NOTICES 
60129 
between the recovered and desired segment 
(e.g.. hybridization and restriction endonu- 
clease fragmentation analysis where appli- 
cable): <d) the genetic stability of the cloned 
fragment; and (e) any alterations in the bio- 
logical properties of the vector and host. 
4. In Section I-E. "exemptions" from the 
Guidelines are discussed. Such experiments 
are not covered by the Guidelines and need 
not be registered with NIH. In Section I-D 
on "prohibitions," the possibility of "excep- 
tions" is discussed. An "exception" means 
that an experiment may be expressly re- 
leased from a prohibition. At that time it 
will be assigned appropriate 
5. Care should be taken to Inactivate re- 
combinant DNA before disposal. Procedures 
for inactivating DNA can be found in the 
"Laboratory Safety Monograph: A Supple- 
ment to the NIH Guidelines for Recombin- 
ant DNA Research." 
6. Laboratory Safety at the Center for Dis- 
ease Control (Sept. 1974). U.S. Department 
of Health. Education and Welfare Publica- 
tion No. CDC 75-8118. 
7. Classification of Etiologic Agents on the 
Basis of Hazard. (4th Edition. July 1974). 
U.S. Department of Health. Education and 
Welfare. Public Health Service. Center for 
Disease Control. Office of Biosafety. Atlan- 
ta. Georgia 30333. 
8. National Cancer Institute Safety Stand- 
ards for Research Involving Oncogenic Vir- 
uses (Oct. 1974). U.S. Department of Health. 
Education and Welfare Publication No. 
(NIH) 75-790. 
9. National Institutes of Health Bioha- 
zards Safety Gutde (1974). U.S. Department 
of Health. Education and Welfare. Public 
Health Service. National Institutes of 
Health. U.S. Government Printing Office. 
Slock No. 1740-00383. 
10. Biohazards in Biological Research 
(1973). A. Heilman. M. N. Oxman. and R 
Pollack (ed.) Cold Spring Harbor Labora- 
tory. 
11. Handbook of Laboratory Safety (1971). 
Second Edition. N. V. Steere <ed.>. The 
Chemical Rubber Co.. Cleveland. 
12. Bodily. H. L. (1970). General Adminis- 
tration of the Laboratory. H. L. Bodily. E. L. 
Updyke, and J. O. Mason (eds.). Diagnostic 
Procedures for Bacterial. Mycotic and Para- 
sitic Infections. American Public Health As- 
sociation. New York. pp. 11-28. 
13. Darlow. H. M. (1969). Safety in the Mi- 
crobiological Laboratory. In J. R. Norris 
and D. W. Robbins (ed.). Methods In Micro- 
biology. Academic Press. Inc. New York. pp. 
169-204. 
14. The Prevention of Laboratory Acquired 
Infection (1974). C. H. Collins, E. O. Hart- 
ley. and R. Pllsworth. Public Health Labora- 
tory Service. Monograph Series No. 6. 
15. Chatlgny. M. A. (1961). Protection 
Against Infection In the Microbiological 
Laboratory: Devices and Procedures. In W. 
W. Umbreit (ed.). Advances In Applied Mi- 
crobiology. Academic Press. New York. N.Y. 
3:131-192. 
16. Design Criteria for Viral Oncology Re- 
search Facilities (1975). U.S. Department of 
Health. Education and Welfare. Public 
Health Service. National Institutes of 
Health. DHEW Publication No. (NIH) 75- 
891. 
17. Kuehne. R. W. (1973). Biological Con- 
tainment Facility for Studying Infectious 
Disease Appl. Microbiol. 26-239-243. 
18. Runkle. R. S.. and O. B. Phillips 
(1969). Microbial Containment Control 
FEDERAL 
Facilities. Van Nostrand Reinhold. New 
York. 
19. Chatlgny. M. A., and D. I. Cllnger 
(1969). Contamination Control in Aerobio- 
logy. In R. L. Dimmick and A. B. Akers 
(eds.). An Introduction to Experimental 
Aerobiology. John Wiley & Sons. New York, 
pp. 194-263. 
19A Horsfall. P. L.. Jr., and J. H. Baner 
(1940). Individual Isolation of Infected Ani- 
mals in a Single Room. J. Bact. 40. 569-580. 
20. Biological safety cabinets referred to 
in this section are classified as Class I. Class 
II. or Class III cabinets. A Class I is a venti- 
lated cabinet for personnel protection 
having an inward flow of air away from the 
operator. The exhaust air from this cabinet 
is filtered through a high-efficiency particu- 
late air (HEPA) filter. This cabinet is used 
In three operational modes: (1) With a full- 
width open front. (2) with an installed front 
closure panel (having four 8-inch diameter 
openings) without gloves, and (3) with an in- 
stalled front closure panel equipped with 
arm-length rubber gloves. The face velocity 
of the inward flow of air through the full- 
width open front is 75 feet per minute or 
greater. A Class II cabinet is a ventilated 
cabinet for personnel and product protec- 
tion having an open front with inward air 
flow for personnel protection, and HEPA fil- 
tered mass recirculated air flow for product 
protection. The cabinet exhaust air is fil- 
tered through a HEPA filter. The face ve- 
locity of the inward flow of air through the 
full-width open front is 75 feet per minute 
or greater. Design and performance specifi- 
cations for Class II cabinets have been 
adopted by the National Sanitation Founda- 
tion. Ann Arbor. Michigan. A Class III cabi- 
net is a closed-front ventilated cabinet of 
gas-tight construction which provides the 
highest level of personnel protection of all 
biohazard safety cabinets. The interior of 
the cabinet is protected from contaminants 
exterior to the cabinet. The cabinet Is fitted 
with arm-length rubber gloves and is operat- 
ed under a negative pressure of at least 0.5 
inches water gauge. All supply air Is filtered 
through HEPA filters. Exhaust air Is fil- 
tered through two HEPA filters or one 
HEPA filter and incinerator before being 
discharged to the outside environment. 
