the University of Alabama in Birmingham / UNIVERSITY STATION / BIRMINGHAM, ALABAMA 35294 
the Medical Center /department of microbiology / May 11, 1979 
Dr. William Gartland 
Director 
Office of Recombinant DNA Activities 
Room 4A52, Building 31 
National Institutes of Health 
Bethesda, Maryland 20014 
Dear Bill: 
I am writing in response to the notice of proposed actions under the NIH 
Guidelines for Research Involving Recombinant DNA Molecules as appeared in 
the Federal Register on April 13, 1979. My comments are as follows: 
Item 5 - Proposed Amendment of Section II-D-l-a-(l) . I am not at all supportive 
of the proposed amendment to the definition of EK1 that would permit the presence 
of autonomous or integrated conjugative plasmids or prophages of generalized trans- 
ducing phages in hosts used for phage vectors. The purpose of this amendment is 
ostensibly to allow use of M13 and other donor-specific DNA-containing phages as 
cloning vectors. However, the change also creates a "loophole", an example of 
which is discussed below. 
In terms of the M13 cloning vector system, there are several features that very 
much decrease its biological containment relative to the EK1 systems employing 
X vectors. First, M13-infected cells do not lyse and continue to multiply while 
extruding M13 particles containing the cloned insert. Second, the production of 
M13 particles by such infected cells is several orders of magnitude greater than 
the production of X by cells lysogenic for X which are permitted under the EK1 
category. This is based on the fact that an M13-infected cell releases at 
least 10-times more phage per unit time than a x-infected cell and the fact that 
all M13-infected cells release phage compared to 1 per 10^ to 10 5 cells lysogenic 
for X. Third, while x-sensitive hosts that are infectable by X are exceedingly 
rare in the natural environment, M13 and other donor-specific DNA phages have a 
much more extensive host range. The most prevalent conjugative plasmids present 
in E_. coli strains in the United States are those in the IncFI, IncFII, IncFIII, 
IncFIV and IncFV groups. All these plasmids specify F pi 1 i and thus endow sen- 
sitivity to M13. Furthermore, although two years ago I and others estimated that 
the presence of derepressed F-type conjugative plasmids were rare among natural 
enteric isolates, recent data indicate that Ent plasmids (which specify entero- 
toxin) and ColV plasmids (which specify some virulence factor) are increasingly 
[260] 
AN AFFIRMATIVE ACTION / EQUAL OPPORTUNITY EMPLOYER 
