Temperature-Gradient gel electrophoresis of nucleic acids: Analysis of conformational transitions, sequence variations, and protein-nucleic acid interactions

1989 ◽  
Vol 10 (5-6) ◽  
pp. 377-389 ◽  
Author(s):  
Detlev Riesner ◽  
Gerhard Steger ◽  
Rolf Zimmat ◽  
Robert A. Owens ◽  
Manfred Wagenhöfer ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Temperature Gradient ◽  
Nucleic Acids ◽  
Gel Electrophoresis ◽  
Conformational Transitions ◽  
Sequence Variations ◽  
Gradient Gel Electrophoresis ◽  
Protein Nucleic Acid ◽  
Temperature Gradient Gel Electrophoresis ◽  
Nucleic Acid Interactions

scholarly journals Detection of Mitochondrial DNA Mutations by Temporal Temperature Gradient Gel Electrophoresis

1999 ◽  
Vol 45 (8) ◽  
pp. 1162-1167 ◽  
Author(s):  
Tian-Jian Chen ◽  
Richard G Boles ◽  
Lee-Jun C Wong
Keyword(s):  
Mitochondrial Dna ◽  
Temperature Gradient ◽  
Gel Electrophoresis ◽  
Direct Sequencing ◽  
Polymorphism Analysis ◽  
Mtdna Mutations ◽  
Sequence Variations ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
Temperature Gradient Gel Electrophoresis

Abstract Background: A unique requirement for the molecular diagnosis of mitochondrial DNA (mtDNA) disorders is the ability to detect heteroplasmic mtDNA mutations and to distinguish them from homoplasmic sequence variations before further testing (e.g., sequencing) is performed. We evaluated the potential utility of temporal temperature gradient gel electrophoresis (TTGE) for these purposes in patients with suspected mtDNA mutations. Methods: DNA samples were selected from patients with known mtDNA mutations and patients suspected of mtDNA disorders without detectable mutations by routine analysis. Six regions of mtDNA were PCR amplified and analyzed by TTGE. Electrophoresis was carried out at 145 V with a constant temperature increment of 1.2 °C/h. Mutations were identified by direct sequencing of the PCR products and confirmed by PCR/allele-specific oligonucleotide or PCR/restriction fragment length polymorphism analysis. Results: In the experiments using patient samples containing various amounts of mutant mtDNA, TTGE detected as little as 4% mutant heteroplasmy and identified heteroplasmy in the presence of a homoplasmic polymorphism. In 109 specimens with 15 different known mutations, TTGE detected the presence of all mutations and distinguished heteroplasmic mutations from homoplasmic polymorphisms. When 11% of the mtDNA genome was analyzed by TTGE in 104 patients with clinically suspected mitochondrial disorders, 7 cases of heteroplasmy (≈7%) were detected. Conclusions: TTGE distinguishes heteroplasmic mutation from homoplasmic polymorphisms and appears to be a sensitive tool for detection of sequence variations and heteroplasmy in patients suspected of having mtDNA disorders.

Snapshot blotting: The transfer of nucleic acids and nucleoprotein complexes from electrophoresis gels to EM grids

1992 ◽  
Vol 50 (1) ◽  
pp. 762-763
Author(s):  
Stephen D. Jett
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acids ◽  
Nucleic Acid Binding ◽  
Mobility Shift ◽  
Carbon Coated ◽  
Nucleoprotein Complexes ◽  
Mobility Shift Assay ◽  
Protein Nucleic Acid ◽  
Gel Mobility Shift ◽  
Nucleic Acid Interactions

The electrophoresis gel mobility shift assay is a popular method for the study of protein-nucleic acid interactions. The binding of proteins to DNA is characterized by a reduction in the electrophoretic mobility of the nucleic acid. Binding affinity, stoichiometry, and kinetics can be obtained from such assays; however, it is often desirable to image the various species in the gel bands using TEM. Present methods for isolation of nucleoproteins from gel bands are inefficient and often destroy the native structure of the complexes. We have developed a technique, called “snapshot blotting,” by which nucleic acids and nucleoprotein complexes in electrophoresis gels can be electrophoretically transferred directly onto carbon-coated grids for TEM imaging.

scholarly journals Application of temperature gradient gel electrophoresis to the characterization of a nitrifying bioaugmentation product

2003 ◽  
Vol 43 (2) ◽  
pp. 277-286 ◽  
Author(s):  
Melissa A. Fouratt ◽  
Jeremy S. Rhodes ◽  
Charles M. Smithers ◽  
Nancy G. Love ◽  
Ann M. Stevens
Keyword(s):  
Temperature Gradient ◽  
Gel Electrophoresis ◽  
Gradient Gel Electrophoresis ◽  
Temperature Gradient Gel Electrophoresis

Temporal Temperature Gradient Gel Electrophoresis

2004 ◽  
pp. 1257-1261
Author(s):  
Lee‐Jun C. Wong ◽  
Özgül Alper ◽  
Hae‐Young Kwon ◽  
Duan‐Jun Tan ◽  
Tian‐Jian Chen
Keyword(s):  
Temperature Gradient ◽  
Gel Electrophoresis ◽  
Gradient Gel Electrophoresis ◽  
Temperature Gradient Gel Electrophoresis
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