Bacteriophage Systems 
Vector 
Host 
Date Certified 
Literature 
References 
Xg[WES AB' 
DPEOsupF 
2-22-1977 
8,9 
XgiWES ■ AB* 
DPSOsupf 
6-22-1977 
12 
AgtZJw'r ■ AB' 
£. coli K-12 
2-22-1977 
10 
Charon 3A \ 
( DP50 
Charon 4A > 
or 
3-1-1977 
11 
Charon 16A ; 
1 DPSOsupf 
Charon 21A 
DPSOsupf 
8-12-1978 
13 
In Volume 1, Number 1 of the Bulletin, Charon 16A was identified as Charon 316A. The designation Charon 16A is currently in use. 
in EK2 systems, recombinant molecules may be introduced into propagation hosts either by transfection with naked DNA or by 
infection with viral particles produced by in vitro packaging. When in vitro packaging is used: 
(1) The packing extract must be free from viable bacteria. 
(2) Any packaging protocol may be used, provided that control experiments on the packaging of EK2 vector DNA meet one of the 
following two criteria; 
(a) The number of amber+ phages produced must be less than 10-^ times the number of amber- phages. If shotgun populations 
are to be propagated in bulk culture or by confluent lysis methods, this measurement must be made on packaged EK2 vector DNA 
propagated to the same extent. 
(b) If the total number of amber- phages produced in a packaging experiment is less than 10®, and if the shotgun population is not 
to be propagated in bulk, the number of observed amber plaques must be zero. 
(3) The above tests must be done on each batch of packaging extract used. 
(4) Any individual clone isolated from the shotgun must be tested for retention of the safety characteristics of the vector before it can 
be used for bulk propagation. 
(5) A description of the packaging protocol should be filed with NIH for information, but NIH approval is not required provided that the 
above numerical criteria are met. 
Certain types of small-scale experiments are allowed in hosts other than DPSOsupf. Specifically, transfection and very small scale 
experiments (i.e., less than 1.0 ml, plating tor single plaques for screening purposes) are allowed in other E. co//K-12 hosts. However, 
plate lysates or other procedures (for instance, growth of small lysates in liquid) which would yield large numbers of phage (1or 
greater) are to be done only in the DPSOsupf host. 
Literature References 
1. Curtiss, R., Ill, Pereirei, D.A., Hsu J.C., Hull, S.C.. Clark, J.E., 
Maturin, L.F., Sr/ Goldschmidt, R.. Moody, R., Inoue, M., and 
Alexander, L., Biological Containment: The Subordination of 
Escherichia Coli K-12, in Recombinant Molecules; Impact on 
Science and Society Edt. R.R. Beers, Jr,, and E.G. Bassett. Pro¬ 
ceedings of the 10th Miles International Symposium, 45-46, 
1977, 
2. Cohen, S.N,, Chang, A.C.Y., Boyer, H., and Helling, R., Con¬ 
struction of biologically functional plasmids in vitro. Proc. Nat. 
Acad, Sci, USA 70: 3240-3244, 1973. 
4. Armstrong, K.A., Hershfield, V., and Helinski, D R,, Gene 
cloning and containment properties of plasmid Col El and its 
derivatives. Science 196: 172-4,1977, 
5. Rodriguez, R.L., Bolivar, F., Goodman, H.M., Boyer, H,, and 
Betlach, M., Construction and characterization of cloning vehi¬ 
cles, in Molecular Mechanisms in the Control of Gene Expres¬ 
sion Edt D.P. Nierlich, W.J. Rutter and C.F. Fox, Proceedings of 
the 5th ICN-UCLA Symposium, 471-77, 1976, 
6. Bolivar, F., Rodriguez, R.I., Betlach, M,C., and Boyer, H.W., 
Construction and characterization of new cloning vehicles: I. 
Ampicillin-resistant derivative of the plasmid pMB9, Gene 2: 
75-93, 1977, 
3, Covey, C., Richardson, D , and Carbon, J,, A Method for the 
detection of restriction sites in bacterial plasmid deoxyribonu¬ 
cleic acid, Molec, Gen. Genet. 145: 155-58, 1976, 
26 
