artefacts
False positive True positive False Negative (locus dropout)
Single-Cell Genomics 1. Cell Isolation 2. Genome Amplification 3. Genome Sequencing 4. Bioinformatics analysis
Our beauty is in our differences Dr Paolo Actis School of Electronic and Electrical Engineering p.actis@leeds.ac.uk @paoloactis Our beauty is in our difference and this is true down to level of individual cells and individual molecules.