Team:Alberta/Safety

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            METHODOLOGY
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<h2>Team Alberta iGEM Safety Proposal</h2>
 +
 
 +
Team Alberta recognizes the important role that safety plays in the
 +
completion of any scientific undertaking. Our team members are fully
 +
committed to not only ensuring the safety of those within our lab but
 +
also addressing any potential risks that our project may pose to the
 +
greater community.
 +
 
 +
<br>
 +
 
 +
Please find below Team Alberta's answers to the new, required iGEM
 +
safety questions.
 +
 
 +
<h3 id=Section_1>1. Would the materials used in your project and/or your final
 +
product pose:</h3>
 +
<h4>a. Risks to the safety and health of team members or others in
 +
the lab?</h4>
-
<div id=highschool4>
+
<div class=safety-img id=labelledflask>
-
[[Image:Manual.jpg|280px|thumb|right|Information about reagents and safety protocols
+
<img src="https://static.igem.org/mediawiki/2011/8/8a/TeamAlberta_labelledflasks.jpg">
-
are all found in an easily accessible binder]]
+
Neurospora is stored in clearly labelled flasks with sponge tops to prevent contamination
</div>
</div>
 +
<p>The materials used in our project pose little to no risks to the
 +
safety and health of our team members or others in the lab and our
 +
final product poses no risk to these individuals.</p>
-
{{Team:Alberta/endLeftSideBar}}
+
<br>
-
{{Team:Alberta/beginRightSideBar}}
+
<p>The organism that our group has genetically manipulated,
-
[[Image:Alberta_Waste.jpg|280px|thumb|left|Clearly labelled waste disposal containers ensure
+
<i>Neurospora crassa</i>, is well-documented as being safe<sup>1</sup>. N. crassa is
-
proper removal of potentially hazardous materials]]
+
a biosafety level 1 organism. Since 1941 the species has been used
 +
extensively in laboratories resulting in the publication of thousands
 +
of research papers. In none of these numerous publications has the
 +
<i>Neurospora</i> genus been implicated in causing disease in either animals
 +
or plants<sup>1-4</sup>.</p>
-
[[Image:TeamAlberta labelledflasks.jpg|280px|thumb|left|Neurospora is stored in clearly
+
<br>
-
labelled flasks with sponge tops to prevent contamination]]
+
-
{{Team:Alberta/endRightSideBar}}
+
<p><i>Neurospora's</i> minimal threat can largely be attributed to it being
 +
an obligate aerobe. Unlike other species such as yeasts, <i>Neurospora</i>
 +
is unable to grow in organs such as the gut or bladder, tissues, or
 +
systemically within an organism<sup>1</sup>. Furthermore, despite <i>Neurospora</i>
 +
being regarded by some as an allergen, medical literature generally
 +
fails to mention the genus<sup>5-8</sup>.</p>
-
<html><div id="center-content" style="margin-top:120px;" class="</html>{{{class|}}}<html>"></html>
+
<br>
 +
<p>Our team acknowledges that though certain fungal species
 +
themselves may not be pathogenic, they may produce dangerous
 +
toxins. Throughout our research into Neurospora our group has
 +
not found any documentation of the species producing dangerous
 +
mycotoxins or any dangerous secondary metabolites<sup>1</sup>. Given that
 +
Neurospora has had such a long history of living in close association
 +
with humans (please see below), it is a generally regarded as being
 +
harmless.</p>
 +
<div class=safety-img id=waste>
 +
<img src="https://static.igem.org/mediawiki/2011/2/25/Alberta_Waste.jpg">
 +
Clearly labelled waste disposal containers ensure proper removal of potentially hazardous materials
 +
<br>
-
<h1>Team Alberta iGEM Safety Proposal</h1>
+
</div>
-
<div id="horiz-line"></div>
+
-
<p> Team Alberta recognizes the important role that safety plays in the completion of any scientific undertaking. Our
+
<p>It should be noted that there are still materials used in our
-
team members are fully committed to not only ensuring the safety of those within our lab but also addressing any potential risks that our project may pose to the greater community. </p>
+
project that may potentially be hazardous. Our esterification procedure
 +
involves the use of strong chemicals and reagents, some of which have
 +
the potential to cause harm. These reagents are primarily volatile and
 +
require careful handling and storage.</p>
 +
<br>
-
<h1>1. Safety</h1>
+
<p>As well, Hygromycin b, an antibiotic and antifungal agent that
-
<div id="horiz-line"></div>
+
is used for selection, is toxic if inhaled. Care must be taken around
 +
powdered or lyophilized Hygromycin B, and precaution should be taken
 +
to cover up respiratory passageways, skin, and the eyes. However,
 +
Hygromycin B is most commonly purchased in liquid form, and in liquid
 +
carries with it significantly less potential for accidental inhalation.</p>
-
<h2>1.1 Neurospora crassa</h2>
+
<h4>b. Risks to the safety and health of the general public if
 +
released by design or accident?</h4>
-
<p> It is well documented that the organism that our group plans to genetically manipulate, Neurospora crassa, is safe<sup>1</sup>. Since 1941 the species has been used extensively in laboratories resulting in the publishing of thousands of research papers. In none of these numerous publications has the Neurospora genus been implicated in causing disease in either animals or plants<sup>1-4</sup>. </p>
+
<p>Our organism poses no threat to the safety and health of the
 +
general public. Neurospora has a well-documented history of living in
 +
close association with humans. It has historically resided in a variety of
 +
businesses including bakeries, lumber yards and plywood factories<sup>1;9-
 +
11</sup>. Growth has also been observed on the stubble of burnt sugar cane
 +
fields and along burnt railway tracks<sup>13</sup>. Thus, it poses no threat to the
 +
health and safety of those in its surroundings.</p>
-
<p>Neurospora’s minimal threat can largely be attributed to it being an obligate aerobe. Resultantly, unlike other species such as yeasts, Neurospora is unable to grow in organs such as the gut or bladder, tissues, or systemically within an organism<sup>1</sup>. Furthermore, despite Neurospora being regarded by some as an allergen, medical literature generally fails to mention the genus<sup>5-8</sup>. </p>
+
<h4>c. Risks to environmental quality if released by design or
 +
accident?</h4>
 +
<p>As mentioned, N. crassa has a long history of living in close
 +
association with humans; however, these instances are largely
 +
documented as having occurred in moist tropical or subtropical
 +
environments. Like many fungi, warm and moist is the favored climate
 +
for Neurospora growth<sup>1</sup>. Thus, if our organism, WT or manipulated,
 +
