OHS017
OHS Risk Assessment and
Control Form
Risk assessment completed by: Kylie Taylor
Staff/student number: 3294023
Faculty/Division: Medicine
Document number
SOMS.CGM.RA029
School/Unit: School of Medical Sciences
Initial Issue date
30/06/09
Current version
1.0
Current Version
Issue date 30/06/09
Next review date
30/06/12
For additional information refer to the OHS Risk Assessment and Control Procedure, the OHS Risk Rating Procedure and the Hierarchy of Risk Controls.
Risk Assessment title::Purification of DNA using PureYield Plasmid Miniprep System (Promega).
Step 1: Identify the activity
Procedure adapted from the Plasmid Miniprep System Handbook. Consult this handbook before using Miniprep kit.
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Prepare an appropriate volume of LB broth .For 1 L of LB dissolve 10 g of tryptone and NaCl and 5 g of yeast extract in 1 L MilliQ water and autoclave.
Allow the media to cool to at least 55°C before inoculating 5 mL of the prepared LB broth with the appropriate antibiotic – normally, 5 µL of 10 mg/mL stock in 5 mL of culture gives an antibiotic
concentration of 10 µg/mL.
Directly inoculate the media by picking a single colony from an agar plate with a sterile toothpick or pipette tip and placing this directly into the media and growing the cells overnight (12 – 16 hrs)
at 37° with vigorous shaking.
For low copy number plasmids, collect cells from up to 3 mL of culture by spinning down in microcentrifuge for 30 s at maximum speed and resuspending the pelleted cells in 600 µL of water or TE
buffer. For high copy number plasmids take 600 µL of bacterial culture.
Add 100 µL Cell Lysis Buffer and mix thoroughly by inverting the tube until the solution is a uniform blue colour.
Add 350 µL cold (4 - 8°) Neutralisation Solution and mix thouroughly by inverting the tube until the solution is a uniform yellow colour.
Centrifuge at maximum speed in a microcentrifuge for 3 mins.
Pipette the supernatant into a PureYield Minicolumn without disturbing the pelleted cell debris.
Place the column into a PureYield Collection Tube and centrifuge at maximum speed in a microcentrifuge for 15 s.
Discard the flowthrough and return the column to the same collection tube. Add 200 µL Endotoxin Removal Wash and centrifuge at maximum speed in a microcentrifuge for153s.
Add 400 µL of Column Wash Solution to the column and centrifuge at maximum speed in a microcentrifuge for 30 s.
Transfer the column to a clean 1.5 mL microcentrifuge tube and add 30 µL Elution Buffer or nuclease-free water to the column.
Incubate for 1 min at room temperature.
Centrifuge at maximum speed in a microcentrifuge for 15 s and store the eluted DNA at -20°.
Describe the location: Room 501, Level 5, Wallace Wurth Building, University of New South Wales.
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Page 1 of 5
Risk Assessment and Control Form
Date Effective: 01/01/2007
Uncontrolled document when printed
Current Version: 2.6, 16/07/2008
Step 2: Identify who may be at risk by the activity
A number of people may be at risk from any activity. This may affect the risk controls needed. These people may include fellow workers, visitors, contractors and the public. The location of the activity
may affect the number of people at risk.
Laboratory staff.
Steps 3 to 7: Identify the hazards, risks, and rate the risks
1. An activity may be divided into tasks. For each task identify the hazards and associated risks.
2. List existing risk controls and determine a risk rating using the UNSW Risk Rating Procedure.
3. Additional risk controls may be required to achieve an acceptable level of risk. Re-rate the risk if additional risk controls used.
Tasks
Hazards
Associated risks
(Step 3)
(Step 4)
Risk rating with existing
controls *
Additional risk controls
required
Risk Rating with
additional controls *
(Step 5)
(Step 6)
(Step 7)
Existing risk controls
C
Preparation of LB media
Culturing of bacteria
Autoclaving media
Bacteria
Orbital incubator
Burns from spilling hot
media or handling hot
glassware
Autoclave media in a
sealed vessel.
Contamination by
bacteria
Culture cells in flasks
with a cap.
Injury caused by
shaker-incubator
Always stop shaker
before opening the
incubator.
L
R
2
D
Low
2
E
Low
(Apply the hierarchy of
risk controls)
C
L
R
Allow glassware to cool
before handling; use
insulating gloves and
carry-baskets to transport
media.
Ensure all users are
properly trained in the
correct use of the
autoclave.
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Risk Assessment and Control Form
Date Effective: 01/01/2007
Uncontrolled document when printed
Current Version: 2.6, 16/07/2008
Collection of bacterial
cells
Lysis of bacterial cells
Disposal of contaminated
media and plates
Centrifuge
Chemicals in lysis
buffers-
Bacteria
Bleach or antiseptic
solution
Injury caused by
incorrect use of
centrifuge
Irritation to skin or eyes
Contamination by
bacteria
Train all users in
correct use of
centrifuge.
Check that the rotor is
in the correct
orientation.
Balance centrifuge
before use.
