Risk Assessments

Here you can find guidelines for the most common risks assosiated with the protocols used in the course of our experiments and the specific risk assessments for each of the protocols listed on the wiki.

Risk Assessments

β-galactosidase Assay

Using Biobricks from the distribution plates

Colony PCR

Chemically competent cell - generation

Electrocompetent cell - generation

Electroporation

Gel electrophoresis

Gel extraction of DNA

Gibson Assembly

Glycerol Stocks

IPTG Induction

LB agar plates - preparation

Miniprep - DNA extraction

PCR using high temperature DNA polymerase

SDS PAGE protein analysis

Restriction Enzyme Digest

Transformation of Bacillus subtilis

Transformation of Escherichia coli

Use of Sodium Fluoride for construct testing

General Risks

It is expected that good laboratory practice will be observed and that the correct PPE (personal protective equipment) will be worn at all times. Further precautions must be taken however in light of some hazards.

Bacteria- Depending on the bacteria used in your experiments, the danger associated with exposure will vary. Our laboratory is only authorised to use Biosafety level 1 (non-pathogenic) bacteria and so in our experiments the risk to scientists is minimal. It is however the policy of the department to treat all bacteria as pathogens. This will be increasingly important if pathogenic strains of bacteria are used.

Infection - the risk of infection in our experiment was not of great concern as non-pathogenic strains were used. Gloves and lab coats were still always worn when working with bacteria (this was also to help prevent contamination of samples). Where there is a greater risk masks should also be work to prevent access via the mouth or airways. Despite wearing gloves while working with bacteria, scientists must still wash their hands before leaving the lab to eliminate the chance of contaminating food.

Disposal - Unused bacteria and any material that has come into contact with the bacteria (including gloves)should be disposed of in the biological waste vessels in the laboratory. When full, these materials are then autoclaved and disposed of in accordance with normal departmental procedures.

Cold - The extreme cold used to store the competent cells until use is capable of causing freeze burns. If prolonged exposure to these temperatures is required the wearing of heat proof gloves is recommended.

Gel preparation - the TAE buffer and agarose are not inherently dangerous but to be effectively mixed together they need to be heated, usually in a microwave. To fully dissolve the agarose, the mixture must be heated to a temperature too hot to touch. The container must therefore be handled with a heat proof glove.

There is also a danger that the mixture could superheat and this is extremely dangerous. A mixture that does not appear to be boiling when taken out of the microwave can boil violently when swirled. The mixture should therefore be heated gradually, in small bursts and then swirled at arms length to heat to the minimum possible temperature to completely dissolve. The lid of the container must be on, but not screwed tight as tight as possible to allow for gas expansion as the mixture heats up.

Glass - Glass is somewhat ubiquitous in laboratories and though it is expected that scientists will work safely, accidents still happen. All glassware in use should be labelled so that in the case of a breakage the carried substance can be identified. Broken glass is obviously a hazard through the liklihood of injury and it should be cleared away as soon as possible. The contents of the vessel will often be of greater concern however. Post breakage procedures will vary in accordance with what was in the container before the break and scientists should familiarise themselves with the disposal measures of their department.

Hazardous Chemicals - some of the chemicals used in these protocols carry specific health risks. These will be listed on an individual basis in the relevant risk assessments below but in all cases it is worth noting that additional PPE may have to be worn and that normal waste disposal procedures may not be appropriate.

Heat- While most PCR is now carried out using specialised machines, it is possible to perform PCR by hand using heat blocks and waterbaths. The high temperatures used in this technique are sufficient to burn the skin so care must be taken when using machinery and transferring samples. The use of tongs or heat proof gloves is recommended in this case.

Voltage - The Genepulse device administers a very high voltage and therefore the electroporation cuvette should only be used with the device in the manner described by the manufacturer. This will normally involve the use of a plastic loading device which connects the cuvette to the electrodes prior to electroporation. If in doubt, seek advice before connecting the cuvette to the device.