were to be released in our dry, temperate environment of Edmonton,
 +
Alberta, Canada, with its bitterly harsh winters, we would expect its
 +
survival and environmental impact to be minimal. Even if it were to
 +
survive, we cannot see it affecting local ecosystems by any appreciable
 +
amount.</p>
-
<p>Our team acknowledges that though certain fungal species themselves may not be pathogenic, they may produce dangerous toxins. Throughout our extensive research into Neurospora our group has not found documentation of the species producing dangerous mycotoxins or any dangerous secondary metabolites<sup>1</sup>. Given the Neurospora has had such a long history of living in close association with humans (please see below) the aforementioned facts are even moreso compelling<sup>1</sup>.</p>
+
<br>
-
<h2>1.2 Reagents and Chemicals</h2>
+
<p>Nonetheless, our alternative biodiesel has the potential to
 +
revolutionize economies throughout the world, as discussed in our
 +
human practices component, and we must therefore regard the effects
 +
of environmental exposure to Neurospora in other climates. Were
 +
Neurospora to be released in other warmer climates it may readily
 +
survive, creating its own niche within the environment. However, the
 +
aforementioned nonpathogenicity of the genus results in this not being
 +
a source of alarm<sup>1-4</sup>.</p>
 +
<br>
-
<p>In light of our procedure involving the use of many reagents and chemicals, each of our team members has received safety training (ex. WHMIS, etc.) on each of the materials we will be using. Moreover, data sheets for each of these materials are stored in an organized binder that is readily accessible to each team member. </p>
+
<p>Further, Team Alberta predicts that our manipulated strain
 +
of Neurospora crassa would be disadvantaged to survive relative
 +
to WT in any environment; our genetic manipulation results in the
 +
organism's energy being diverted away from essential metabolic
 +
functions, inhibiting organismal growth and manipulation. This puts it
 +
at a disadvantage against natural competing fungi.</p>
-
<p>Our overall experimental procedure involves many steps. Most notably, the esterification portion of our procedure involves the greatest use of chemicals and reagents. We have therefore greatly scrutinized all steps within this procedure for potential hazards and have appropriately addressed all concerns (please see the outline below). </p>
+
<h4>d. Risks to security through malicious misuse by individuals,
 +
groups or states?</h4>
-
[[Image:TeamAlbertaReagentsAndChemicals.png|500px]]
+
<p>Unforeseen alteration of biopart functions may represent issues
 +
of concern. However, in our case if one or several of our bioparts were
 +
changed to alter their function or stop working as intended, limited to
 +
no safety issues would arise. Our knockout of the FadD gene results
 +
in there being limited potential for its alteration of function. Further,
 +
our manipulation of thioesterases similarly represents limited safety
 +
concerns due to their prevalence in all organisms. If further alterations
 +
were to be made to thioesterase genes and resulted in overproduction,
 +
our organism would likely die or have a limited existence; if further
 +
alterations to thioesterase genes were to prevent their function, then
 +
the organism may return to a more WT phenotype<sup>12</sup>.</p>
 +
<br>
-
<h2>1.3 Environmental Exposure</h2>
+
<p>The misuse of our bioconverter represents a potential threat
 +
to safety. The esterification process requires heat and flammable
 +
chemicals contained within a sealed vessel and thus the potential for
 +
serious injury exists.</p>
-
<p>Effects of unforeseen release of Neurospora crassa into the environment
+
<h3 id=Section_2>2. If your response to any of the questions above is yes:</h3>
-
must be considered. We asked ourselves what would happen if either our WT or
+
<h4>a. Explain how you addressed these issues in project design
-
manipulated organism were to be released into the environment. </p>
+
and while conducting laboratory work.</h4>
-
<p>Neurospora has a well-documented history of living in close association
+
<p>To deal with such issues our team greatly scrutinized all
-
with humans. It is commonly regarded as having resided in a variety of
+
steps within our procedure for potential hazards and subsequently
-
businesses including bakeries, lumber yards and plywood factories1;<sup>9-11</sup>. Growth
+
collectively addressed all areas of concern. Please see the outline
-
has also been observed on the stuble of burnt sugar cane fields and along burnt
+
below.</p>
-
railway tracks<sup>13</sup>. However, these instances are largely documented as having
+
-
occurred in moist tropical or subtropical environments, the favored climate for
+
-
Neurospora growth<sup>1</sup>. Thus, if our organism, WT or manipulated, were to be
+
-
released in our dry, temperate environment, Edmonton, Alberta, Canada, with its
+
-
bitterly harsh winters, we would expect its survival and therefore impact to be
+
-
minimal. </p>
+
-
<p>Nonetheless, as we discussed in the human practices component of our
+
<br>
-
project, our alternative biodiesel has the potential to revolutionize economies
+
-
throughout the world and we must therefore regard the effects of environmental
+
-
exposure to Neurospora in other climates. Were Neurospora to be released in
+
-
other warmer climates it may readily survive, creating its own niche within the
+
-
environment. The aforementioned nonpathogenicity of the genus results in this not being a source of alarm<sup>1-4</sup>. </p>
+
-
<p>Team Alberta further predicts that our manipulated strain of Neurospora
+
<img width=650px src="https://static.igem.org/mediawiki/2011/7/74/TeamAlbertaReagentsAndChemicals.png">
-
crassa would be disadvantaged to survive relative to WT in any environment: our
+
-
genetic manipulation results in the organism’s energy being diverted away from
+
-
other essential functions, inhibiting organismal growth and manipulation. </p>
+
-
<h2>1.4 Biopart Malfunction</h2>
+
<br>
-
<p>Unforeseen alteration of biopart functions may represent issues of
+
<p>Our careful consideration of the potential safety concerns
-
concern. However, in our case if one or several of our bioparts were to change
+
associated with the completion of our project and our careful
-
their function or stop working as intended, limited safety issues would arise.
+
documentation and safety training have allowed us to work both
-
Our knockout of the FAD gene results in there being limited potential for its
+
responsibly and cautiously, limiting the probability of injuries.</p>
-