21. Hershfleld. V.. H. W. Boyer. C. Yan- 
ofsky, M. A. Lovett, and D. R. Helinski 
(1974). Plasmid Col El as a Mo’ecular Vehi- 
cle for Cloning and Amplification of DNA. 
Proc. Nat. Acad. Sci. USA 71 3455-3459. 
22. Wensink, P. C.. D. J. Finnegan. J. E. 
Donelson. and D. S. Kogness (1974). A 
System for Mapping DNA Sequences in the 
Chromosomes of Drosophila Melanogaster. 
Cell X 315-335. 
23. Tanaka. T.. and B. Welsblum (1975). 
Construction of a Colicin El-R Factor Com- 
posite Plasmid In Vitro. Means for Amplifi- 
cation of Deoxyribonucleic Acid. J. Barter- 
iol. 121. 354-362. 
24. Armstrong. K. K., V. Hershfleld. and 
D. R. Helinski (1977). Gene Cloning and 
Containment Properties of Plasmid Col El 
and Its Derivatives. Science 196. 172-174. 
25. Bolivar. F.. R. L. Rodriguez. M. C. Bet- 
lach. and H. W. Boyer (1977). Construction 
and Characterization of New Cloning Vehi- 
cles: I. Ampicillin- Resistant Derivative of 
pMB9. Gene 2, 75-93. 
26. Cohen. S. N.. A. C. W. Chang. H. 
Boyer, and R. Helling (1973). Construction 
of Biologically Functional Bacterial Plas- 
mids in Vitro. Proc. Natl. Acad. Scl. USA 70. 
3240-3244. 
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27. Bolivar. F.. R. L. Rodriguez, R. J. 
Greene. M. C. Batlach. H. L. Reyneker, H. 
W. Boyer. J. H. Crosa, and S. Falkow (1977). 
Construction and Characterization of New 
Cloning Vehicles: II. A Multi-Purpose Clon- 
ing System. Gene 2. 95-113. 
28. Thomas. M.. J. R. Cameron, and R. W. 
Davis (1974). Viable Molecular Hybrids of 
Bacteriophage Lambda and Eukaryotic 
DNA. Proc. Nat. Acad. Sci. USA 71 4579- 
43583. 
29. Murray. N. E.. and K. Murray (1974). 
Manipulation of Restriction Targets in 
Phage Lambda to Form Receptor Chromo- 
somes for DNA Fragments. Nature 251. 476- 
481. 
30. Rambach, A., and P. Tiollais (1974). 
Bacteriophage Hatring EcoRI Endonuclease 
Sites Only in the Non-Essential Region of 
the Genome Proc. Nat. Acad. Sci. USA 71, 
3927-3930. 
31. Blattner, F. R.. B. G. Williams, A. E. 
Bleche. K. Denniston-Thompson, H. E. 
Faber, L. A. Furlong. D. J. Gunwald. D. O. 
Kiefer. D. D. Moore. J. W. Shumm. E. L. 
Sheldon, and O. Smithies (1977). Charon 
Phages: Safer Derivatives of Bacteriophage 
Lambda for DNA Cloning. Science 196, 163- 
169. 
32. Donoghue. D. J.. and P. A. Sharp 
(1977). An Improved Lambda Vector: Con- 
struction of Model Recombinants Coding for 
Kanamycin Resistance. Gene 1. 209-227. 
33. Leder. P., D. Tiemeier and L. Enquist 
(1977). EK2 Derivatives of Bacteriophage 
Lambda Useful in the Cloning of DNA from 
Higher Organisms: Thi gt WES System. Sci- 
ence 196. 175-177. 
33A. Skalka, A. (1978). Current Status of 
Coliphage EK2 Vectors. Gene 3. 29-35. 
33B. Szybalski. W.. A. Skalka. S. Gottes- 
man, A. Campbell, and D. Botstein (1978). 
Standardized Laboratory Tests for EK2 Cer- 
tification. Gene 3. 36-38. 
34. We arc specifically concerned with the 
remote possibility that potent toxins could 
be produced by acquiring a single gene or 
cluster of genes. See also footnote 2A. 
35. Defined as observable under optimal 
laboratory conditions by transformation, 
transduction, phage infection, and/or conju- 
gation with transfer of phage, plasmid, and/ 
or chromosomal genetio information. Note 
that this definition of exchange may be less 
stringent than that applied to exempt or- 
ganisms under Section I-E-4. 
36. As classified in the Second Report of 
the International Committee on Taxonomy 
of Viruses: Classification and Nomenclature 
of Viruses. Frank Fenner. Ed. Intervirology 
7(19-115) 1976. (As noted In the Prohibition 
Section, the use of viruses classifiedil] as 
Class 3. 4, or 5. other than VSV. is prohibit- 
ed.) 
37. The cDNA copy of the viral mRNA 
must be >99% pure; otherwise as for shot- 
gun experiments with eukaryotic cellular 
DNA. 
37A. For the purpose of these Guidelines, 
viruses of the families Papovaviridae, Aden- 
oviridae. and Herpetoviridae (36) should be 
considered as "transforming" viruses. While 
only certain of these viruses have been asso- 
ciated with cell transformation in vivo or in 
vitro. It seems prudent to consider all mem- 
bers to be' potentially capable of transfor- 
mation. In addition, those viruses of the 
family Poxviridae that produce prolifera- 
tive responses— i.e.. myxoma, rabbit and 
squirrel fibroma, and Yaba viruses— should 
be considered as "transforming." 
22, 1978 
