Virtual laboratory

DNA and genes

Genotyping, breeding of genetically modified lines, CRISPR genome editing, mutagenesis and cloning.

1. Chromatin immunoprecipitation (ChIP-qPCR)Chromatin immunoprecipitation to test whether the transcription factor CREB, or an activity-dependent histone acetylation, is present at the promoter of the Fos gene and increases when neurons are stimulated.Guided experiment
2. Restriction cloning of a gene into a plasmidConstruction of a recombinant plasmid carrying a fluorescent-protein gene, from the choice of vector and restriction enzymes to a sequence-verified clone.Guided experiment
3. Breeding and genotyping a mouse lineBreeding and genotyping of one litter of a knockout mouse line, from the choice of breeders to a genotyped cohort.Guided experiment
4. Gene knockout in cultured cells with CRISPR–Cas9Knockout of a gene in a cultured cell line with CRISPR–Cas9, from the choice of guide RNA to a clone that lacks the protein.Guided experiment
5. Changing one amino acid: site-directed mutagenesisOne defined point mutation is made in a gene carried on a standard laboratory plasmid, from the choice of the amino-acid change to its confirmation in a sequencing trace.Guided experiment
6. Mouse genotyping by PCR and gel electrophoresisGenotyping of a mouse litter for a knockout or floxed allele by PCR and agarose gel electrophoresis, with blind scoring and a test against Mendelian expectation.Simulator
7. Breeding a conditional knockout lineA Cre–loxP breeding programme planned week by week, from founder crosses to an age-matched cohort with littermate controls, with a count of the surplus animals.Simulator
8. CRISPR–Cas9 genome editingDesign of SpCas9 guides against a simulated gene, editing of cells or mouse zygotes, and validation of knockouts and point mutations by T7E1 assay and amplicon sequencing.Simulator
9. DNA mutation and site-directed mutagenesisTranslation of a cloned gene, classification of mutations, and a defined point mutation taken through primer design, PCR, DpnI treatment, transformation and sequence-verified screening.Simulator

RNA

Methods that measure which genes are expressed, where and in what amount, from RNA extraction to single-cell sequencing.

1. RNA in situ hybridisationSingle-molecule fluorescent in situ hybridisation to identify which neurons in a region contain the mRNA of an excitatory marker, which contain the mRNA of the inhibitory marker <i>Gad1</i>, and whether the two overlap.Guided experiment
2. Measuring gene expression by RT-qPCRThe messenger RNA of one gene is measured in the hippocampus of two groups, from frozen tissue to a statistical comparison: RNA extraction and quality control, reverse transcription, primer validation, the qPCR run, the threshold and melt curve, the choice of reference genes and the analysis.Guided experiment
3. Finding cell types with single-cell RNA sequencingA single-cell RNA sequencing experiment is run on the cortex of control and treated mice, from the design and tissue dissociation to the naming of cell types and a test of whether the treatment changes the share and the gene expression of microglia.Guided experiment
4. Single-cell RNA sequencingSingle cells from mouse cortex, analysed from quality control to cell types, with conditions compared using animals as replicates.Simulator

Proteins and synapses

Detection, measurement and manipulation of proteins, receptors and synapses: Western blotting, receptor pharmacology and synaptic plasticity.

1. Co-immunoprecipitation of synaptic proteinsCo-immunoprecipitation to test whether the scaffold protein PSD-95 and the NMDA receptor subunit GluN2B are associated in brain tissue.Guided experiment
2. Measuring protein concentration by sandwich ELISAMeasurement of the concentration of BDNF in two groups of mice or people by sandwich ELISA, from sample preparation to the interpretation of the standard curve.Guided experiment
3. Counting dendritic spines with Golgi–Cox stainingA test of whether dendritic spines are less dense on pyramidal neurons in one group than in another, by Golgi–Cox staining, blind spine counts and an analysis of nested data.Guided experiment
4. Measuring how strongly a drug acts: concentration–responseThe potency and efficacy of a reference agonist and two coded test compounds are measured on cells that carry their receptor, from weighing the powder to a statistical comparison, and two antagonists are then classified by Schild analysis.Guided experiment
5. Long-term potentiation in a hippocampal sliceSynaptic strength is recorded at the CA3 to CA1 synapses of a hippocampal slice, strengthened with a burst of high-frequency stimulation, and tested for its dependence on NMDA receptors, with the analysis a laboratory would use.Guided experiment
6. Isolating synapses: synaptosome and postsynaptic density fractionationPurify synaptosomes and the postsynaptic density from mouse or human brain tissue, prove with marker proteins what each fraction contains, and ask whether a receptor subunit is redistributed to synapses in one group compared with another.Guided experiment
7. Measuring a protein by Western blottingOne protein is measured in the hippocampus of two groups of mice, from frozen tissue to a statistical comparison: lysis, protein assay and loading, electrophoresis and transfer, antibodies and exposure, blind quantification and analysis.Guided experiment
8. Western blotting: from brain tissue to quantified bandsLysis of brain tissue, a BCA protein assay, SDS-PAGE, transfer, probing and imaging of a simulated protein, and blind quantification of the bands across groups of animals.Simulator
9. Receptor pharmacology in a cell-based assayConcentration–response curves of agonists and antagonists in a 96-well calcium-flux assay, with estimates of potency, efficacy and antagonist affinity.Simulator
10. Synaptic plasticity in hippocampal slicesField EPSPs at CA3 to CA1 synapses, the induction of long-term potentiation or depression, and a test of its dependence on NMDA receptors.Simulator

Cells and circuits

Recording, imaging and control of single neurons and small populations: patch clamp, optogenetics and calcium imaging.