mutation and subsequent alteration of function. Further, our manipulation of
+
-
thioesterases similarly represents limited safety concerns due to their prevalence
+
-
in all organisms. If mutations were to occur in thioesterase genes and result
+
-
in overproduction, our organism would likely die or have a limited existence; if
+
-
mutation to thioesterase genes were to prevent its function, then the organism
+
-
may return to a more WT phenotype<sup>12</sup>.</p>
+
-
<h2>1.5 Device Malfunction</h2>
+
<br>
-
<p>The malfunction of our bioconverter represents a potential threat to safety.
+
<p>All MSDS data sheets were followed in working with strong acids,
-
The esterification process requires heat and flammable chemicals contained
+
bases, volatile chemicals, or otherwise potentially harmful reagents. All
-
within a sealed vessel and thus the potential for serious injury undoubtedly
+
antibiotics and antifungals were stored cold and in sealed containers,
-
exists. We addressed these areas of concern in our product design by
+
and any reacting chemicals were kept in separate storage areas. All
-
incorporating various safeguards. For example, our bioconverter is designed to
+
experiments done involving the use of open flame, electrical power
-
automatically turn off upon exceeding the recommended temperature and
+
sources, or flammable substances were treated with the utmost
-
pressure past specific margins.</p>
+
caution. All experiments with distillation used properly fitted apparatus
 +
and were designed to eliminate all possibility of heating closed
 +
containers. (For details on the safe handling of Hygromycin b, please
 +
see question 3).</p>
 +
<br>
-
<h2>1.6 Cumulative Risk Assessment</h2>
+
<p>Safety issues arising from the completion of our bioreactor were
 +
addressed in the stages of its design. Several important safeguards
 +
were incorporated. For example, our bioconverter is designed to
 +
automatically turn off upon exceeding the recommended temperature
 +
and pressure past specific margins.</p>
-
<center><b>Risk = Hazards x Probability<sup>13</sup> </b></center>
+
<h4>b. Describe and document safety, security, health and/or
 +
environmental issues as you submit your parts to the Registry.</h4>
-
<h2>Hazards</h2>
+
<p>In submitting parts to the registry, Team Alberta has actively
 +
undertaken the practice of describing and documenting any safety,
 +
security, health and/or environmental issues of our parts as they are
 +
submitted to the registry. None of our parts confer any pathogenicity
 +
or otherwise harmful effects upon either Neurospora crassa or
 +
Escherichia coli.</p>
-
<p>Any of the above mentioned areas of concern represent potential hazards
+
<br>
-
that may be associated with our project. Throughout our project’s design and
+
-
continued development, these were continually addressed and limited, where
+
-
possible.</p>
+
-
<h2>Probability</h2>
+
<p>One of our parts will confer Hygromycin B resistance to
 +
Neurospora crassa, and such strains that contain this part should be
 +
noted as being resistant to that particular antifungal agent.</p>
-
<p>The probability of our project resulting in any harm to members of our lab,
+
<h3 id=Section_3>3. Under what biosafety provisions will / do you operate?</h3>
-
    the community, or the environment is very low. This minimal probability can be
+
-
    attributed to our team’s careful consideration of the potential safety concerns
+
-
    associated with the completion of our project; our careful documentation and
+
-
    safety training have allowed us to work both responsibly and cautiously thus
+
-
    limiting the probability of injuries. Of course, if these practices were to subside,
+
-
    the probability of injury would proportionally increase. </p>
+
-
    <p>As mentioned before, our team plans to develop a contained bioconverter
+
<h4>a. Does your institution have its own biosafety rules and if so
-
    within which all the processes needed to carry out the production of our
+
what are they? Provide a link to them online if possible.</h4>
-
    alternative biodiesel are to be carried out; our team further aspires to scale-up
+
-
    this device for industrial production. Therefore, our project does not require the
+
-
    exposure or release of our engineered organism to people or the environment
+
-
    further limiting the probability of unfavorable events.</p>
+
-
    <h1>2. Documentation and management of safety issues</h1>
+
<p>Yes, the University of Alberta contains a specified set of
-
    <div id="horiz-line"></div>
+
Biosafety Rules that are outlined within the Biosafety Manual that is
 +
published by the Office of Environmental Health and Safety. A link to
 +
his valuable resource can be found here:
 +
<a href="http://www.ehs.ualberta.ca/en/EHSDivisions/~/media/8C72A3F626FB47B6BD70EC79C94CD1CE.ashx">
 +
http://www.ehs.ualberta.ca/en/EHSDivisions/~/media/8C72A3F626FB47B6BD70EC79C94CD1CE.ashx</a></p>
-
    <h2>2.1 Safe Handling of Biobrick Parts and Devices</h2>
+
<h4>b. Does your institution have an Institutional Biosafety
 +
Committee or equivalent group? If yes, have you discussed
 +
your project with them? Describe any concerns or changes
 +
that were made based on this review.</h4>
 +
<p>Team Alberta consulted the University of Alberta's Office of
 +
Environmental Health and Safety throughout the planning of our
 +
project. The Biosafety Division of this office provides both technical
 +
expertise and support to labs throughout campus and thus ensures
 +
a safe working and learning environment for both staff and students.
 +
In addition, the division also monitors compliance with federal and
 +
provincial legislation<sup>16</sup>.</p>
-
    <p>None of the genetic parts or devices that we have used or made this year
+
<br>
-
    raise any safety issues with the exception of our inserting a Hygromycin b resistance gene. The most predominant threat that this component possesses
+
-
    stems from our daily use of Hygromycin b in selection procedures. Hygromycin b
+
-
    is listed as potentially being fatal if absorbed through the skin or if swallowed. In
+
-
    addition, it is listed as being a hazard for birth defects, eye irritation, and
+
-
    respiratory tract irritation<sup>14</sup>. Our team diligently handled this area of concern by
+
-
    always wearing lab coats, safety goggles, and surgical masks when working with
+
-
    this potent antibiotic. Further, when our genetic parts are assembled into
+
-
    devices, no additional areas of concern arise. </p>
+
-
    <p>It should also be noted environmental exposure of our Hygromycin b
+
<p>Our team submitted a written overview of the relevant safety
