What is the gt in lambda gt10 or gt11?
smithwhi at students.msu.edu
Thu Mar 17 15:13:00 EST 1994
In Article <2m2aoq$2q0 at news.acns.nwu.edu> "h-seifert at nwu.edu (Hank Seifert)" says:
> >> Right you are. I was just talking off the top of my head and
> >> as usual was wrong. However in the spirit of discussion,
> >> should terms from genetics be used to describe new techniques
> >> that arise from molecular biology? The process of transducing
> >> a new piece of DNA is not unique to the lambda-gt-series of
> >> bacteriophage but also applies to M13 phage and all other phage
> >> derived cloning vectors. None of these vectors have a 'gt'
> >> designation.
> >The tradition of including 'gt' as part of the name of a particular phage
> >strain was a part of the history of lambda phage genetics whereas this
> >tradition was absent in M13. This is simply because only lambda was ever
> >able to demonstrate generalized transduction prior to the age of in vitro
> >manipulation. If you use M13 to clone - in contrast to subcloning - you
> >deserve all the problems you encounter.
> I'm confused, when was lambda showed to participate in
> generalized transduction?
The definition of transduction is the transfer of genetic information
from one bacterium to another via a phage. There is nothing in the
definition concerning the construction of the transducing phage. I
incorrectly stated that lambda performed generalized transduction
prior to the era of in vitro manipulation.
> >The process of cloning into a phage
> >> vector is very distinct from generalized transduction
> >Is it? Just because you perform all but one of the steps in a cell-free
> >environment does not justify overlooking the identity with the in vivo
> >process. Otherwise one would have to create a new name for the process of
> >encapsidating the lambda genomes in vitro to distinguish that from the name
> >originally developed for the process in vivo - packaging the phage genome.
> These are not the same things. The process of packaging lambda in
> vitro is mechanistically similar to that in vivo. The
> mechanisms used to allow "natural" transduction by P1, P22,
> or Mu phage are very different than cloning into gt10 or gt11.
This obsession with "natural"ness is the whole problem. Is the
hydration of CO2 on the protein surface of carbonic anhydrase -
carbonate dehydratase more or less natural than the process occurring
in protein-free solution? Should the there be two names for the
> >> result in the new generations of researchers,
> >> many of whom never heard of generalized transduction, thinking that
> >> the term is specific for cloning. The 'gt' label was good
> >> advertising for Young and Davis, but is it good terminology?
The vocabulary is explicitly designated in the original literature -
PNAS 71,4579 (1974). Since you claim to have read the original
literature I will provide the quote for those of us who obviously
don't have either the time or the inclination to read and forget.
"METHODS AND MATERIALS
TERMINOLOGY. The term lambda gt (generalized transducer) designates
that portion of the lambda genome that is common to all the hybrid
DNA molecules and contains all of the essential genes for plaque
formation." (So much for the "garbage truck" theory put forth in an
> >The vocabulary is not good with respect to gt10 since the presence of an
> >insert renders the phage incapable of forming a lysogen in the absence of cI
> >protein supplied in trans. The ability to form a lysogen is an obligatory
> >intermediate in generalized transduction by lambda. However Young and Davis
> >followed a tradition initiated by others in the development of cloning
> >vectors which have been supeceded by those currently in vogue with their
> >trademarked names. (Don't get me started about this silliness.)
> >> I'm also concerned about the use
> >> of other genetic terms in this context. Genetic
> >> transformation has a defined meaning in bacterial genetics
> >> (it was the first proof that DNA is the genetic material) but
> >> now means the introduction of plasmids into E. coli by
> >> chemical or physical means.
In the definition of transformation there is nothing concerning the
method or route or protocol to get the DNA into the cell. Check the
glossary of any genetics textbook. Back to your obsession with
"natural"ness - whatever the hell that is.
> >If you had read the original literature, the definition of transformation
> >has always included the concept of introduction of DNA into the receptive
> >recipient. There never was a proscription concerning the source or the nature
> >of the DNA. If you had read some more original literature, you would have
> >noticed that the process of Ca-dependent introduction of DNA into E coli was
> >originally studied with lambda as a tracer - and was described as
> >transfection. When the generality of the process was demonstrated, the name
> >was changed to transformation - since the result was a heritable alteration
> >of the recipient bacteria.
> I have read most, if not all, of the original papers. I agree
> that if a change in genotype and phenotype occurs that
> transformation is a proper term. I think, however, that we
> should be able to create new terminology to describe the very
> different processes of artificial transformation and natural
Back to the "natural"ness silliness. If a population of bacteria have
0.0001% of the cells competent to take up DNA, is it a violation
of the vocabulary/thought police to think that a protocol which
yields a population of bacteria with 99% of the cells competent to
take up DNA is not natural? Or is the mere fact that human
intervention occurred sufficient to taint the process and thus
requires a new vocabulary to name the process?
> >To make matters worse, the same
> >> process in eukaryotic cells is called transfection! This term was
> >> originally reserved for the introduction of viral DNA into a
> >> cell.
> >If you had read the original literature, the process of introducing DNA into
> >a eukaryotic cell was studied with viral DNA as a tracer. Therefore the
> >vocabulary is exactly correct. The fact that the process fails to
> >discriminate against nonviral DNA is no reason to rename the technique. Also,
> >if the technique is used to create cells with a heritable change, most
> >individuals withwhom I have had contact use the vocabulary of transformation.
> >NO PROBLEM!!!!
> I just experienced a minor heat shock! Most people in the
> world use the term transfection to describe the introduction of
> DNA into eukayotic cells (just read this newsgroup!). I read a
A good question - is the name determined by the process of getting
the DNA into the cell or the result of the process of getting the DNA
into the cell? People I have worked with have used the latter
criterion. Transient expression is transfection - heritable change is
transformation. No problem!
> suggestions several years ago that all introduction of DNA
> into cells by artificial means be termed transfection. I would
> prefer to reserve this for viral DNA but still think that
> artificial introduction of DNA and natural uptake of DNA should
> have seperate terms. This is also true for the new
> observations that muscle cells can efficiently take up DNA and express
> genes encoded therein.
> > While it is too late to changes these terms, their use
> >> leads to inaccurate communication and can produce sloppy
> >> thinking. These new processes should have new names to
> >> distinguish them from their related genetic systems? Are there
> >> good alternate names for artificial plasmid transformation,
> >> eukaryotic cell transfection, and cloning into phage vectors?
> >Maybe if you spent a little of your "net time" with any introductory genetics
> >text while considering the subtle points of identity between superficially
> >different events, we would be relieved of this burden to rename the wheel.
> Perhaps if you would consider whether our present terminology
> is appropriate instead of launching a personal attack or
> relying on historical precedent,
As if any sort of nomenclature or vocabulary you would consider
acceptable would have no relation to any historical precedents. If
you think that my attack upon your obsession with "natural"ness is
personal - that's too bad.
we could have a
> reasonable discussion on the merits of these terms.
> Hank Seifert
More information about the Methods