Male Specific Bacteriophages and EK1 Containment 
The Ml3, fd, and other related single-strand 
bacteriophages in conjunction with £. coli K-12 
hosts have been examined for their suitability 
as EK1 systems. Since these bacteriophages 
are male-specific, they are not presently 
approvable for use with their natural hosts as 
the Guidelines do not allow EK1 hosts that con¬ 
tain conjugation-proficient plasmids. 
However, these bacteriophages in conjunc¬ 
tion with E. coii K-12 strains that do not contain 
conjugation-proficient plasmids (i.e., E') are 
acceptable for experiments that require EK1 
biological containment. Since the host strains 
are not the natural hosts, the means of infection 
would involve transfection. Procedures for 
transfection are similar to those for transforma¬ 
tion of E. coll. Because the phages are exuded 
into the medium without cell lysis, large yields 
can be obtained. 
In addition based on the recommendations of 
the NIH Recombinant DMA Advisory Commit¬ 
tee (RAC) and its bacteriophage subcommit¬ 
tee, these bacteriophages are now also 
approved for use with bacteria that contain 
conjugation-defective plasmids: 
Conjugation-deficient mutants, such as the 
traD and tral mutants of the E factor may be 
used with Ef bacteriophages if the mutants 
have been shown to exhibit low levels of 
transfer (of the order of 10-^ or less) and also 
have low reversion rates (such as found for 
deletion or double-mutants). 
This action is based on the intent of the Guide¬ 
lines; that is, the hosts shall not contain 
conjugation-proficient plasmids. These restric¬ 
tions on the transfer defective mutants combined 
with the natural containment features of these 
bacteriophages in their hosts provide more than 
adequate EK1 containment. 
The use of these or any bacteriophages in the 
presense of conjugationally-proficient plasmids 
has not been approved for EK1 containment. This 
requires further examination by the RAC and will 
be discussed at the next meeting of the RAC as 
part of a general proposal to allow such use. 
[A complete report of the use of the single-strand 
male-specific bacteriophages as vectors in EK1 
systems is available from the Office of Recombi¬ 
nant DNA Activities.] 
dA-dT Tailing with Certified Vectors 
The dA-dT tailing method can be used to enable use of this method is considered to lead only to a 
the joining of DNA fragments of EK2 vectors. The minor modification of a certified vector. 
X2282 in EK2 Experiments 
The E. coli K-12 strain X2282 has been given 
approval for limited use as an EK2 host. This 
ThyA+ variant of XI 776 may only be used for EK2 
containment in experiments involving the cloning 
and expression of the dihydrofolate reductase 
gene.x2282 is not certified for general use in EK2 
systems. 
Data to be Submitted for the Certification of 
EK2 Host-Plasmid Systems 
The Host-plasmid Subcommittee of the RAC 
recently reevaluated the criteria for acceptance of 
EK2 systems. Many of the following instructions to 
investigators concerning data to be submitted, 
were described in an earlier Bulletin (Vol. 1. No. 2. 
Winter, 1978, p. 5 see II, B), The major changes 
include shorter matings (see B), testing an addi¬ 
tional plasmid (see II, C), and the absence of tri- 
parental matings. 
I. Survival 
(A) Time-course experiments must show that 
survival of the host-plasmid falls to 10-^ or lower 
within 24 hours of incubation in vitro under non- 
permissive conditions (i.e., conditions that have 
been demonstrated to obtain in the mammalian 
intestinal tract). 
(B) The above loss of survival must depend on 
at least two phenotypic traits which contribute 
17 