-
    resistant strain does not pose a significant threat as our organism remains
+
considerations associated with our project to this office. Upon
-
    sensitize to other antifungal treatments such as Bleomycin<sup>15</sup>.</p>
+
perusal of this document, the Biosafety Officer provided us with
 +
recommendations to fully ensure that our project met all biosafety
 +
requirements. Please see the Biosafety Officer's recommendations
 +
below and how we met each of them:</p>
-
     <h2>2.2 Registry Documentation</h2>
+
<ol style="list-style-type:lower-roman">
 +
<li>
 +
     <p>It was recommended that if our lab were to grow N. Crassa beyond
 +
a 10L culture volume that we would have to obtain permission from
 +
the federal government, as such a culture would be considered large
 +
scale production. The Biosafety Officer therefore recommends avoiding
 +
production of cultures beyond a 9.5L volume. In addition, it was
 +
recommended that our lab prepare a Biological Spill Remediation kit
 +
according to the Standard Operating Protocols (SOP) of the university.</p>
-
    <p>The potential safety issues arising from inserting a Hygromycin b
+
<p>To avoid having to proceed with a lengthy governmental registration
-
    resistance gene will be carefully documented when our created part is submitted
+
process in the relatively short timeframe of the iGEM competition, our
-
    to the registry. We plan to outline the aforementioned hazards and proper
+
lab will ensure that no culture we prepare will be larger than 9.5L.
-
    handling procedures in our submission.</p>
+
To date, the largest culture that we have prepared has been less
 +
than 1.5L. As well, the members of our lab assembled the materials
 +
required for a Biological Spill Remediation kit, which will allow for the
 +
safe and efficient cleanup of any potential spills.</p>
 +
</li>
 +
<li>
 +
<p>The Biosafety officer recommended that our lab ensure all cultures
 +
of N. crassa be rendered inert prior to disposal.</p>
-
    <h2>2.3 Application to other teams</h2>
+
<p>The members of our lab have already been adhering to this practice
 +
by treating all cultures of N. crassa with a household bleach mixture
 +
for a minimum of one hour prior to disposal. Our bleach mixture
 +
contains a minimum of one part bleach to three parts culture, which is
 +
consistent with the guidelines outlined by the Biosafety Division of the
 +
Environmental Health and Safety Office.</p>
 +
</li>
-
    <p>Team Alberta feels that the best way that other teams can learn from our
+
<li>
-
    experiences with Hygromycin b and engineering Hygromycin b resistance is from
+
<p>The Biosafety Officer recommended that all individuals working
-
    the careful documentation of our work. As previously mentioned, Team Alberta
+
within our lab should wear personal protective equipment (PPE),
-
    intends to outline the potential hazards associated with Hygromycin b exposure
+
comprised of fully-fastened laboratory coats or gowns, disposable
-
    and procedures for its proper handling in our registry submission. This will afford
+
gloves, safety glasses, closed-toe shoes and floor-length pants.</p>
-
    future iGEM teams ease of access to information in determining how to safely
+
-
    handle the potent antibiotic and subsequently resistant strains.</p>
+
-
    <h1>3. Project Evaluation by University of Alberta Environmental Health and Safety</h1>
+
<p>The members of our lab have been adhering to this common
-
    <div id="horiz-line"></div>
+
laboratory practice since the very beginning.</p>
 +
</li>
-
    <p>Team Alberta consulted the University of Alberta’s Office of Environmental
+
<li>
-
    Health and Safety throughout the planning of our project. The Biosafety Division
+
<p>Due to the use of the antibiotic and antifungal Hygromycin b, the
-
    of this office provides both technical expertise and support to labs throughout
+
Biosafety Officer recommended that all work with this antibiotic be
-
    campus and thus ensures a safe working and learning environment for both staff
+
carried out in a fume hood. He further recommended that members
-
    and students. In addition, the division also monitors compliance with federal and
+
of our team coming in contact with this antibiotic be fit-tested for N95
-
    provincial legislation<sup>16</sup>. </p>
+
respirators in the event that this antibiotic cannot be handled within
 +
the fume hood.</p>
-
    <p>We have currently submitted a written overview of the relevant safety
+
<p>The members of our team who are not fit-tested for N95 respirators
-
    considerations associated with our project and we are patiently awaiting any final
+
will not work with Hygromycin b. The two members of our team that
-
    feedback that the Biosafety Officer may provide us with. Please check back
+
will work with this antibiotic have been fit-tested with N95 respirators
-
    shortly for an update.</p>
+
so that they may safely handle this compound, regardless of its form.
 +
All work with Hygromycin b will be carried out within a fumehood.</p>
 +
</li>
 +
</ol>
-
    <h1>4. Safety Considerations for Future iGEM Competitions</h1>
+
<h4>c. Will / did you receive any biosafety and/or lab training
-
    <div id="horiz-line"></div>
+
before beginning your project? If so, describe this training.</h4>
-
    <p>In order to motivate future iGEM teams to increasingly emphasize safety
+
<div class=safety-img id=manual>
-
    in their daily laboratory practices, we propose that each team completes a
+
<img src="https://static.igem.org/mediawiki/2011/c/c7/Manual.jpg">
-
    detailed safety report for a minimal medal requirement. </p>
+
Information about reagents and safety protocols are all found in an easily accessible binder
 +
</div>
-
    <p>Our team recognizes that biosafety engineering has the potential to make
+
<p>The members of our lab have all had variable levels of laboratory
-
    many devices and systems safer. For example, knockouts may be created
+
experience upon beginning the summer. Though most members
-
    through partial or full deletions as oppose to single nucleotide alterations that
+
had some level of biosafety training from previous lab experiences,
-
    are susceptible to mutation, auxtotrophic stains may be created, or antibiotic
+
including WHMIS certification, some members did not have any
-
    resistance may be removed. In our project, we are crossing out extraneous
+
previous safety training. Because of this uneven experience, all
-
    basta resistance found in mus strains from our final organism. </p>
+
members of our team received a laboratory tour at the beginning of
 +
