4.3. Genetically Modified Organisms (GMO)


Locations:
These laboratories are approved for containment level 2
Laboratory building: Laboratories on the 3rd, 5th, 6th and 8th floors.
BuS: All laboratories
Premises for which approval is granted, are marked with yellow biohazard signs.

 

 

 

Contact person for GMO at the department: siv Lise Bedringaas, e-mail

 

Relevant definitions:

Genetically modified organisms:  Genetically modified organisms are defined as microorganisms (GMM), plants and animals where the genetic composition has been changed through the use of gene- or cell technology.

Microorganisms: Microorganisms are defined as any cellular or non-cellular microbiological unit capable of multiplying itself or of transferring genetic material. The definition of microorganisms includes: Viruses, bacteria, unicellular plants and animals, plant- and animal cells (including human cells) in cultures and microscopic yeast- and mold. The definition does not cover plasmids or other DNA outside of the cell.

Contained use:Contained use is defined as any work operation where genetically modified organisms are created, cultured, stored, destroyed or used in other ways, in a closed system where physical barriers are used in order to limit the organisms’ contact with humans and the environment. Transport of genetically modified organisms between approved laboratories within the same institution, for example, other laboratories within a university area, is also considered encased use.

Deliberate release: Deliberate release is defined as any creation and use of genetically modified organisms not classified as contained use.

Accident: Any incident involving accidental release of GMM upon confined use and which may cause immediate or consequential danger to human and / or animal health and / or the environment,

 

Laws and Regulations

For GMO work, we must comply with the Gene Technology Act and its regulations, which require reports / approvals of both projects and laboratory facilities. The act also stipulates requirements for how the work is carried out.

 

Use of GMO  at K2

Genetically modified microorganisms (GMMs) and -animals are research methods used by the institute.

Examples of GMM may be the cultivation of plasmids in bacteria or gene transfection of cell lines using viral vectors.

Examples of transgenic animals may be mice which have inserted RNA or DNA, or knock-out mice where a gene is turned off.

 

When conducting day-to-day GMO work in the laboratory there are four things one must adhere to:

  1. That the laboratories and facilities are approved for work with GMO. At present K2 has approved laboratories for working with GMM at containment – / Biosafety  level1 and  2. Approved areas are marked with the yellow Biohazard signs informing of this.
  2. That there exists reports/approval for the work you shall carry out. The leader of the relevant project is responsible for this. The project leader is also responsible for ensuring that those working on the project are informed that the work contains GMM and what precautions and work routines to follow. In the report/approval there should be attached a preliminary assessment of use with regard to risk of disease/injury to humans, animals, plants or damage to the environment.
  3. That work is conducted in an appropriate manner according to safety guidelines (see below).
  4. That a work journal is kept. Document applications regarding use of GMO and GMM and record procedures so that they may be produced upon request/for inspection.

For all activities involving GMM, the principles for good microbiological practice regarding good safety and hygiene in the work place should be followed.

 

Training and competence

Everyone who works with GMO must have necessary and documented training. This implies that one must have knowledge of:

  • Laws and regulations.
  • Cell culture and sterile technique.
  • Risks associated with the GMO work.
  • Waste handling.
  • How to respond to spill / accident.

The GMO course at the faculty is mandatory. First course in English will be held in spring 2019.

 

 

Personal Protective Equipment

  • When working with viral vectors use only the Biosafety cabinet/LAF-bench dedicated for this use.
  • Use proper protective coat (one that protects the front part of the body) and glows (A double set of glows or “Touch N Tuff” is recommended).
  • Protect yourself with plaster if you have any wounds or scratches on your hands.
  • Use blue shoe cover.
  • Make a safety and spill kit that contains equipment needed for spill or accidents.

 

Containment level and Risk class

The approved laboratories at K2 have authorization for work with GMO at containment levels 1 and 2. In these classes, the risk of harm to humans, animals or the environment is considered to be insignificant or small. The requirements for the building (laboratories), waste management, internal control and emergency preparedness are intensified by increasing risk classification. See GMM Regulations Appendix 1 A and C.

To determine the containment level, the properties of GMM will be assessed by the ability of the following factors to cause disease in humans, animals, or plants:

  • Recipient microorganism
  • Parent microorganism
  • Vector
  • Gene insert
  • GMM

Microorganisms that most likely do not cause disease are classified as risk class 1. See GMM regulations Annex III, point B (4).

For other risk classes, see List of classified biological factors (contagious groups) to assess the level of occupational exposure to GMM.

Transport

For GMM in risk class 1 and 2, transport and import (shipments) of up to 10 liters is permitted without any special approval as long as the regulations for labelling and packaging are followed.

