Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting.

Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Research Abstract Details 

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Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Abstract Text:

B R Murphy,H S Moayedpardazi,A M Gewirtz,S L Diamond,E A Pierce,

Single-stranded oligodeoxynucleotide (ssODN) gene targeting may facilitate animal model creation and gene repair therapy. Lipofection of ssODN can introduce point mutations into target genes. However, typical efficiencies in mouse embryonic stem cells (ESC) are <10(-4), leaving corrections too rare to effectively identify. We developed ESC lines with an integrated mutant neomycin resistance gene (Tyr22Ter). After targeting with ssODN, repaired cells survive selection in G418. Correction efficiencies varied with different lipofection procedures, clonal lines, and ssODN designs, ranging from 1 to 100 corrections per million cells plated. Uptake studies using cell sorting of Cy5-labelled ssODN showed 40% of the corrections concentrated in the best transfected 22% of cells. Four different basepair mismatches were tested and results show that the base-specificity of the mismatch is critical. Dual mismatch ssODN also showed mismatch preferences. These ESC lines may facilitate development of improved ssODN targeting technologies for either animal production or ex vivo gene therapy.

Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Publishing Authors By Initials

BR Murphy,HS Moayedpardazi,AM Gewirtz,SL Diamond,EA Pierce,

For similar investigative techniques: genetic techniques: gene transfer techniques: transfection research abstracts see: investigative techniques: genetic techniques: gene transfer techniques: transfection research

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Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Gene therapy

VOLUME: 14

Page Numbers: 304-15

Journal Abbreviation: Gene Ther.

ISSN: 0969-7128

DAY: 5

MONTH: 10

YEAR: 2006

Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 9421525

Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Keywords Mesh Terms:

KEYWORDS: Transfection

MESH TERMS: methods

Chemical & Substance for Abstract: Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting. Information

Substance Name: Neomycin

Registry Number: 1404-04-2

Grant and Affiliation Information for Delivery and mechanistic considerations for the production of knock-in mice by single-stranded oligonucleotide gene targeting.

AFFILIATION: Department of Chemical and Biomolecular Engineering, Institute for Medicine and Engineering, University of Pennsylvania, Philadelphia, PA, USA.

Country: England

AGENCY: United States NCI

GRANT: P01-CA72765

ACRONYM: CA

MEDLINETA: Gene Ther

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