Team:Chiba/Safety

Basic Safety Questions for iGEM 2013

iGEM 2013 Biosafety Form 1

Q1. Please describe the chassis organism(s) you will be using for this project. If you will be using more than one chassis organism, provide information on each of them:

	Species	Strain no/name	Risk Group	Risk group source link	Disease risk to humans?If so, which disease?
1	E. coli	XL10 Gold	1	www.absa.org/riskgroups/bacteria	Yes.May cause irritation to skin.
2	E. coli	BL21	1	www.absa.org/riskgroups/bacteria	Yes.May cause irritation to skin.
3	E. coli	Shuffle competent	1	http://www.absa.org/riskgroups/index.html	Yes.May cause irritation to skin.
4	S.cerevisiae	W303	1	http://www.absa.org/riskgroups/index.html	No.
5	S.cerevisiae	H207	1	http://www.absa.org/riskgroups/index.html	No.
6	S.cerevisiae	BY4741	1	http://www.absa.org/riskgroups/index.html	No.
7	S.cerevisiae	YPH499	1	http://www.absa.org/riskgroups/index.html	No.

Q2. Highest Risk Group Listed:
Ans. 1



Q3. List and describe all new or modified coding regions you will be using in your project. (If you use parts from the 2013 iGEM Distribution without modifying them, you do not need to list those parts.)
Ans.

	Part number	Where did you get the physical DNA for this part (which lab, synthesis company, etc)	What species does this part originally come from?	What is the Risk Group of the species?	What is the function of this part, in its parent species?
1	BBa_K1057001	BBa_1746908	Aequorea victoria	1	Fluorescent protein(super folder GFP)
2	BBa_K1057002	Synthesized, DNA 2.0	Homo sapiens	1	Iron storange container proteins(2 orfs)
3	BBa_K1057003	Synthesized, DNA 2.0	Homo sapiens	1	Aid ferritin to store irons(chaperon)
4	BBa_K1057004	Synthesized, DNA 2.0	Homo sapiens	1	Three proteins coding for iron storage proteins and Fe chaperon
5	BBa_K1057005	Purchased from Addgene	Streptococcus pyogenes	2	Cas9 protein (exponuclease; useas CRISPi regulator protein)
6	BBa_K1057006	Modification from the material purchased from Addgene	Streptococcus pyogenes(partially)	2	sgRNA targeting to Fe uptake regulator fur and exportor fiefF in E. coli)
7	BBa_K1057007	Modification from the material purchased from Addgene	Streptococcus pyogenes(partially)	2	sgRNA targeting thioredoxin reductase in E.coli
8	BBa_K1057008	Modification from the material purchased from Addgene	Streptococcus pyogenes(partially)	2	sgRNA targeting glutathione reductase in E.coli

Q4. Do the biological materials used in your lab work pose any of the following risks? Please describe.
a. Risks to the safety and health of team members or others working in the lab?
Ans. No risk. Genes and modification version used in this project are half from human and are no reported toxicity to human being or other living system.

b. Risks to the safety and health of the general public, if released by design or by accident?
Ans. None.

c. Risks to the environment, if released by design or by accident?
Ans. None.

d. Risks to security through malicious misuse by individuals, groups, or countries?
Ans. None. To remove genes and organisms used in this project from our lab is restricted. In addition, we control them not to be taken out by other people.


Q5. If your project moved from a small-scale lab study to become widely used as a commercial/industrial product, what new risks might arise? (Consider the different categories of risks that are listed in parts a-d of the previous question.) Also, what risks might arise if the knowledge you generate or the methods you develop became widely available? (Note: This is meant to be a somewhat open-ended discussion question.)
Ans. Production and dispersion of amount of ferritin could result in the decrease from environment.　Moreover, because this project use iron solution, if this project is carried out in large scale, huge amount of iron solution occurs and they probably pollute environment.


Q6. Does your project include any design features to address safety risks? (For example: kill switches, auxotrophic chassis, etc.) Note that including such features is not mandatory to participate in iGEM, but many groups choose to include them.
Ans. No. Due to the nature of the DNA we use in this project, we did not find need for safety devices such as kill-swithces (spontaneous mutations anyway kill off the suicide devices…)


Q7. What safety training have you received (or plan to receive in the future)? Provide a brief description, and a link to your institution’s safety training requirements, if available.
Ans.1. Official seminars on recombinant DNA and its biosafety rules in Japan.
Ans.2. Special lectures and coaching held by our mentors, Prof. Daisuke Umeno and Prof. Shigeko Kawai.


Q8. Under what biosafety provisions will / do you work?
a. Please provide a link to your institution biosafety guidelines.
http://www.jm.chiba-u.jp/kenkyu/005/newpage1.htm

b. Does your institution have an Institutional Biosafety Committee, or an equivalent group? If yes, have you discussed your project with them? Describe any concerns they raised with your project, and any changes you made to your project plan based on their review.
Ans. Yes. One of our mentors, Prof. Daisuke Umeno, is the board member of the Chiba University Biosafety Committee.

c. Does your country have national biosafety regulations or guidelines? If so, please provide a link to these regulations or guidelines if possible.
Ministry Education:http://www.bch.biodic.go.jp/bioethics/anzen.html
Ministry Environment:http://www.lifescience.mext.go.jp/english/e_index.html

d. According to the WHO Biosafety Manual, what is the BioSafety Level rating of your lab? (Check the summary table on page 3, and the fuller description that starts on page 9.) If your lab does not fit neatly into category 1, 2, 3, or 4, please describe its safety features.
Ans. BSL2

e. What is the Risk Group of your chassis organism(s), as you stated in question 1? If it does not match the BSL rating of your laboratory, please explain what additional safety measures you are taking.
Ans. We use Saccharomyces cerevisiae(BSL1) and E.coli(K-12,BSL1), only and the place of experiment(Umeno lab, BSL2) is suitable manipulating them.