 

When transporting GMM between different approved units/rooms make sure that this is carried out in a way that minimizes the probability for accidents/spills.

Transport of risk class 1 or 2 requires:

  • Inside and an outer container, both must be waterproof.
  • Absorbent material between inner and outer container.
  • The container should be transported on a trolley.

Transport of GMM in higher risk classes, and more than 10 liters in risk class 2 will require more substantial approval and is not covered here.


Protocol: All transport and import must be recorded and the following should contain the following:

  • Type GMO
  • Date of shipment and receipt
  • Acceptance inspection
For more information regarding transport, see the regulation for GMO  labelling etc. (No).

 

Waste handling

Actions that are mandatory by law with contained use of GMO / GMM:

Prevention of emissions: GMM should be inactivated through the use of recognized methods for waste disposal and emissions, including waste water. If necessary, check whether or not viable organisms outside the primary physical encasing come into being. Here one has to use incineration, autoclaving or other disinfection methods as outlined on the page  “General work procedures“.

Antibiotic resistance genes should be handled so that these genes are destroyed, for example through fragmenting prior to emission into the surroundings. Incineration is one way of ensuring this, but autoclaving also causes gene inactivation and is best for large volumes.

Approved methods of inactivation are:
  • Virkon, (Incubate for about 12 hours) in combination with
  • burning, or 
  • autoclaving (autoclaving alone is not always enough to fragment plasmid DNA

 

Spill of liquids or other substances containing GMO

Depending on the amount and place of spill, assess the incident with respect to decontamination, evacuating the room and reporting.  Everyone should be familiar with the lab’s spill response.

 

Recommended Action List:

  1. Let the Biosafety cabinet continue to operate
  2. Decontaminate the surface by gently cover the spill with absorbent paper towels sprayed / soaked in Virkon. Allow sufficient decontamination time (approximately 20 minutes) before clean up.
  3. Clean the surface with Antibac (or 70% ethanol ) and discard soaked paper towels in a biohazard waste bag in the cabinet.

With major spills (> 25 ml): Firstly, evacuate the room, close the door and remove any contaminated clothing including laboratory coat, Wash face and hands, and post a warning sign.

With severe spills (Outside the biosafety cabinet): Firstly, put on a mask to minimize the chance of inhaling the aerosol, alert anyone in the vicinity, ascertain extent of spill, wash your hands and face thoroughly using soap and disinfectant.  Notify the PI or the laboratory (research)h manager.

Reporting accidents/spills

Major spill (> 25 ml inside the LAF bench or <10 ml outside the LAF bench):

The spill must be reported to:

  • The PI
  • HSE Department (non-conformities form)

 

Any cases of accident or severe spill (> 10 ml) of GMOs outside the biosafety cabinet that might cause danger to human or animals health and/or environment.

(c.f. The GMO regulation § 19)

The spill must be reported to:

The following information must be provided:
1. Detailed circumstances of the accident.
2. Identity and amount of GMM released.
3. All information necessary to assess the impact of the accident on health and the environment.
4. Which actions have been effectuated.

The project or group leader are responsible for submitting the incident report to the NDH.

 

Waste disposal

Virus culture: Liquid waste, solid waste such as pipettes, tips, tubes, dishes etc. that have been in contact with virus must be immersed completely in a Virkon waste container for approximately 12 hours. Dispose the waste according to internal instructions.

  • After incubation, liquid (disposed into a liquid-waste bottle/container) and solid waste shall be disposed into cardboard boxes for bio waste labeled “Prøverør/Agarskål” and burned.
  • Alternatively, all fluids and solid waste may be autoclaved immediately after the work ends.

Bacterial culture:

  • All bacterial waste should be autoclaved.
  • All reusable centrifugation tubes / other equipment should be autoclaved and washed in the dishwasher.

 

Madatory documentation

 

  • Risk assessment of all GMOs including own work.
  • Procedures: There must be available information about safety, waste management and work operations.
  • Journal of all work with GMM.
  • Registration of all biological factors including GMM in EcoOnline.
  • Applications and approvals for the use of GMOs should be kept collectively and readily available.

 

Risk of infectious transmission using retroviral / lentiviral vectors (viruses)

Infection from needle sticks and cuts causes the highest risk factor for transmission of infection when working with Lentiviral vectors in the laboratory. The other form of exposure is from airborne aerosols via the respiratory tract, for example due to spills or to hard pipetting.

Exposure can lead to infections with transmission of viral genetic material that may result in:

  • mutations
  • development of oncogenesis 
  • generation of replication competent lentivirus (RCL)

 

Links: