These Frequently Asked Questions (FAQs) are designed to help you complete the Biological Use Authorization
(BUA) applications. The below FAQs correspond to questions on the full BUA Application. The FAQs also pertain to the Request for Change to BUA Application; although, the numbering of the referenced questions differs. See the IBC webpage for information about the review process and to download the applications.
Index
Q. What trainings do I need to have completed?

A. The Instatutional Biosafety Committee (IBC) requires every person working on IBC approved protocols to have completed the Blood Borne Pathogens and Researcher Biosafety modules. These two courses are available on the CITI webpage . If you do not have a CITI account, please click the register here link to create your account. If you are already registered, login with your credentials. Please be sure that you enter your UNCG Employee/Student ID number. If you are NOT affiliated with UNCG, please enter 9 zeros (000000000) as your ID number.
Q7.
I am asked to list the species of all immunodeficient animals used in my research. Does this apply only to animals that are constitutively immunodeficient, or does it include animals that are treated with drugs or other agents that temporarily cause them to be immunodeficient?
A.
For the purpose of this form, the term 'immunodeficient animal' applies to all animals whose immune systems have been altered by genetic engineering, spontaneous mutations, or chemicals. In other words, the term 'immunodeficient animal' applies to all animals that are immunodeficient, regardless of the mechanism by which they were rendered immune-deficient or the period of time for which they are immunodeficient.
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Qs17-20.
Throughout the Culture of Primary Cells or Cell Lines section I am asked to list the 'type' and 'source' of my cells/cell lines. What do you mean by 'type' and 'source' of cells/cell lines?
A.
'Type' of primary cells means human or animal primary cells taken directly from blood or living tissue
(e.g., peripheral blood mononuclear cells, T cells, dendritic cells, etc.).
'Type' of cell lines means human or animal cell lines that may or may not be well characterized [e.g., human derived HEK293 cell lines, Epstein-Barr Virus (EBV)-transformed lymphoblastoid cell lines, embryonic stem cell (ESC) lines, induced pluripotent stem cell (iPSC) lines, etc.].
if your cell lines contain Return to Index
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Recombinant and Synthetic
, be sure to include them in the Gene Delivery Methods
table (question 44), even if they were engineered prior to the application.
By 'Source,' we mean the source from where you obtained the primary cells/cell lines (e.g., cells were isolated in my laboratory; cells were obtained from another UW PI; cells were obtained from a collaborator at
Fred Hutchinson Cancer Research Center (FHCRC); cells were purchased them from a commercial vendor-
American Type Culture Collection (ATCC)].
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Q.
What information should I provide if I work with human tissue, blood, or body fluids, or culture of human primary cells or cell lines?
A.
If you answered 'Yes' to questions 11 and/or 17 for work with human blood, tissue, or body fluids, or culture of human primary cells or cell lines, respectively, the
. Annual BBP training is also required. For information about the UW BBP program, see the UNCG EHS webpage .
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Q.
My project involves work with non-recombinant microorganisms as well as recombinant microorganisms. In which sections of the BUA application should I list them?

A.
For work involving non-recombinant microorganisms, to include wild-type and naturally occurring mutant species, complete question 22. In your answer, provide the genus and species of the non-recombinant microorganisms and describe the laboratory activities for which they will be utilized (e.g., Trypanosoma cruzi - agent of Chagas disease- is grown in vitro and used in mice for compound efficacy testing). If you are working with naturally occurring mutant species, you must also provide information about the naturally occurring mutations and their effects on the microorganisms. For examples of non-recombinant
microorganisms, see the FAQ for Q22 .
If you administer non-recombinant microorganisms to animals, complete question 23. In your answer, specify the non-recombinant microorganisms to be administered, the animal species, the route of administration, and the dose and/or concentration.
For work involving recombinant microorganisms complete question 24, all applicable questions of the
Recombinant and Synthetic DNA and RNA section (questions 31-43), and the Gene Delivery Methods table
(question 44). For the definition and examples of recombinant microorganisms, see the FAQ for Q24 .
Questions 25 and 26 must be answered for work with both recombinant and non-recombinant microorganisms.
If you are working with a large number of recombinant and/or non-recombinant organisms, please provide an attached list of the organisms.
Be aware that your application may be returned to you as incomplete if any applicable information is omitted.
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Q22.
Can you provide some examples of non-recombinant microorganisms?
A.
Non-recombinant microorganisms include the following:

Wild-type, naturally occurring species of bacteria, viruses, yeasts, fungi, parasites, and prions
(e.g., Salmonella typhimurium, Bacillus cereus, Treponema pallidum ).

