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Internalization of oligonucleotides ...

Don, Michelle.

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Internalization of oligonucleotides into bacterial cells.

Record Type:	Electronic resources : Monograph/item
Title/Author:	Internalization of oligonucleotides into bacterial cells./
Author:	Don, Michelle.
Published:	Ann Arbor : ProQuest Dissertations & Theses, : 2009,
Description:	66 p.
Notes:	Source: Masters Abstracts International, Volume: 71-10.
Contained By:	Masters Abstracts International71-10.
Subject:	Molecular biology. -
Online resource:	https://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=1470805
ISBN:	9781109339482

Internalization of oligonucleotides into bacterial cells.
Don, Michelle.

Internalization of oligonucleotides into bacterial cells. - Ann Arbor : ProQuest Dissertations & Theses, 2009 - 66 p.

Source: Masters Abstracts International, Volume: 71-10.

Thesis (M.S.)--California State University, Fullerton, 2009.

The enzyme aminoglycoside 6'-N-acetyl transferase (AAC(6')-Ib) encoded by the transposon Tn1331 on the pJHCMW1 plasmid, mediates resistance to amikacin and other aminoglycoside antibiotics. Through the use of RNase H mapping and computer prediction software, the secondary structure of aac(6')-Ib mRNA was determined and regions accessible for interaction with antisense oligonucleotides were identified. Oligonucleotides designed to target these regions have been used to inhibit the synthesis of AAC(6')-Ib, decreasing the level of resistance to amikacin. However, a successful and reproducible method of delivery of oligonucleotides to bacterial cells is not yet available. Liposome-mediated delivery of fluorescent labeled phosphorothioate oligonucleotide analogues and plasmid DNA to bacterial cells was attempted in this study. Fluorescent microscopy and antibiotic resistance plates were used as evidence to show internalization of the oligonucleotides or plasmid DNA. A method of liposome encapsulation of DNA was established, however, liposome-mediated delivery of DNA to Escherichia coli and Bacillus subtilis cells was unsuccessful. Delivery of naked phosphorothioate oligonucleotide analogues to Acinetobacter baumanii was successful and internalization was visualized through fluorescence microscopy. This work provides a methodology to test internalization of oligonucleotides into bacterial cells, and contributes to the overall understanding of liposomes as vehicles of DNA delivery and provides the first evidence that Acinetobacter baumanii, as it is the case of A. calcoaceticus and A. baylyi, is naturally competent.

ISBN: 9781109339482Subjects--Topical Terms:

517296
Molecular biology.

Internalization of oligonucleotides into bacterial cells.
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500 $a Publisher info.: Dissertation/Thesis.
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520 $a The enzyme aminoglycoside 6'-N-acetyl transferase (AAC(6')-Ib) encoded by the transposon Tn1331 on the pJHCMW1 plasmid, mediates resistance to amikacin and other aminoglycoside antibiotics. Through the use of RNase H mapping and computer prediction software, the secondary structure of aac(6')-Ib mRNA was determined and regions accessible for interaction with antisense oligonucleotides were identified. Oligonucleotides designed to target these regions have been used to inhibit the synthesis of AAC(6')-Ib, decreasing the level of resistance to amikacin. However, a successful and reproducible method of delivery of oligonucleotides to bacterial cells is not yet available. Liposome-mediated delivery of fluorescent labeled phosphorothioate oligonucleotide analogues and plasmid DNA to bacterial cells was attempted in this study. Fluorescent microscopy and antibiotic resistance plates were used as evidence to show internalization of the oligonucleotides or plasmid DNA. A method of liposome encapsulation of DNA was established, however, liposome-mediated delivery of DNA to Escherichia coli and Bacillus subtilis cells was unsuccessful. Delivery of naked phosphorothioate oligonucleotide analogues to Acinetobacter baumanii was successful and internalization was visualized through fluorescence microscopy. This work provides a methodology to test internalization of oligonucleotides into bacterial cells, and contributes to the overall understanding of liposomes as vehicles of DNA delivery and provides the first evidence that Acinetobacter baumanii, as it is the case of A. calcoaceticus and A. baylyi, is naturally competent.
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