{"data":{"id":"us/40-cfr-725.421","jurisdiction":"us","citation":"40 CFR 725.421","heading":"Introduced genetic material.","body":"For a new microorganism to qualify for either exemption under this subpart, introduced genetic material must meet all of the criteria listed in this section.\n(a) Limited in size. The introduced genetic material must consist only of the following:\n(1) The structural gene(s) of interest.\n(2) The regulatory sequences permitting the expression of solely the gene(s) of interest.\n(3) Associated nucleotide sequences needed to move genetic material, including linkers, homopolymers, adaptors, transposons, insertion sequences, and restriction enzyme sites.\n(4) The nucleotide sequences needed for vector transfer.\n(5) The nucleotide sequences needed for vector maintenance.\n(b) Well-characterized. For introduced genetic material, well-characterized means that the following have been determined:\n(1) The function of all of the products expressed from the structural gene(s).\n(2) The function of sequences that participate in the regulation of expression of the structural gene(s).\n(3) The presence or absence of associated nucleotide sequences and their associated functions, where associated nucleotide sequences are those sequences needed to move genetic material including linkers, homopolymers, adaptors, transposons, insertion sequences, and restriction enzyme sites.\n(c) Poorly mobilizable. The ability of the introduced genetic material to be transferred and mobilized is inactivated, with a resulting frequency of transfer of less than 10−8 transfer events per recipient.\n(d) Free of certain sequences. (1) The introduced genetic material must not contain a functional portion of any of the toxin-encoding sequences described in this paragraph (d).\n(i) For the purposes of this section, a functional portion of a toxin-encoding sequence means any sequence which codes for a polypeptide that has one of the following effects:\n(A) It directly or indirectly contributes to toxic effects in humans. Directly contributes to toxic effects in humans means those sequences encoding polypeptides that have direct toxicity to target cells. An example of a sequence which directly contributes to toxic effects in humans is one which encodes the portion of diphtheria toxin, listed in paragraph (d)(2) of this section, capable of interacting with elongation factor 2, leading to inhibition of protein synthesis in target respiratory, heart, kidney, and nerve tissues. Indirectly contributes to toxic effects in humans means a sequence whose encoded polypeptide is not directly toxic to target cells, yet still adversely affects humans. An example of a sequence which indirectly contributes to toxic effects is the sequence which encodes the portion of the botulinum toxin, listed in paragraph (d)(3) of this section, capable of blocking the release of acetylcholine from gangliosides. Botulinum toxin affects neuromuscular junctions by its blockage of acetylcholine release, leading to irreversible relaxation of muscles and respiratory arrest.\n(B) It binds a toxin or toxin precursor to target human cells.\n(C) It facilitates intracellular transport of a toxin in target human cells.\n(ii) While these toxins are listed (with synonyms in parentheses) in paragraphs (d)(2) through (d)(7) of this section according to the source organism, it is use of the nucleotide sequences that encode the toxins that is being restricted and not the use of the source organisms. The source organisms are listed to provide specificity in identification of sequences whose use is restricted. Although similar or identical sequences may be isolated from organisms other than those listed below in paragraphs (d)(2) through (d)(7) of this section, these comparable toxin sequences, regardless of the organism from which they are derived, must not be included in the introduced genetic material.\n(2) Sequences for protein synthesis inhibitor.\nSequence Source Toxin Name\nCorynebacterium diphtheriae \u0026 C. ulcerans Diphtheria toxin\nPseudomonas aeruginosa Exotoxin A\nShigella dysenteriae Shigella toxin (Shiga toxin, Shigella dysenteriae type I toxin, Vero cell toxin)\nAbrus precatorius, seeds Abrin\nRicinus communis, seeds Ricin\n(3) Sequences for neurotoxins.