the project. This included an overview of general laboratory safety
 +
procedures and protocols. Additionally, all team members were briefed
 +
on new safety procedures as our project developed. For example, upon
 +
beginning work with our organism, N. crassa, all lab members were
 +
briefed on its safe disposal.</p>
-
    <p>Bearing in mind the increasing importance of safety engineering, Team
+
<h4>d. Does your country have national biosafety regulations or
-
    Alberta further proposes that future iGEM competitions require teams to engineer
+
guidelines? If so, provide a link to them online if possible.</h4>
-
    their parts to have several safeguards in order to achieve a gold medal. Such
+
-
    components being medal requirements, we feel that it would increasingly
+
-
    motivate iGEM participants to more actively address potential safety issues in
+
-
    their future work, iGEM or otherwise. Moreover, this would encourage teams to
+
-
    be more creative with respects to organismal engineering.</p>
+
-
    <h1>References</h1>
+
<p>Yes, the Public Health Agency of Canada outlines national
-
    <div id="horiz-line"></div>
+
biosafety guidelines. Specifically, the Laboratory Biosafety and
-
1. Perkins DD and Davis RH:  <b>Evidence for Safety of Neurospora Species for Academic and Commercial Use.</b> <i>Applied and Environmental Microbiology 2006</i>, <b>66</b> (12): 5107-5109.
+
Biosecurity Division of this agency is responsible for publishing
 +
biosafety regulations. A quick synopsis of these guidelines may be
 +
found here:
-
2. Davis RH: <i>Neurospora: contributions of a model organism.</i> Oxford University Press US; 2000.
+
<a href="http://www.phac-aspc.gc.ca/lab-bio/regul/index-eng.php">http://www.phac-aspc.gc.ca/lab-bio/regul/index-eng.php</a></p>
-
3. Perkins DD: <b>Neurospora: the organism behind the molecular revolution.</b> <i>Genetics</i> 1992, <b>130</b>: 687-701.
+
<h3 id=Section_4>4.Optional Question: Do you have other ideas on how to deal
 +
with safety or security issues that could be useful for future
 +
iGEM competitions? How could parts, devices and systems be
 +
made even safer through biosafety engineering?</h3>
-
4. Perkins DD: <b>Neurospora genetics at the turn of the century</b>. <i>Fungal Genetics Newsletter</i> 2000, <b>47</b>: 83-88.
+
<p>In order to motivate future iGEM teams to further emphasize
 +
safety in their daily laboratory practices, we propose that each team
 +
completes a detailed safety report for a minimal medal requirement.
 +
Bearing in mind the increasing importance of safety in the rapidly
 +
expanding synthetic biology field, Team Alberta further proposes that
 +
future iGEM competitions require teams to engineer their parts and
 +
organisms with several safeguards in place in order to achieve a gold
 +
medal. If these components were medal requirements, we feel that it
 +
would greatly motivate iGEM participants to address potential safety
 +
issues, and not just obvious ones, far ahead of time. This attention
 +
towards possible problems, and the solutions around them, will pay
 +
immense rewards in their future work, iGEM or otherwise. Moreover,
 +
this would encourage teams to be more creative with respects to
 +
synthetic biology and genetic engineering.</p>
-
5. Frank MM, Austen KF, Claman HN, Unanue ER: <i>Samter’s immunological diseases, 5<sup>th</sup> ed.</i> Little, Brown & Co US; 1995.
+
<br>
-
6. Kay AB: <i>Allergy and allergic diseases.</i> Blackwell US; 1997.
+
<p>Our team recognizes that biosafety engineering has the potential
 +
to make many devices and systems safer. For example, essential
 +
gene knockouts can be created through partial or full amino acid
 +
deletions instead of single nucleotide alterations that are susceptible
 +
to mutation. Auxotrophic (or nutrient-deficient) strains can be created
 +
to ensure that an organism has no chance to exist outside of the
 +
laboratory environment, ensuring a minimal chance of contaminating
 +
the outside world. Organisms can even be designed to positively
 +
or negatively respond to varying wavelengths of light and electrical
 +
stimuli. All this and more should be taken into account when designing
 +
any organic circuit.</p>
-
7. Middleton E, Reed CE, Ellis EF, Adkinson, NF, Yunginger, Busse, WW: <i>Allergy: principles and practice, 5<sup>th</sup> ed.</i> Mosby US; 1998.
+
    <h3 id=References>References</h3>
-
8. Patterson R, Grammer LC, Greenberger PA: <i>Allergic diseases: diagnosis and management, 5<sup>th</sup> ed.</i> Lippncott-Raven US; 1997.  
+
<ol>
 +
    <li>
 +
Perkins DD and Davis RH: <b>Evidence for Safety of Neurospora
 +
Species for Academic and Commercial Use.</b><i> Applied and
 +
Environmental Microbiology </i>2006, 66 (12): 5107-5109.
 +
    </li>
-
9. Shear CL, Dodge BO: <b>Life histories and heterothallism of the red bread-mold fungi of the Monilia sitophila group.</b> <i>Journal Agricultural Research</i> 1927, <b>34</b>: 1019-1042.
+
    <li>
 +
Davis RH: <b>Neurospora: contributions of a model organism. </b><i>Oxford
 +
University Press</i> US; 2000.
 +
    </li>
-
10. Yassin S, Wheals A: <b>Neurospora species in bakeries.</b> <i>Journal Applied Bacteriology</i> 1992, <b>72</b>: 337-380.
+
    <li>
 +
Perkins DD: Neurospora: <b>the organism behind the molecular
 +
revolution. </b><i>Genetics </i>1992, 130: 687-701.
 +
    </li>
-
11. Shaw DE: <b>Honeybees collecting Neurospora spores from steam Pinus logs in Queensland.</b> <i>Mycologist</i> 1993, <b>7</b>: 182-185.
+
    <li>
 +
Perkins DD: <b> Neurospora genetics at the turn of the century.
 +
</b><i>Fungal Genetics Newsletter</i> 2000, 47: 83-88.
 +
    </li>
-
12.  Nelson DL, Cox MM: <i>Lehninger Principles of Biochemistry.</i> W.H Freeman US; 2009.
+
    <li>
 +
Frank MM, Austen KF, Claman HN, Unanue ER: <b>Samterâs
 +
immunological diseases, 5th ed. </b><i> Little, Brown & Co</i> US; 1995.
 +
    </li>
-
13. iGEM website: https://2011.igem.org/Safety
+
    <li>
 +
Kay AB: <b>Allergy and allergic diseases. </b><i> Blackwell</i> US; 1997.
 +
    </li>
-
14. Hygromycin material and safety data sheet: http://www.invivogen.com/MSDS/
+
    <li>
-
HygromycinB_solution_v4_MSDS.pdf
+
Middleton E, Reed CE, Ellis EF, Adkinson, NF, Yunginger, Busse,
 +
WW: <b>Allergy: principles and practice, 5th ed. </b><i> Mosby </i>US; 1998.
 +
    </li>
-
15. Kafer E, Luk D: <b>Sensitivity of bleomycin and hydrogen peroxide of DNA repair-defective mutants in Neurospora crassa.</b> <i>Mutant Research</i> 1989, <b>217</b>(1): 75-81.  
+
    <li>
 +
Patterson R, Grammer LC, Greenberger PA: <b>Allergic diseases:
 +
diagnosis and management, 5th ed. </b><i> Lippncott-Raven </i> US; 1997.
 +
    </li>
 +
 