Naturally occurring mutant species of bacteria, viruses, yeasts, fungi, parasites, and prions
(e.g., Bacillus Calmette-
Guérin, Chlamydia trachomatis
LGV strains).
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Q24.
Can you give some examples of recombinant microorganisms?
A.
Recombinant microorganisms include bacteria, viruses, yeasts, fungi, and/or parasites whose
genetic material has been altered using Return to Index
Recombinant and Synthetic
technology. Examples of recombinant
microorganisms are Listeria monocytogenes expressing ovalbumin, Toxoplasma gondii luciferase expressing PRU-Luc-GFP type II strain, and gene cloning done in E. coli K12. The above definition also applies to all forms of viral vectors used for gene transfer, genetic engineering, and gene therapy.
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Q.
How do you define recDNA molecules?
A.
Recombinant and synthetic nucleic acid molecules or "recDNA" molecules, including those that are chemically or otherwise modified analogs of nucleotides (e.g., morpholinos) or both, are defined in the context of the NIH Guidelines as follows. i. Molecules that (a) are constructed by joining nucleic acid molecules and (b) can replicate in a living cell (i.e., recombinant nucleic acids). ii. Nucleic acid molecules that are chemically or by other means synthesized or amplified, including those that are chemically or otherwise modified but can base pair with naturally occurring nucleic acid molecules (i.e., synthetic nucleic acids). iii. Molecules that result from the replication of those described in (i) or (ii) above.
The above definition reflects the March 2013 amendment to the NIH Guidelines .
Examples of recDNA molecules outside of living cells that fall under the above definition include the following.

Cloning DNA in bacterial plasmids
 siRNA

Oligonucleotide primers
 naked DNA from PCR or DNA sequencing
 base-pair analogs

Note that naked DNA molecules in test-tubes are exempt from the NIH Guidelines ; however, when in an organism, naked DNA molecules are not exempt.
All forms of recDNA are treated as biohazards regardless of whether the recDNA molecules are (a) purchased from a vendor (e.g., purchase a human cell line containing a plasmid), (b) constructed in your laboratory, or (c) used to transfect cells.
All recDNA, irrespective of exemption from the NIH Guidelines, must be treated as a biohazard. See the IBC website for the IBC's working definition of biohazardous agents.
Containment practices for work with recDNA in BSL-1 or higher laboratories are as stipulated in the UNCG Biosafety Manual. Information about identifying, segregating, decontaminating, and properly packaging and disposing of biohazardous waste, including recDNA waste, is provided on the UNCG EHS website.
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Q32.
My project involves the use of synthetic DNA probes. Which questions from the Recombinant and Synthetic
DNA and RNA section (questions 31-43) apply to my work?
A.
If you project involves construction and/or use of synthetic DNA probes, mark 'yes' to question 32 and describe the applications for which these probes are used (e.g., oligonucleotides for PCR primers).
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Q35.
Which agents are exempt from the NIH Guidelines ?
A.
According to NIH Guidelines , Section III-F , the following agents are exempt with some exceptions , as stipulated in NIH Guidelines , Appendix C . Review by the IBC is not required for this work; however, you must still register work with all exempt agents with EH&S, by first submitting your completed BUA application.
Exempt agents include the following:

Those that are not in organisms or viruses (e.g., human cells transfected with plasmid DNA or
RNA, plasmid DNA or RNA that is not in viral vectors).

Those that consist entirely of DNA segments from a single nonchromosomal or viral DNA source, though one or more of the segments may be a synthetic equivalent.

Those that consist entirely of DNA from a prokaryotic host including its indigenous plasmids or viruses when propagated only in that host (or a closely related strain of the same species), or when transferred to another host by well-established physiological means.

Those that consist entirely of DNA from a eukaryotic host including its chloroplasts, mitochondria, and plasmids (but excluding viruses) when propagated only in that host (or a closely related strain of the same species).

Those that consist entirely of DNA segments from different species that exchange DNA by known physiological processes, though one or more of the segments may be a synthetic equivalent.
See NIH Guidelines , Appendix A for a list of natural exchangers that are exempt.

Those that do not present a significant risk to health or the environment as determined by the NIH
Director and following appropriate notice and opportunity for public comment.

Experiments involving Escherichia coli K12 host-vector systems. Some examples of E. coli K12 strains are DH10B, DH5-alpha, A19, and ECL1. The Uniprot website provides a reference list of E. coli 12 strains.

Experiments involving Saccharomyces cerevisiae and Saccharomyces uvarum host-vector systems.

Experiments involving Kluyveromyces lactis , host-vector systems.

Experiments involving Bacillus subtilis or Bacillus licheniformis host-vector systems

Transgenic rodents as described in the FAQs for Transgenic Animals .
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A. Yes, list all viral vector work in question 36. You must also list all other work with recombinant microorganisms that is not exempt from the NIH Guidelines (for information and a list of exempt agents,
see the FAQ for Q35.
In your answer, include the genus, species, and strains for the viral vectors and
other recombinant microorganisms.

If working with lentiviral vectors, clarify if there are non-HIV pseudotyped, replication deficient.

If working with gammaretroviral vectors, describe the vector pseudotype (e.g., ecotropic, amphotropic, VSV-G).

For uncommon pseudotype systems, also indicate the host range of that pseudotype [especially if able to infect human cells].