Faculty Advisor Name: Daisuke Umeno

iGEM 2013 Biosafety Form 2 Part 2 This form must be completed separately for each organism or part above RG1. Please cite sources, including web links as applicable, to support your statements. 1.Organism name and strain name or number. Homo sapiens 2. Organism Risk Group: 1 3. If you are using this organism as a chassis, write "chassis". If you are using a genetic part from the organism, give the name of the part and a brief description of what it does and why you are using it. BBa_K1057003, BBa_K1057004 Human ferritin genes consist of ferritin heavy chain FTH and ferritin light chain FTL. 4. How did you physically acquire the organism or part? Synthesis (DNA2.0) 5. What potential safety/health risks to team members, other people at your institution, or the general public could arise from your use of this organism/part? None 6. What measures do you intend to take to ensure that your project is safe for team members, other people at your institution, and the general public? Never been reported to be toxic. Also, this is a coding sequence that boost ferric ions. 7. If you are using only a part from the organism, and you believe the part by itself is not dangerous, explain why you believe it is not dangerous. The function of these genes is only as a container of ferric ions. 8. Why do you need to use this organism/part? Is there an organism/part from a less dangerous Risk Group that would accomplish the same purpose? The core parts for accumulation and storage of Fe(ii)/(iii) are from one of the safest organisms E. coli or from H. sapiens. Also, these genes are well described and have shown to be functional at least in S. cerevisiae. 9. Is the organism/part listed under the Australia Group guidelines, or otherwise restricted for transport? If so, how will your team ship this part to iGEM and the Jamborees? No. 10. Please describe the BioSafety Level of the lab in which the team works, or description of safety features of lab (Refer to Basic Safety form, question 8. d.). If you are using organisms with a BSL level greater than you lab, please explain any additional safety precautions you are taking. BSL2 Faculty Advisor Name: Daisuke Umeno

iGEM 2013 Biosafety Form Part 2

This form must be completed separately for each organism or part above RG1. Please cite sources, including web links as applicable, to support your statements.
1.Organism name and strain name or number.
Homo sapiens 2. Organism Risk Group:
1 3. If you are using this organism as a chassis, write "chassis". If you are using a genetic part from the organism, give the name of the part and a brief description of what it does and why you are using it.
BBa_K1057002, BBa_K1057004
Human iron chaperone PCB1 that accelerate the accumulation of ferric ions into ferritin.
4. How did you physically acquire the organism or part?
Synthesis (DNA2.0) 5. What potential safety/health risks to team members, other people at your institution, or the general public could arise from your use of this organism/part?
None.

6. What measures do you intend to take to ensure that your project is safe for team members, other people at your institution, and the general public?
Never been reported to be toxic. Also, this is a coding sequence that boost the ferritin function.

7. If you are using only a part from the organism, and you believe the part by itself is not dangerous, explain why you believe it is not dangerous.
The function of these genes is only as a container of ferric ions.

8. Why do you need to use this organism/part? Is there an organism/part from a less dangerous Risk Group that would accomplish the same purpose?
The core parts for accumulation and storage of Fe(ii)/(iii) are from one of the safest organisms E. coli or from H. sapiens. Also, these genes are well described and have shown to be functional at least in S. cerevisiae.

9. Is the organism/part listed under the Australia Group guidelines, or otherwise restricted for transport? If so, how will your team ship this part to iGEM and the Jamborees? No. 10. Please describe the BioSafety Level of the lab in which the team works, or description of safety features of lab (Refer to Basic Safety form, question 8. d.). If you are using organisms with a BSL level greater than you lab, please explain any additional safety precautions you are taking. BSL2 Faculty Advisor Name: Daisuke Umeno

iGEM 2013 Biosafety Form Part 2 This form must be completed separately for each organism or part above RG1. Please cite sources, including web links as applicable, to support your statements. 1.Organism name and strain name or number. Streptococcus pyogenes 2. Organism Risk Group: 1 3. If you are using this organism as a chassis, write "chassis". If you are using a genetic part from the organism, give the name of the part and a brief description of what it does and why you are using it. BBa_K1057005 Inactivated CAS9 gene used for site-specific repression of the target genes. 4. How did you physically acquire the organism or part? Purchased from AddGene. 5. What potential safety/health risks to team members, other people at your institution, or the general public could arise from your use of this organism/part? None. 6. What measures do you intend to take to ensure that your project is safe for team members, other people at your institution, and the general public? Never been reported to be toxic. Also, this is just an inactivated. 7. If you are using only a part from the organism, and you believe the part by itself is not dangerous, explain why you believe it is not dangerous. The function of these genes is only to bind some specific site of DNA. 8. Why do you need to use this organism/part? Is there an organism/part from a less dangerous Risk Group that would accomplish the same purpose? Genes from this organism is the only known system that can be used for CRISPRi technique. 9. Is the organism/part listed under the Australia Group guidelines, or otherwise restricted for transport? If so, how will your team ship this part to iGEM and the Jamborees? No. 10. Please describe the BioSafety Level of the lab in which the team works, or description of safety features of lab (Refer to Basic Safety form, question 8. d.). If you are using organisms with a BSL level greater than you lab, please explain any additional safety precautions you are taking. BSL2 Faculty Advisor Name: Daisuke Umeno