Close lid of centrifuge
Wear
gloves and safety
tightly.
glasses when handling
(See ‘Centrifuge Use’
buffers
SWP and RA)
3
D
Medi
um
1
D
Low
Wear gloves and safety
glasses when handling
chemicals and
contaminated media.
Dispose of contaminated
plates via the autoclaved
waste stream.
2
D
Low
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Page 3 of 5
Risk Assessment and Control Form
Date Effective: 01/01/2007
Uncontrolled document when printed
Current Version: 2.6, 16/07/2008
Step 8 Documentation and supervisor approval
Completed by: Kylie Taylor
(signature)
Authorised by: Peter Gunning
(signature)
Date:16/10/2009
Step 9: Implement the additional risk controls
identified
Indicate briefly what additional risk controls from Step 6 above were implemented, when and by whom.
Risk control:
Date:
Implemented by:
Risk control:
Date:
Implemented by:
Risk control:
Date:
Implemented by:
Risk control:
Date:
Implemented by:
Step 10: Monitor and review the risk controls
It is important to monitor risk controls and review risk assessments regularly. Review is required when there is a change in the process, relevant legal changes, and where a cause for concern has
arisen. Reviews could be scheduled on an annual basis. If the risk assessment has substantially changed a new risk assessment is warranted.
Review date:
Reviewed by:
Authorised by:
Review date:
Reviewed by:
Authorised by:
Review date:
Reviewed by:
Authorised by:
Review date:
Reviewed by:
Authorised by:
Review date:
Reviewed by:
Authorised by:
Documentation
It is a requirement that legal and advisory documentation that supports this risk assessment be listed. Such documentation includes Acts, Regulations, Australian Standards and Codes of Practice,
where applicable.
NSW OHS Act 2000
NSW OHS Regulation 2001
Code of Practice for the Labelling of Workplace Substances
AS/NZS 2243.2:2006. Safety in laboratories. Part 2: Chemical aspects
Australian Standard AS2243.3-2002. Safety in laboratories. Part 3: Microbiological aspects and containment facilities.
Australian Standard AS2243.6-1990. Safety in laboratories. Part 6: Mechanical Aspects.
Australian Standard AS2243.7-1991. Safety in laboratories. Part 7: Electrical Aspects.
AS/NZS 2161.1:2000 Occupational Protective Gloves – Selection, Use and Maintenance
AS/NZS 1336:1997 Recommended Practices for Occupational Eye Protection
UNSW Hazardous Waste Disposal Procedure
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Page 4 of 5
Risk Assessment and Control Form
Date Effective: 01/01/2007
Uncontrolled document when printed
Current Version: 2.6, 16/07/2008
UNSW Concise OHS Risk Rating Table
OHS697
What you need to do
1. Consider what can go wrong that can hurt someone
2. Determine what the most likely outcome would be - Consequences
3. Determine how likely those consequences are - Likelihood
4. Calculate the risk rating
5. Required action
How severely could someone be hurt
death or permanent disability to one or more persons
hospital admission required
medical treatment required
first aid required
injuries not requiring first aid
CONSEQUENCES:
Severe
Major
Moderate
Minor
Insignificant
How likely are those consequences?
expected to occur in most circumstances
will probably occur in most circumstances
could occur at some time
is not likely to occur in normal circumstances
may occur only in exceptional circumstances
LIKELIHOOD:
Almost certain
Likely
Possible
Unlikely
Rare
CONSEQUENCES
Insignificant
1
Minor
2
Moderate
3
Major
4
Severe
5
M
H
H
VH
VH
M
M
H
H
VH
Possible
C
L
M
H
H
VH
Unlikely
D
L
L
M
M
H
Rare
E
L
L
M
M
M
LIKELIHOOD
Almost
certain
A
Likely
B
Risk level
Very high
High
Medium
Low
Required action
Act immediately:
The proposed task or process activity must not proceed. Steps must be taken to lower the risk level to as
low as reasonably practicable using the hierarchy of risk controls.
Act today:
The proposed activity can only proceed, provided that:
(i) the risk level has been reduced to as low as reasonably practicable using the hierarchy of
risk controls;
(ii) the risk controls must include those identified in legislation, Australian Standards, Codes of
Practice etc.
(iii) the risk assessment has been reviewed and approved by the Supervisor and
(iv) a Safe Working Procedure or Safe Work Method has been prepared.
(v) The supervisor must review and document the effectiveness of the implemented risk
controls.
Act this week:
The proposed task or process can proceed, provided that:
(i) the risk level has been reduced to as low as reasonably practicable using the hierarchy of
risk controls;
(ii) the risk assessment has been reviewed and approved by the Supervisor and
(iii) a Safe Working Procedure or Safe Work Method has been prepared.
Act this month:
Managed by local documented routine procedures which must include application of the hierarchy of
controls.
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Page 5 of 5
UNSW Concise OHS Risk Rating Table
Effective date: 01/01/2007
Uncontrolled document when printed
Current Version: 2.6,16/07/2008