 +
    <li>
 +
Shear CL, Dodge BO: <b>Life histories and heterothallism of the
 +
red bread-mold fungi of the Monilia sitophila group. </b><i>Journal
 +
Agricultural Research </i>1927, 34: 1019-1042.
 +
    </li>
 +
 
 +
    <li>
 +
Yassin S, Wheals A: <b>Neurospora species in bakeries. </b><i>Journal
 +
Applied Bacteriology </i>1992, 72: 337-380.
 +
    </li>
 +
 
 +
    <li>
 +
Shaw DE: <b>Honeybees collecting Neurospora spores from
 +
steam Pinus logs in Queensland. </b><i> Mycologist </i>1993, 7: 182-185.
 +
    </li>
 +
 
 +
    <li>
 +
Nelson DL, Cox MM: <b>Lehninger Principles of Biochemistry.</b><i> W.H
 +
Freeman </i>US; 2009.
 +
    </li>
 +
 
 +
    <li>
 +
<b>iGEM website:</b><i> https://2011.igem.org/Safety </i>
 +
    </li>
 +
 
 +
    <li>
 +
<b>Hygromycin material and safety data sheet: </b><br><i>
 +
http://www.invivogen.com/MSDS/HygromycinB_solution_v4_MSDS.pdf</i>
 +
    </li>
 +
 
 +
    <li>
 +
Kafer E, Luk D: <b>Sensitivity of bleomycin and hydrogen
 +
peroxide of DNA repair-defective mutants in Neurospora
 +
crassa. </b><i>Mutant Research</i> 1989, 217(1): 75-81.
 +
    </li>
 +
 
 +
    <li>
 +
<b>University of Alberta Environmental Health and Safety website:</b><i>
 +
http://www.ehs.ualberta.ca/ </i>
 +
    </li>
 +
</ol>
 +
 
 +
    </div>
 +
</div>
-
16. University of Alberta Environmental Health and Safety website: http://www.ehs.ualberta.ca/
+
</html>
-
{{Team:Alberta/endMainContent}}
+
{{Team:Alberta/footer|achievements=selected}}

Latest revision as of 16:41, 26 September 2011

METHODOLOGY

Team Alberta iGEM Safety Proposal

Team Alberta recognizes the important role that safety plays in the completion of any scientific undertaking. Our team members are fully committed to not only ensuring the safety of those within our lab but also addressing any potential risks that our project may pose to the greater community.
Please find below Team Alberta's answers to the new, required iGEM safety questions.

1. Would the materials used in your project and/or your final product pose:

a. Risks to the safety and health of team members or others in the lab?

Neurospora is stored in clearly labelled flasks with sponge tops to prevent contamination

The materials used in our project pose little to no risks to the safety and health of our team members or others in the lab and our final product poses no risk to these individuals.


The organism that our group has genetically manipulated, Neurospora crassa, is well-documented as being safe1. N. crassa is a biosafety level 1 organism. Since 1941 the species has been used extensively in laboratories resulting in the publication of thousands of research papers. In none of these numerous publications has the Neurospora genus been implicated in causing disease in either animals or plants1-4.


Neurospora's minimal threat can largely be attributed to it being an obligate aerobe. Unlike other species such as yeasts, Neurospora is unable to grow in organs such as the gut or bladder, tissues, or systemically within an organism1. Furthermore, despite Neurospora being regarded by some as an allergen, medical literature generally fails to mention the genus5-8.


Our team acknowledges that though certain fungal species themselves may not be pathogenic, they may produce dangerous toxins. Throughout our research into Neurospora our group has not found any documentation of the species producing dangerous mycotoxins or any dangerous secondary metabolites1. Given that Neurospora has had such a long history of living in close association with humans (please see below), it is a generally regarded as being harmless.

Clearly labelled waste disposal containers ensure proper removal of potentially hazardous materials

It should be noted that there are still materials used in our project that may potentially be hazardous. Our esterification procedure involves the use of strong chemicals and reagents, some of which have the potential to cause harm. These reagents are primarily volatile and require careful handling and storage.


