ENVIRONMENTAL HEALTH & SAFETY

Recombinant or Synthetic Nucleic Acid Molecules

Institutional Biosafety Committee (IBC) approval is required prior to the initiation of most NON-exempt Recombinant or Synthetic Nucleic Acid Molecule experiments.

For information and registration procedures for Recombinant or Synthetic Nucleic Acid Molecule experiments, contact Paul Skoglund at ps5d@virginia.edu or 243-0726.

Additional information available at:

• NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules
• NIH/OSP FAQs about experiments that are exempt from the NIH Guidelines

Guidance on the use of viral vectors can be found at:

• Biosafety Considerations for Research with Lentiviral Vectors

Guidance on animal experiments involving Recombinant or Synthetic Nucleic Acid Molecules can be found at:

• Animal experiments covered under the NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules.

A brief description of NON-Exempt and Exempt recombinant or synthetic nucleic acid molecule experiments are provided below.

NON-Exempt Experiments - Recombinant or Synthetic Nucleic Acid Molecules

1. Gene transfer experiments in humans.
• Section III-C-1. Requires IBC; RAC and Institutional Review Board approval before initiation.
• Example: Use of a defective adenoviral vector to deliver the CFTR gene intranasally into patients with Cystic Fibrosis; Introduction of a HSV-TK transduced cellline into patients with epithelial ovarian carcinoma, followed by therapy with Gancyclovir.
2. Cloning of toxin molecules with LD50 of less than 100 nanograms/kg body weight.
• Section III-B-1. E.g. microbial toxins such as the botulinum toxins; tetanus toxin; diphtheria toxin; and Shigella dysenteriae neurotoxin. Requires NIH/OSP and IBC approval before initiation.
• Example: Cloning toxin genes (or using plasmids that express genes that encode toxins with low LD50’s) such as Botulinum, Tetrodotoxin, Ricin, T-2, Saxitoxin, Abrin, Tetanus,Shigella Dysenteriae, Pertussis, Staph Aureus Beta, ShigaToxin, and Conotoxins.
3. Experiments involving the introduction of Recombinant or Synthetic Nucleic Acid Molecules into Risk Group 2 organisms (includes adenovirus and murine retroviral vectors) or higher.
• Section III-D-1. Requires IBC approval before initiation.
• Example: Using Adenovirus, Adenovirus-luciferase or adeno-associated virus to transfect cells; Typically involves use of pathogens or defective pathogen vectors (with or without helper virus), such as Adenovirus, Adeno-Associated virus, Baculovirus, Herpes virus, Lentivirus, Retrovirus, Vaccinia and Vesicular Stomatitis Virus, Shigella, Salmonella, Yersinia, and E. histolytica.
4. Experiments in which DNA from human or animal pathogens (Risk Group 2 and higher) is introduced into a non-pathogenic host-vector system.
• Section III-D-2. Requires IBC approval before initiation.
• Example: Yersinia pseudotuberculosis genes encoding outer membrane adhesins are cloned into plasmid vectors for re-introduction into mutant strains of the same bacteria or E. coli.
5. Experiments Involving Cultures of More than 10 Liters.
• Section III-D-6. Requires IBC approval before initiation.
• Example: Use of a 10-Liter fermentor or growing up five 2-Liter flasks of Recombinant or Synthetic Nucleic Acid Molecule culture (i.e. E. coli K-12) qualifies as a large scale experiment.
6. Experiments involving the use of infectious DNA or RNA viruses or defective DNA or RNA viruses (Risk Group 2 or higher) in the presence of helper virus in tissue culture systems.
• Section III-D-3. Requires IBC approval before initiation.
• Example: Insertion of KSHV or RRV genes into defective lentiviral vectors.
7. Experiments Involving the Formation of Recombinant or Synthetic Nucleic Acid Molecules containing no more than 2/3 of the genome of any eukaryotic virus (with some restrictions). It must be demonstrated that cells lack helper virus.
• Section III-E-1. Requires IBC notification simultaneous with initiation.
• Example: Inserting DNA sequences that encode reporters that are measured (lacZ, luciferase, eGFP, dsRed2, etc), or that encode enzymes that are potentially therapeutic (nitric oxide synthases, superoxide dismutase, siRNA) against mRNAs that promote disease, etc into viral vectors that retain no more than 2/3 of the original viral genomic sequence. The cDNAs will be driven by the following promoters: CMV IE, RSV LTR, and cardiac Troponin T (cTnT).
8. Experiments involving whole plants.
• Section III-E-2 or D-5. Investigators must refer to the NIH guidelines to determine if experiments are exempt. Requires IBC notification simultaneous with initiation.
• Example: Creation of transgenic plants; Inserting a gene for herbicide tolerance in food or ornamental plants.
9. The deliberate transfer of a drug resistance trait to microorganisms that are not known to acquire the trait naturally; if such acquisition could compromise the use of the drug to control disease agents in humans; veterinary medicine; or agriculture.
• Section III-A-1-a. Requires RAC review.
• Example: Transferring a drug resistance trait that is used, had previously been used, may be used (outside the U.S.), or that is related to other drugs that are used to treat or control disease agents. These include: Transfer of Erythromycin resistance into Borrelia burgdorferi; Transfer of Pyrimethamine resistance into Toxoplasma gondii; Transfer of Chloramphenicol resistance into Rickettsia conorii; Transfer of Tetracycline resistance into Porphyromonas gingivalis.
10. Whole animals in which the animal's genome has been altered by the stable introduction of Recombinant or Synthetic Nucleic Acid Molecules, or RNA derived therefrom, into the germ-line (transgenic animals).
• Section III-D-4. Requires IACUC/IBC approval.
• Example: Creation of transgenic animals (mice, rats, zebra fish, drosophila,etc.), or knockout animals that leave genetic material in the animal as part of the silencing of the gene. Note: the purchase (or transfer to your lab) of previously created transgenic rodents is exempt from the regulations.

Exempt Experiments - Recombinant or Synthetic Nucleic Acid Molecules

1. Are not in organisms or viruses.
2. Use as host vector systems E. coli K 12; Saccharomyces cerevisiae; Saccharomyces uvarum; or Bacillus subtilis; and their plasmids.
3. Use of Recombinant or Synthetic Nucleic Acid Molecules molecules containing less than one-half of any eukaryotic genome that are propagated and maintained in cells in tissue culture.
4. 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.
5. 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.
6. Consist entirely of DNA from a eukaryotic host including its chloroplasts; mitochondria; or plasmids (but excluding viruses) when propagated only in that host (or a closely related strain of the same species).