Complexation of Lipofectamine and Cholesterol-Modified DNA Sequences Studied by Single-Molecule Fluorescence Techniques

2007 ◽  
Vol 8 (11) ◽  
pp. 3382-3392 ◽  
Author(s):  
Anca Margineanu ◽  
Steven De Feyter ◽  
Sergey Melnikov ◽  
Damien Marchand ◽  
Arthur van Aerschot ◽  
...  
Keyword(s):  
Single Molecule ◽  
Dna Sequences ◽  
Single Molecule Fluorescence ◽  
Fluorescence Techniques ◽  
Modified Dna

scholarly journals Liz Rhoades: Single molecules tell many stories

2010 ◽  
Vol 191 (6) ◽  
pp. 1046-1047
Author(s):  
Caitlin Sedwick
Keyword(s):  
Protein Folding ◽  
Single Molecule ◽  
Single Molecules ◽  
Single Molecule Fluorescence ◽  
Fluorescence Techniques

Rhoades studies protein folding and binding using single-molecule fluorescence techniques.

scholarly journals Multi-site internal modification of long DNA substrates for single-molecule studies

2016 ◽  
Author(s):  
Armando de la Torre ◽  
Yoori Kim ◽  
Andrew A. Leal ◽  
Ilya J. Finkelstein
Keyword(s):  
Single Molecule ◽  
Dna Sequences ◽  
Nuclear Antigen ◽  
Triplet Repeat ◽  
Chemical Modifications ◽  
Site Specific ◽  
Specific Modification ◽  
Modified Dna ◽  
Single Molecule Studies ◽  
Dna Substrates

AbstractSingle-molecule studies of protein-nucleic acid interactions frequently require site-specific modification of long DNA substrates. DNA isolated from bacteriophage λ (λ-DNA) is a convenient source of high quality long (48.5 kb) DNA. However, introducing specific DNA sequences, tertiary structures, and chemical modifications into λ-DNA remains technically challenging. Most current approaches rely on multi-step ligations with low yields and incomplete products. Here, we describe a molecular toolkit for rapid preparation of modified λ-DNA. A set of PCR cassettes facilitates the introduction of recombinant DNA sequences into λ-DNA with 90-100% yield. Furthermore, various DNA structures and chemical modifications can be inserted at user-defined sites via an improved nicking enzyme-based strategy. As a proof-of-principle, we explore the interactions of Proliferating Cell Nuclear Antigen (PCNA) with modified DNA sequences and structures incorporated within λ-DNA. Our results demonstrate that PCNA can load on both 5’-ssDNA flaps and a 13xCAG triplet repeat. However, PCNA remains trapped on the 13xCAG structure, confirming a proposed mechanism for triplet repeat expansion. Although we focus on λ-DNA, this method is applicable to all long DNA substrates. We anticipate that this molecular toolbox will be broadly useful for both ensemble‐ and single-molecule studies that require site-specific modification of long DNA substrates.

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