As well, Hygromycin b, an antibiotic and antifungal agent that is used for selection, is toxic if inhaled. Care must be taken around powdered or lyophilized Hygromycin B, and precaution should be taken to cover up respiratory passageways, skin, and the eyes. However, Hygromycin B is most commonly purchased in liquid form, and in liquid carries with it significantly less potential for accidental inhalation.

b. Risks to the safety and health of the general public if released by design or accident?

Our organism poses no threat to the safety and health of the general public. Neurospora has a well-documented history of living in close association with humans. It has historically resided in a variety of businesses including bakeries, lumber yards and plywood factories1;9- 11. Growth has also been observed on the stubble of burnt sugar cane fields and along burnt railway tracks13. Thus, it poses no threat to the health and safety of those in its surroundings.

c. Risks to environmental quality if released by design or accident?

As mentioned, N. crassa has a long history of living in close association with humans; however, these instances are largely documented as having occurred in moist tropical or subtropical environments. Like many fungi, warm and moist is the favored climate for Neurospora growth1. Thus, if our organism, WT or manipulated, were to be released in our dry, temperate environment of Edmonton, Alberta, Canada, with its bitterly harsh winters, we would expect its survival and environmental impact to be minimal. Even if it were to survive, we cannot see it affecting local ecosystems by any appreciable amount.


Nonetheless, our alternative biodiesel has the potential to revolutionize economies throughout the world, as discussed in our human practices component, and we must therefore regard the effects of environmental exposure to Neurospora in other climates. Were Neurospora to be released in other warmer climates it may readily survive, creating its own niche within the environment. However, the aforementioned nonpathogenicity of the genus results in this not being a source of alarm1-4.


Further, Team Alberta predicts that our manipulated strain of Neurospora crassa would be disadvantaged to survive relative to WT in any environment; our genetic manipulation results in the organism's energy being diverted away from essential metabolic functions, inhibiting organismal growth and manipulation. This puts it at a disadvantage against natural competing fungi.

d. Risks to security through malicious misuse by individuals, groups or states?

Unforeseen alteration of biopart functions may represent issues of concern. However, in our case if one or several of our bioparts were changed to alter their function or stop working as intended, limited to no safety issues would arise. Our knockout of the FadD gene results in there being limited potential for its alteration of function. Further, our manipulation of thioesterases similarly represents limited safety concerns due to their prevalence in all organisms. If further alterations were to be made to thioesterase genes and resulted in overproduction, our organism would likely die or have a limited existence; if further alterations to thioesterase genes were to prevent their function, then the organism may return to a more WT phenotype12.


The misuse of our bioconverter represents a potential threat to safety. The esterification process requires heat and flammable chemicals contained within a sealed vessel and thus the potential for serious injury exists.

2. If your response to any of the questions above is yes:

a. Explain how you addressed these issues in project design and while conducting laboratory work.

To deal with such issues our team greatly scrutinized all steps within our procedure for potential hazards and subsequently collectively addressed all areas of concern. Please see the outline below.



Our careful consideration of the potential safety concerns associated with the completion of our project and our careful documentation and safety training have allowed us to work both responsibly and cautiously, limiting the probability of injuries.


All MSDS data sheets were followed in working with strong acids, bases, volatile chemicals, or otherwise potentially harmful reagents. All antibiotics and antifungals were stored cold and in sealed containers, and any reacting chemicals were kept in separate storage areas. All experiments done involving the use of open flame, electrical power sources, or flammable substances were treated with the utmost caution. All experiments with distillation used properly fitted apparatus and were designed to eliminate all possibility of heating closed containers. (For details on the safe handling of Hygromycin b, please see question 3).


Safety issues arising from the completion of our bioreactor were addressed in the stages of its design. Several important safeguards were incorporated. For example, our bioconverter is designed to automatically turn off upon exceeding the recommended temperature and pressure past specific margins.

b. Describe and document safety, security, health and/or environmental issues as you submit your parts to the Registry.

In submitting parts to the registry, Team Alberta has actively undertaken the practice of describing and documenting any safety, security, health and/or environmental issues of our parts as they are submitted to the registry. None of our parts confer any pathogenicity or otherwise harmful effects upon either Neurospora crassa or Escherichia coli.


One of our parts will confer Hygromycin B resistance to Neurospora crassa, and such strains that contain this part should be noted as being resistant to that particular antifungal agent.

3. Under what biosafety provisions will / do you operate?

a. Does your institution have its own biosafety rules and if so what are they? Provide a link to them online if possible.

Yes, the University of Alberta contains a specified set of Biosafety Rules that are outlined within the Biosafety Manual that is published by the Office of Environmental Health and Safety. A link to his valuable resource can be found here: http://www.ehs.ualberta.ca/en/EHSDivisions/~/media/8C72A3F626FB47B6BD70EC79C94CD1CE.ashx

b. Does your institution have an Institutional Biosafety Committee or equivalent group? If yes, have you discussed your project with them? Describe any concerns or changes that were made based on this review.

Team Alberta consulted the University of Alberta's Office of Environmental Health and Safety throughout the planning of our project. The Biosafety Division of this office provides both technical expertise and support to labs throughout campus and thus ensures a safe working and learning environment for both staff and students. In addition, the division also monitors compliance with federal and provincial legislation16.


Our team submitted a written overview of the relevant safety considerations associated with our project to this office. Upon perusal of this document, the Biosafety Officer provided us with recommendations to fully ensure that our project met all biosafety requirements. Please see the Biosafety Officer's recommendations below and how we met each of them:

  1. It was recommended that if our lab were to grow N. Crassa beyond a 10L culture volume that we would have to obtain permission from the federal government, as such a culture would be considered large scale production. The Biosafety Officer therefore recommends avoiding production of cultures beyond a 9.5L volume. In addition, it was recommended that our lab prepare a Biological Spill Remediation kit according to the Standard Operating Protocols (SOP) of the university.