\nSequence Source Toxin Name\nClostridium botulinum Neurotoxins A, B, C1, D, E, F, G (Botulinum toxins, botulinal toxins)\nClostridium tetani Tetanus toxin (tetanospasmin)\nProteus mirabilis Neurotoxin\nStaphylococcus aureus Alpha toxin (alpha lysin)\nYersinia pestis Murine toxin\nSnake toxins\nBungarus caeruleus Caeruleotoxin\nBungarus multicinctus Beta-bungarotoxin (phospholipase)\nCrotalus spp. Crotoxin (phospholipase)\nDendroaspis viridis Neurotoxin\nNaja naja varieties Neurotoxin\nNotechia scutatus Notexin (phospholipase)\nOxyuranus scutellatus Taipoxin\nInvertebrate toxins\nChironex fleckeri Neurotoxin\nAndroctnus australis Neurotoxin\nCentruroides sculpturatus Neurotoxin\n(4) Sequences for oxygen labile cytolysins.\nSequence Source Toxin Name\nBacillus alve Alveolysin\nBacillus cereus Cereolysin\nBacillus laterosporus Laterosporolysin\nBacillus thuringiensis Thuringiolysin\nClostridium bifermentans Lysin\nClostridium botulinum Lysin\nClostridium caproicum Lysin\nClostridium chauvoei Delta-toxin\nClostridium histolyticum Epsilon-toxin\nClostridium novyi Gamma-toxin\nClostridium oedematiens Delta-toxin\nClostridium perfringens Theta-toxin (Perfringolysin)\nClostridium septicum Delta-toxin\nClostridium sordellii Lysin\nClostridium tetani Tetanolysin\nListeria monocytogenes Listeriolysin (A B)\nStreptococcus pneumoniae Pneumolysin\nStreptococcus pyogene Streptolysin O (SLO)\n(5) Sequences for toxins affecting membrane function.\nSequence Source Toxin Name\nBacillus anthracis Edema factor (Factors I II); Lethal factor (Factors II III)\nBacillus cereus Enterotoxin (diarrheagenic toxin, mouse lethal factor)\nBordetella pertussis Adenylate cyclase (Heat-labile factor); Pertussigen (pertussis toxin, islet activating factor, histamine sensitizing factor, lymphocytosis promoting factor)\nClostridium botulinum C2 toxin\nClostridium difficile Enterotoxin (toxin A)\nClostridium perfringens Beta-toxin; Delta-toxin\nEscherichia coli \u0026 other Enterobacteriaceae spp. Heat-labile enterotoxins (LT); Heat-stable enterotoxins (STa, ST1 subtypes ST1a ST1b; also STb, STII)\nLegionella pneumophila Cytolysin\nVibrio cholerae \u0026 Vibrio mimicus Cholera toxin (choleragen)\n(6) Sequences that affect membrane integrity.\nSequence Source Toxin Name\nClostridium bifermentans \u0026 other Clostridium spp Lecithinase\nClostridium perfringens Alpha-toxin (phospholipase C, lecithinase); Enterotoxin\nCorynebacterium pyogenes \u0026 other Corynebacterium spp. Cytolysin (phospholipase C), Ovis toxin (sphingomyelinase D)\nStaphylococcus aureus Beta-lysin (beta toxin)\n(7) Sequences that are general cytotoxins.\nSequence Source Toxin Name\nAdenia digitata Modeccin\nAeromonas hydrophila Aerolysin (beta-lysin, cytotoxic lysin)\nClostridium difficile Cytotoxin (toxin B)\nClostridium perfringens Beta-toxin; Epsilon-toxin; Kappa-toxin\nEscherichia coli \u0026 other Enterobacteriaceae spp. Cytotoxin (Shiga-like toxin, Vero cell toxin)\nPseudomonas aeruginosa Proteases\nStaphylococcus aureus Gamma lysin (Gamma toxin); Enterotoxins (SEA, SEB, SEC, SED SEE); Pyrogenic exotoxins A B; Toxic shock syndrome toxins (TSST-1)\nStaphylococcus aureus \u0026 Pseudomonas aeruginosa Leucocidin (leukocidin, cytotoxin)\nStreptococcus pyogenes Streptolysin S (SLS); Erythrogenic toxins (scarlet fever toxins, pyrogenic exotoxins)\nYersinia enterocolitica Heat-stable enterotoxins (ST)","path":["Title 40—Protection of Environment","CHAPTER I—ENVIRONMENTAL PROTECTION AGENCY","SUBCHAPTER R—TOXIC SUBSTANCES CONTROL ACT","PART 725—REPORTING REQUIREMENTS AND REVIEW PROCESSES FOR MICROORGANISMS","Subpart G—General Exemptions for New Microorganisms"],"source_url":"https://www.ecfr.gov/api/versioner/v1/full/2026-08-25/title-40.xml","current_through":"2026-08-25","vintage":"","retrieved_at":"2026-08-27T02:26:04Z","sha256":"e21d7b9de2b5078792ea29be9f6e289067821afa5e94943d653c5b486dac4e03","source_id":"us-cfr","stale":true,"prev":"us/40-cfr-725.420","next":"us/40-cfr-725.422"},"notice":"GroundRules: Original legal text. Not legal advice."}