In addition to question 36, the Gene Delivery Methods table (question 44) must also be completed for work involving gene delivery via viral vectors and other recombinant microorganisms. You must use RefSeq gene names when listening specific genes.
Instructions for verifying correct RefSeq gene names are included in the FAQs for
Q44B .
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Q44B.
I am asked to provide gene inserts and key regulatory elements. Can I use gene names other than RefSeq gene names in column B of the Gene Delivery Methods table?
A.
No, gene names other than RefSeq gene names may not be used in this column. The IBC requires that you provide only RefSeq gene names. Some examples of RefSeq gene names include 'GFP', 'MYC', and
'COP4'. To verify that your gene names are correct follow the instructions below.
1. Access the National Center for Biotechnology Information (NCBI) RefSeqGene website .
2. Enter your gene name. Select 'Search' to search the RefSeq database.
3. The search results for your gene will display. If your gene name appears as blue linked text, you have entered a correct RefSeq gene name. If your gene name does not appear in the search results or appears under aliases, you have not entered a RefSeq gene name. You must determine the correct RefSeq gene name before entering the gene name into Column B or elsewhere on your application.
If your research involves more genes than can be easily listed in column B, list the functional categories or the categories of the encoded proteins in question 46.
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Q.
What are the exemptions for transgenic rodents?
A.
The following are exempt as stipulated in the NIH Guidelines , Section III-F .
1. The purchase or transfer of transgenic rodents.
2. Generation of BSL-1 transgenic rodents via breeding, with the following exceptions . Exceptions are as stipulated in NIH Guidelines , Appendix C . o Breeding of rodents that have a gene encoding more than fifty percent of an exogenous eukaryotic virus o Breeding of rodents in which the transgene is under the control of a gammaretroviral long terminal repeat (LTR)

See the reference table from NIH OBA for a listing of animal experiments covered under the NIH
Guidelines with references to the corresponding NIH Guidelines sections and biosafety levels.
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Q.
My work involves the generation of BSL-1 transgenic mice via breeding. Is this work exempt from the NIH
Guidelines ?
A.
Experiments involving the breeding of transgenic rodents who can be housed at BSL-1 containment are exempt from the NIH Guidelines only under the following conditions.

The transgenic rodents do not contain a gene encoding for more than one-half of a viral genome.

The transgene is not under the control of a gammaretroviral long term repeat (LTR).
The PI is responsible for assuring that the transgenes do not include gammaretroviral LTRs of more than one-half of a viral genome. There are a growing number of examples in which PIs have not reported this correctly. PIs are obligated to review the primary literature associated with the transgenic rodents that they wish to use in order to verify this. Special rules apply to the use of transgenic animals that fall under the above categories and failure to accurately disclose their use can put your funding and the funding of the
Institution at risk.
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Q57a.
I am asked to identify the biosafety levels of my laboratory. I am not sure about the biocontainment levels for my laboratories. Can you explain the different biocontainment levels?
A.
Question 57.a requires you to select the applicable biosafety levels for your laboratory. The four levels of physical containment, designated as BSL-1, BSL-2, BSL-3, and BSL-4, are as prescribed in NIH Guidelines ,
Appendix G and in the CDC Biosafety in Microbiological and Biomedical Laboratories , Section IV . Biosafety level classification of physical containment is based on laboratory practices, containment equipment, and special laboratory design. The following table on the UNCG IBC webpage provides the biosafety levels, their definitions, and some examples. Note that no BSL-3 or BSL-4 research is conducted at UNCG.
As mandated by the NIH Guidelines , PIs must complete the CITI Researcher Biosafety and CITI Blood
Borne Pathogens training modules prior to their initiation of work with biohazardous agents and every three years thereafter. The training provides relevant information about laboratory biosafety levels. It is recommended that you take the training before completing this application. Be aware that your application cannot be approved until completion of training is verified.
If this question is completed incorrectly, a biosafety officer will notify you and help you to determine the correct biosafety levels for your laboratory.
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Q57b.
I am asked to identify the biosafety levels of my animal facilities. I am not sure about the biocontainment levels for my animal work. Can you explain the different animal biocontainment levels.
A.
Animal Biosafety Level (ABSL) classification refers to facilities, practices, and operational requirements applicable to work with animals that have been exposed to biohazardous agents. Animals exposed to biohazardous agents assigned to BSLs 1-4 are generally assigned into ABSLs 1-4, respectively. ABSLs correspond to increased levels of protection for research staff and the environment, and are recommended as the minimal standard for activities involving laboratory animals exposed to biohazardous agents. Note that no ABSL-4 research is conducted at the UW. See the reference table from NIH OBA for a listing of animal experiments covered under the NIH Guidelines with references to the corresponding NIH
Guidelines sections and biosafety levels.
Practices and facility requirements for biosafety laboratories apply to research with animals as well. The animal care and use environment can present unique hazards not found in standard microbiological research laboratories (e.g., animals may generate aerosols, bite and scratch, be infected with a zoonotic agent, etc.) Keeping this in mind, all additional animal facility standard operating procedures must be followed.
If this question is completed incorrectly, a biosafety officer will notify you and help you to determine the correct animal biosafety levels for your laboratory.
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