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Title: Methods and compositions for amplifying DNA clone copy number

Abstract

A method for retrofitting DNA in a single-copy or high-copy vector, such as a fosmid or BAC, whereby an artificial transposon is used to introduce a conditional multi-copy origin of replication (“ori”) into the DNA in said vector. Following random in vitro or in vivo transposition of the ori-containing transposon into DNA in the single-copy or low-copy vector, the resulting insertion clones are introduced into a special host strain that contains a gene which encodes a polypeptide required for replication from the multi-copy ori. However, since the gene for this polypeptide is expressed from a tightly-regulated inducible promoter, the polypeptide is not expressed in the absence of inducer. On addition of inducer to the culture medium, the host cell synthesizes the polypeptide, which in turn activates replication from the multi-copy ori, thereby increasing the amount of clone DNA synthesized by the cell.

Inventors:
; ; ;
Issue Date:
Research Org.:
Epicentre Technologies Corp., Madison, WI (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1531519
Patent Number(s):
7138267
Application Number:
10/116,886
Assignee:
Epicentre Technologies Corporation (Madison, WI)
Patent Classifications (CPCs):
C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES
DOE Contract Number:  
FG02-00ER83003
Resource Type:
Patent
Resource Relation:
Patent File Date: 2002-04-04
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Jendrisak, Jerome J., Hoffman, Leslie M., Fiandt, Michael J., and Haskins, Darin. Methods and compositions for amplifying DNA clone copy number. United States: N. p., 2006. Web.
Jendrisak, Jerome J., Hoffman, Leslie M., Fiandt, Michael J., & Haskins, Darin. Methods and compositions for amplifying DNA clone copy number. United States.
Jendrisak, Jerome J., Hoffman, Leslie M., Fiandt, Michael J., and Haskins, Darin. Tue . "Methods and compositions for amplifying DNA clone copy number". United States. https://www.osti.gov/servlets/purl/1531519.
@article{osti_1531519,
title = {Methods and compositions for amplifying DNA clone copy number},
author = {Jendrisak, Jerome J. and Hoffman, Leslie M. and Fiandt, Michael J. and Haskins, Darin},
abstractNote = {A method for retrofitting DNA in a single-copy or high-copy vector, such as a fosmid or BAC, whereby an artificial transposon is used to introduce a conditional multi-copy origin of replication (“ori”) into the DNA in said vector. Following random in vitro or in vivo transposition of the ori-containing transposon into DNA in the single-copy or low-copy vector, the resulting insertion clones are introduced into a special host strain that contains a gene which encodes a polypeptide required for replication from the multi-copy ori. However, since the gene for this polypeptide is expressed from a tightly-regulated inducible promoter, the polypeptide is not expressed in the absence of inducer. On addition of inducer to the culture medium, the host cell synthesizes the polypeptide, which in turn activates replication from the multi-copy ori, thereby increasing the amount of clone DNA synthesized by the cell.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {2006},
month = {11}
}

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Works referenced in this record:

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