Multiple host expression vector

Details Multiple host expression vector Multiple host expression vector

2000-04-13

2003-07-08

Guzo, David (Department: 1636)

Chemistry: molecular biology and microbiology

Vector, per se

C536S023100, C536S024100

Reexamination Certificate

active

06589783

ABSTRACT:

CROSS-REFERENCE TO RELATED APPLICATIONS
STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT
BACKGROUND OF THE INVENTION
In the practice of modem biotechnology it has become common practice to create artificial gene constructions which are capable of expressing a foreign protein in a host which normally does not produce that protein. The process of producing a protein from a gene in a host is referred to as protein expression.
Interest in expressing proteins in foreign hosts has led to the development of a market in pieces of DNA known as expression vectors. Expression vectors, often but not invariably sold in the form of circular pieces of DNA known as plasmids, contain all of the genetic elements on them to express a protein coding sequence inserted into a defined location within the expression vector. Typically one would create the coding sequence for a protein that one wants to express and then insert that coding sequence into a defined location in the expression vector. The expression vector would already contain within it a promoter adapted for use in the host for which the expression vector is defined and may also include suitable downstream transcription terminators or polyadenylation sequences sufficient to terminate transcription, a necessary requirement in a eukaryotic, but not a prokaryotic host. Such expression vectors often would also have a selectable marker gene for one or more forms of antibiotic resistance so that the bacterial host containing the expression vector can readily be identified.
Different categories of host organisms require different promoters. For example, prokaryotic promoters such as those functional to cause gene expression in bacteria, are almost invariably non-functional in eukaryotic organisms such as vertebrate or plant cells or yeast. Even among eukaryotic organisms, most plant promoters do not work in animal cells and most animal promoters do not work in plant cells. Accordingly it is typically the case that an expression vector is designed for a specific host or class of hosts and has promoters and other genetic elements specifically designed for that specific host.
One prior vector is known that can be used in two hosts. The pDual vector from Stratagene includes promoters suitable for both mammalian cell expression and bacterial expression. No prior vector is known capable of operation in more than two categories of hosts.
BRIEF SUMMARY OF THE INVENTION
The present invention is summarized in that a multiple host expression vector has built within it a facility for expression of a inserted protein coding sequence in bacterial cells, vertebrate cells, and in baculovirus infected insect cells. This permits the same expression vector to be used in bacterial, insect, or vertebrate-cells to produce a protein of interest.
The vector of the present invention advantageously has diverse promoters in the orientation of vertebrate, then bacterial, then baculovirus, since then the bacterial and baculovirus promoters can be incorporated into 5′ untranslated mRNA that does not affect efficient expression of an inserted gene sequence in vertebrate-cells.
The present invention is also summarized in that the p10 baculovirus promoter is used in such a multiple host expression vector, the p10 baculovirus promoter permitting high expression of inserted protein coding sequences in baculovirus infected insect cells without adversely effecting the functioning of other promoters contained within the vector.
Other objects, advantages and features of the present invention will become apparent from the following specification when taken in conjunction with the accompanying drawings.


REFERENCES:
patent: 6194158 (2001-02-01), Kroes et al.
pDual Expression System, Stratagene on-line catalog, http://www.stratagene.com/vectors/expression/pdual.htm; Mar. 23, 2001.*
Novy et al. FASEB Journal 13(7):A1552, abstract# 1269, Apr. 1999.*
Novy et al. Mol. Biol. Cell 10(Suppl):270a, abstract#1562, Nov. 1999.*
Hasnain et al. Gene 190:113-118 1997.*
Binder et al. Plant Mol. Biol. 32:303-314 1996.*
Romanos et al. Yeast 8:423-488 1992.*
pBAC-2cp Transfer Plasmid, Novagen Catalog p. TB128 1/96.
pcDNA3.1/Hygro Vectors, Invitrogen abstract.
pCI Vector Circle Map, Sequence and Restriction Sites, Abstract 9/94.
pDual Expression System, Statagen Catalog (Aug. 9, 1999).
pET-28a-c(+) Vectors, Novagen Catalog p. TB074 12/98.

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