    To avoid having to proceed with a lengthy governmental registration process in the relatively short timeframe of the iGEM competition, our lab will ensure that no culture we prepare will be larger than 9.5L. To date, the largest culture that we have prepared has been less than 1.5L. As well, the members of our lab assembled the materials required for a Biological Spill Remediation kit, which will allow for the safe and efficient cleanup of any potential spills.

  2. The Biosafety officer recommended that our lab ensure all cultures of N. crassa be rendered inert prior to disposal.

    The members of our lab have already been adhering to this practice by treating all cultures of N. crassa with a household bleach mixture for a minimum of one hour prior to disposal. Our bleach mixture contains a minimum of one part bleach to three parts culture, which is consistent with the guidelines outlined by the Biosafety Division of the Environmental Health and Safety Office.

  3. The Biosafety Officer recommended that all individuals working within our lab should wear personal protective equipment (PPE), comprised of fully-fastened laboratory coats or gowns, disposable gloves, safety glasses, closed-toe shoes and floor-length pants.

    The members of our lab have been adhering to this common laboratory practice since the very beginning.

  4. Due to the use of the antibiotic and antifungal Hygromycin b, the Biosafety Officer recommended that all work with this antibiotic be carried out in a fume hood. He further recommended that members of our team coming in contact with this antibiotic be fit-tested for N95 respirators in the event that this antibiotic cannot be handled within the fume hood.

    The members of our team who are not fit-tested for N95 respirators will not work with Hygromycin b. The two members of our team that will work with this antibiotic have been fit-tested with N95 respirators so that they may safely handle this compound, regardless of its form. All work with Hygromycin b will be carried out within a fumehood.

c. Will / did you receive any biosafety and/or lab training before beginning your project? If so, describe this training.

Information about reagents and safety protocols are all found in an easily accessible binder

The members of our lab have all had variable levels of laboratory experience upon beginning the summer. Though most members had some level of biosafety training from previous lab experiences, including WHMIS certification, some members did not have any previous safety training. Because of this uneven experience, all members of our team received a laboratory tour at the beginning of the project. This included an overview of general laboratory safety procedures and protocols. Additionally, all team members were briefed on new safety procedures as our project developed. For example, upon beginning work with our organism, N. crassa, all lab members were briefed on its safe disposal.

d. Does your country have national biosafety regulations or guidelines? If so, provide a link to them online if possible.

Yes, the Public Health Agency of Canada outlines national biosafety guidelines. Specifically, the Laboratory Biosafety and Biosecurity Division of this agency is responsible for publishing biosafety regulations. A quick synopsis of these guidelines may be found here: http://www.phac-aspc.gc.ca/lab-bio/regul/index-eng.php

4.Optional Question: Do you have other ideas on how to deal with safety or security issues that could be useful for future iGEM competitions? How could parts, devices and systems be made even safer through biosafety engineering?

In order to motivate future iGEM teams to further emphasize safety in their daily laboratory practices, we propose that each team completes a detailed safety report for a minimal medal requirement. Bearing in mind the increasing importance of safety in the rapidly expanding synthetic biology field, Team Alberta further proposes that future iGEM competitions require teams to engineer their parts and organisms with several safeguards in place in order to achieve a gold medal. If these components were medal requirements, we feel that it would greatly motivate iGEM participants to address potential safety issues, and not just obvious ones, far ahead of time. This attention towards possible problems, and the solutions around them, will pay immense rewards in their future work, iGEM or otherwise. Moreover, this would encourage teams to be more creative with respects to synthetic biology and genetic engineering.


Our team recognizes that biosafety engineering has the potential to make many devices and systems safer. For example, essential gene knockouts can be created through partial or full amino acid deletions instead of single nucleotide alterations that are susceptible to mutation. Auxotrophic (or nutrient-deficient) strains can be created to ensure that an organism has no chance to exist outside of the laboratory environment, ensuring a minimal chance of contaminating the outside world. Organisms can even be designed to positively or negatively respond to varying wavelengths of light and electrical stimuli. All this and more should be taken into account when designing any organic circuit.

References

  1. Perkins DD and Davis RH: Evidence for Safety of Neurospora Species for Academic and Commercial Use. Applied and Environmental Microbiology 2006, 66 (12): 5107-5109.
  2. Davis RH: Neurospora: contributions of a model organism. Oxford University Press US; 2000.
  3. Perkins DD: Neurospora: the organism behind the molecular revolution. Genetics 1992, 130: 687-701.
  4. Perkins DD: Neurospora genetics at the turn of the century. Fungal Genetics Newsletter 2000, 47: 83-88.
  5. Frank MM, Austen KF, Claman HN, Unanue ER: Samterâs immunological diseases, 5th ed. Little, Brown & Co US; 1995.
  6. Kay AB: Allergy and allergic diseases. Blackwell US; 1997.
  7. Middleton E, Reed CE, Ellis EF, Adkinson, NF, Yunginger, Busse, WW: Allergy: principles and practice, 5th ed. Mosby US; 1998.
  8. Patterson R, Grammer LC, Greenberger PA: Allergic diseases: diagnosis and management, 5th ed. Lippncott-Raven US; 1997.
  9. Shear CL, Dodge BO: Life histories and heterothallism of the red bread-mold fungi of the Monilia sitophila group. Journal Agricultural Research 1927, 34: 1019-1042.
  10. Yassin S, Wheals A: Neurospora species in bakeries. Journal Applied Bacteriology 1992, 72: 337-380.
  11. Shaw DE: Honeybees collecting Neurospora spores from steam Pinus logs in Queensland. Mycologist 1993, 7: 182-185.
  12. Nelson DL, Cox MM: Lehninger Principles of Biochemistry. W.H Freeman US; 2009.
  13. iGEM website: https://2011.igem.org/Safety
  14. Hygromycin material and safety data sheet:
    http://www.invivogen.com/MSDS/HygromycinB_solution_v4_MSDS.pdf
  15. Kafer E, Luk D: Sensitivity of bleomycin and hydrogen peroxide of DNA repair-defective mutants in Neurospora crassa. Mutant Research 1989, 217(1): 75-81.
  16. University of Alberta Environmental Health and Safety website: http://www.ehs.ualberta.ca/