1. Two-photon calcium imaging of visual cortexTwo-photon calcium imaging of the responses of neurons in layer 2/3 of mouse primary visual cortex to gratings of different orientation, from the choice of indicator to the statistics.Guided experiment
2. Primary neuronal culture, immunocytochemistry and Sholl analysisA test of whether a compound changes dendritic branching in cultured neurons, from the coated coverslip to a Sholl analysis with the independent culture as the unit of analysis.Guided experiment
3. Controlling a neuron with light in a brain sliceThe experiment measures how a light-gated protein changes the firing of neurons in an acute brain slice: the opsin is matched to the question, expression is checked, photocurrents and light-driven spikes are recorded, the controls are run and the result is analysed with the animal as the unit.Guided experiment
4. Recording a neuron with patch clampA whole-cell patch-clamp recording is made step by step, as on a real rig: the pipette is prepared, a cell is found and sealed, the quality of the recording is checked, and the firing and ionic currents of the cell are measured and tested with two classic channel blockers.Guided experiment
5. Patch-clamp electrophysiologyThe voltage and ion currents of a single simulated neuron, and the effects of drugs that block its channels.Simulator
6. OptogeneticsExcitation and silencing of neurons with light-gated proteins, and the limits of each tool.Simulator
7. Two-photon calcium imagingImaging of a population of neurons in visual cortex and of the response of each neuron to oriented gratings.Simulator

Tissue

Preparation, staining and imaging of sections of brain and spinal cord to identify which cells are present and which are active.

1. Fluorescence microscopy: c-Fos immunofluorescenceFluorescence microscopy of c-Fos, NeuN, DAPI and a Nissl stain in separate colour channels, in mouse or rat brain, mouse spinal cord or human post-mortem brain.Guided experiment
2. Counting neurons without bias: design-based stereologyThe experiment tests whether a group of brains has fewer neurons than a control group, for example fewer dopaminergic (TH-positive) neurons in the substantia nigra pars compacta of a mouse model of Parkinson's disease, by the optical fractionator with the animal as the unit of analysis.Guided experiment

Brain regions and the whole brain

Measurement of activity in brain regions and across the whole brain: high-density probes, EEG, fMRI and transcranial magnetic stimulation.

1. Recording the P300 with EEGRecording of the P300, the brain’s response to rare target tones, in one volunteer and then in a group, from consent to statistical analysis.Guided experiment
2. Mapping brain activity with fMRIMapping of the brain area that controls finger movement with fMRI, in one simulated volunteer and then in a group, with the participant as the unit of analysis.Guided experiment
3. Recording many neurons at once with a silicon probeThe spikes of several neurons are recorded at once with a high-density silicon probe in a simulated mouse cortex and sorted into single neurons, with every decision from the insertion of the probe to the analysis made at the bench.Guided experiment
4. Measuring motor cortex excitability with TMSTMS is applied to people only by trained staff, under a protocol approved by an ethics committee; every volunteer here is simulated. The session covers screening and consent, EMG from a hand muscle, the hotspot, the resting motor threshold, and the excitability of the motor cortex in one person and then in a group.Guided experiment
5. High-density recording and spike sortingRecording of many neurons with a silicon probe and sorting of their spikes, checked against the known true assignment.Simulator
6. EEG and event-related potentialsBrain responses to rare and frequent sounds, and the averaging that reveals the P300.Simulator
7. fMRI experiment designThe timing of an fMRI experiment and its effect on the power to detect brain activity.Simulator
8. Transcranial magnetic stimulation and motor evoked potentialsA TMS session with simulated participants: screening, hotspot, motor threshold, recruitment curve and paired-pulse inhibition, recorded by EMG.Simulator

Behaviour

Behavioural experiments with groups of animals: random allocation, blinding, tracking and statistical analysis.

1. Testing spatial memory in the Morris water mazeA blinded Morris water-maze study of whether an NMDA-receptor antagonist impairs spatial learning in mice.Guided experiment
2. Morris water mazeA blinded water-maze study of spatial learning in groups of virtual mice, from random allocation and training to the probe trial, exclusions and a mixed ANOVA.Simulator

Computational models

Computer models of neurons and networks, built and tested against data.

1. Building and testing a spiking network modelA computer model of a cortical circuit is built as a modeller builds one: one model neuron is tested, a thousand are connected, the balance of excitation and inhibition is set, the states of the network and their reliability are measured, and the model is compared with published theory.Guided experiment
2. Spiking network modelA network of excitatory and inhibitory spiking neurons, used to test across replicate networks how the balance of excitation and inhibition shapes its activity.Simulator

Cognitive experiments

Twelve classic experiments in perception, attention, learning and memory, in which the visitor takes part as the participant.