Cybertory: virtual molecular biology lab

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Sequence analysis - Direct link to the tools (transcription-translation, sequence pattern search, sequence massager)

General instructions for the use of Cybertory.

Features:

• Digestion of DNA with restriction enzymes (81 enzymes available).
• PCR amplification by multiplex PCR of DNA segments that include STR polymorphic markers from CODIS (6 available) and a sex marker.
• PCR amplification by multiplex PCR of several polymorphic markers and species-specific sequences.
• Electrophoresis of DNA fragments on agarose or polyacrylamide gel and ethidium bromide staining.
• DNA sequencing using dideoxynucleotides and electrophoresis. Label with either a radioisotope, a fluorochrome or 4 fluorochromes.

Possible applications:

• Molecular diagnostics with RFLP or PCR of genetic polymorphism.
• Forensic analysis of DNA.
• Paternity test using multiplex PCR of polymorphic STR genetic markers.
• Molecular analysis of foodstuff adulteration.
• Restriction mapping.

Workbooks:

	1A	1B	1C	1D	2A	2B	3	4	5	6	7	8	9
using electrophoresis
using restriction enzymes
using PCR

General instructions for the use of Cybertory.

1. Genetic diagnosis; β^A/β^S polymorphism in the beta globin gene, responsible for drepanocytosis and drepanocytic anaemia (sickle cell anaemia):

   1A		Testing restriction enzymes for the analysis of this polymorphism by RFLP. doi:10.5281/zenodo.1299796
   		Some enzyme sets suitable for this practical.
   1B		Molecular genetic diagnosis of the polymorphism by RFLP. doi:10.5281/zenodo.1299811
   1C		DNA sequencing to analyse this polymorphism (dideoxynucleotides method). doi:10.5281/zenodo.1299816
   1D		Introduction to genomic sequence databases and analysis of sequences related to this polymorphism. doi:10.5281/zenodo.1299825

2. Forensic analysis: genetic identification of individuals:

   2A		Using RFLP. doi:10.5281/zenodo.1299843
   2B		Using multiplex PCR on STR markers (CoDIS). doi:10.5281/zenodo.3607339

3. Paternity test using multiplex PCR on STR markers (CoDIS). https://doi.org/10.5281/zenodo.1299837
4. Genetic diagnosis, personalised medicine. Identification of polymorphic variants on cytochrome P450 (CYP2C9), using multiplex PCR. doi:10.5281/zenodo.1299846
5. Genetic diagnosis of celiac disease. Identification of polymorphic variant associated to celaic disease, using multiplex PCR on the DQ2 and DQ8 markers. doi:10.5281/zenodo.1299854
6. Dairy product analysis. Identification of the source of milk (cow, sheep or goat) by analysis of mitochondrial DNA. doi:10.5281/zenodo.1299860
7. Vegetable oil analysis. Identification of a poossible adulteration of olive oil with other specis by analysis of chloroplast DNA. (not yet available)
8. Viral infection test. Identification of infection by coronavirus or by common flu virus using multiplex PCR on cDNA obtained from samples. doi:10.5281/zenodo.3730995
9. Genetic diagnosis. Detection of a mutation in protooncogene RET causing multiple endocrine neoplasia, type 2A (MEN2A). doi:10.5281/zenodo.5168483

Simulation of celulose acetate electrophoresis of proteins

Features:

• Electrophoresis of either serum proteins or LDH isoenzymes on cellulose acetate strips.
• Only the final result is displayed.
• Reveal serum proteins by staining with Coomassie Blue, Amido Black or Ponceau Red, or by autoradiography.
• Reveal LDH with an enzyme assay.
• A single sample per run.
• Predefined profiles, both normal and pathological, or custom profile depending on given proportion of each component.
• Automatic plot of both the band pattern on the cellulose acetate strip and its densitogram. Possibility to overlay successive densitograms in the same plot for comparison.

Possible applications:

• Clinical interpretation of serum protein profiles.
• Differential tissue profile of LDH isoenzymes.
• Clinical interpretation of LDH isoenzyme profiles.

Access:

Access the simulator for serum proteins and for LDH isoenzymes.

Simulation of SDS-PAGE electrophoresis of proteins

Features:

• Electrophoresis of proteins on a polyacrylamide gel with SDS; direct detection.
• Several acrylamide concentrations are available: 7.5; 10; 12 and 15%.
• 4 voltages are available to run the separation.
• One lane for standards and another for a sample.
• 7 standard proteins of known size and 10 unknown samples (with a single protein each).
• The progress of the separation is displayed.
• Automatic drawing of the calibration line and aided interpolation.

Possible applications:

• Estimation of the molecular mass of the unknown proteins.

Workbooks:

These are integrated into the simulation page:

Practice problem 1: influence of the voltage applied.

Practice problem 2: influence of the acrylamide concentration.

Practice problem 3: estimation of molecular mass of the unknown proteins.

Practice problem 4: determination of quaternary structure (I).

Practice problem 5: determination of quaternary structure (II).

Access:

Acces the simulator.

UV-VIS spectrophotometry laboratories

Simulator of ultraviolet absorption spectra for proteins and nucleic acids

Access:

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Features:

• Protein and DNA absorption spectrum between 230 and 340 nm.
• Absorbance values at 260 and 280 nm, and the ratio between both.
• Predefined profiles: proteins, DNA and four mixtures.
• Custom profiles based on proportion provided for each type of molecule.
• Possible to superimpose sucessive spectra to compare them.

Applications:

• Demonstration of the analytical value of the technique for checking DNA contamination with proteins or vice versa.

Virtual UV-VIS spectrophotometer

Access:

Access the virtual laboratory.

Features:

• Absroption spectrum of diverse substances between 200 and 800 nm.
• Preparation of dilutions.

Applications:

• Relation between absorbance and concentration.
• Fundamentals of a protein quantitation assay.
• Determination of absorption spectra; search for λ_max

Colorimetric assay for enzyme activity

Access:

Access the virtual laboratory.

Features:

• Enzyme activity assay.
• Effect of pH and temperature on the activity of an enzyme.

Applications:

• Search for optimal conditions for the enzyme.

Simulation of column cromatography of proteins

Features:

• Several types of stationary phase, for:
  • molecular exclusion (gel filtration)
  • anionic and cationic exchange
  • affinity
• Analisis of samples made of 3 proteins.
• Choose among 13 predefined proteins or proteins with user-provided properties (M_r, pI).
• Shows progress of the protein bands along the column as well as así recording of chromatogram (absorbance versus elution volume).

Possible applications:

• To illustrate the mechanism of separation on each type of matrix. Effect of pH on ion exchange.
• Assay of conditions for resolution of proteins in a sample (for example, for analysis or for purification).

Access:

Access the simulator.

Working plans:

Project: design of problem-solving activities using the simulator.

Other labs

These are ready but not yet translated into English. You can check the Spanish versions or, if you are interested on them, for setting an English translation.

• More UV-VIS spectrophotometric assays:
  • Forms of hemoglobin.
  • Plant pigments.
  • With calibration curve: glucose, proteins (Lowry), creatinine.
  • Kinetic measurement: gamma-glutamyltranspeptidase.
  • MTT metabolic viability (on 24-well plates)
  • Ketone bodies (on reactive strips)
• Circular dichroism spectra of proteins.
• Ion exchange chromatography to measure % HbA1c.
• Thin-layer chromatoraphy (amino acis, lipids, plant pigments)