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Biotech / Medical : Targeted Gene Repair
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To: John McCarthy who started this subject9/15/2002 8:25:17 PM
From: John McCarthy   of 22
 
TARGETED GENE THERAPY

Subject:SFHR - (small fragment homologous replacement )
(The following material is presented in Ascending Date Order)

This post is generated by a program that reads a database and generates this file. Errors will result from incorrect database material. Updates to the database automatically reflected in this file.

Researcher Name shown is my estimate of Principal Researcher

DB Key:2001-SFHR-B
Doc:Abstract
Vector:SFHR Small Fragment DNA
Experiment:Failed or Not Applicable
Of Note:Epithelial cells

2/1/2001-----Goncz KK-----University of Vermont
Title:Targeted correction of a defective selectable marker gene in human epithelial cells by small DNA fragments
Link:http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11237674&dopt=Abstract

Snippet:
The present study evaluates the conditions that modulate SFHR-mediated correction of a defective Zeocin antibiotic resistance (Zeo(r)) gene that has been inactivated by a 4-bp insertion. The conditions include delivery systems, plasmid-to-fragment ratio, fragment length, and fragment strandedness (single- or double-stranded DNA). Targeting fragments comprise the wild-type Zeo(r) gene sequence and were either 410 (Zeo1) or 458 bp (Zeo3). Expression vectors containing the corrected Zeo(r) gene were isolated as episomal plasmids or were allowed to stably integrate into cultured human airway epithelial cells. Correction of the Zeo(r) gene was phenotypically defined as restoration of resistance to Zeocin in either bacteria or epithelial cell clones. Extrachromosomal gene correction was assayed using polymerase chain reaction amplification, restriction enzyme digestion, DNA sequencing, and Southern blot hybridization analysis of DNA from isolated prokaryotic and eukaryotic clones. Neither random sequence alteration in the target episomal gene nor random integration of the small fragments was detected. Targeted correction efficiencies of up to 4% were attained. These studies provide insight into parameters that can be modulated for the optimization of SFHR-mediated targeting.
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